REFERENCE TO SEQUENCE LISTING SUBMITTED ELECTRONICALLY

[001] The disclosure is being filed along with a Sequence Listing in ST.26 XML format. The Sequence Listing is provided as a file titled “30436 WO” created 13 September 2023 and is 74.6 kilobytes (kb) in size. The Sequence Listing information in the ST.26 XML format is incorporated herein by reference in its entirety.

TECHNICAL FIELD

[002] The disclosure relates generally to biology and medicine, and more particularly it relates to synthetic nucleic acids and their use for treating diseases and disorders associated with triggering receptor expressed on myeloid cells 2 (TREM2), especially as a gene therapy for treating neurological disease such as Alzheimer’s Disease (AD), Adult-Onset Leukoencephalopathy with Axonal Spheroids and Pigmented Glia (ALSP) or Nasu-Hakola Disease (NHD).

BACKGROUND

[003] TREM2 is a cell surface transmembrane glycoprotein expressed primarily in cells of myeloid lineage, including microglia. In humans, the complete absence of TREM2 has been shown to cause NHD, a rare neurodegenerative disease with late-onset dementia, demyelination and cerebral atrophy. See, e.g., Paloneva et al. (2002) Am. J. Hum. Genet. 71 :656-662 and Paloneva et al. (2003) J. Exp. Med. 198:669-675.

[004] Mutations in TREM2 also have been associated with risk of developing AD. See, e.g. , Guerreiro et al. (2013) N. Engl. J. Med. 368: 117-127. AD is an irreversible, progressive brain disorder characterized by the presence of abnormal protein deposits throughout the brain, which inhibit neuronal function, disrupt connections between neurons, and ultimately result in cell death. These deposits comprise plaques of amyloid-beta and tangles formed by phosphorylated-tau proteins. Individuals with mild AD experience memory loss, leading to wandering, difficulty handling money, repeating questions, and personality and behavior changes. Moreover, individuals with moderate AD exhibit increased memory loss, leading to confusion and difficulty recognizing friends and family, inability to learn new things, hallucinations, delusions, and paranoia. Furthermore, individuals with severe AD cannot communicate and are completely dependent on others for their care. Ultimately, protein plaques and tangles spread throughout the brain, leading to significant tissue shrinkage.

[005] Mutations in CSF1R can lead to a microgliopathy known as Colony Stimulating Factor 1 Receptor (CSFIR)-Related Adult-Onset Leukoencephalopathy (CRL), which is a subclass of ALSP. Interestingly, TREM2 activates a similar signaling pathway resulting in intracellular and functional responses that phenocopy CSF1R activation. It is believed that increased cellular TREM2 can attenuate CSF1R loss-of-function and can attenuate CRL- associated pathology. See, e.g., Tchessalova et al. (2022) Alzheimer's Dement. 18:e061595.

[006] A number of therapeutic approaches to increase TREM2 are known, including small molecules or monoclonal antibodies to modulate the downstream signaling of TREM2. More recently, even gene therapy approaches have been described to increase TREM2 and/or its activity. See, e.g., Inti. Patent Application Publication No. WO 2019/070894.

[007] There is a need, however, for other disease-modifying therapies to increase TREM2 and/or its activity in individuals having TREM2-associated diseases and/or disorders.

BRIEF SUMMARY

[008] To address this need, the disclosure describes synthetic nucleic acids for use in treating TREM2-associated diseases and disorders. In one instance, the synthetic nucleic acid includes a nucleotide sequence that encodes TREM2 (i.e., a TREM2-encoding transgene) and that has at least about 90% (e.g., at least about 90%, 91%, 92, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100%) sequence identity to any one of SEQ ID NOS:3 to 6. In some instances, TREM2-encoding transgene can be codon-optimized. Alternatively or additionally, the TREM2-encoding transgene can be CpG minimized or CpG depleted. In certain instances, the nucleotide sequence for the TREM2-encoding transgene is SEQ ID NO:3, 4, 5 or 6.

[009] In another instance, the synthetic nucleic acid can be an expression construct including a first nucleotide sequence for at least one expression control element operably linked to a second nucleotide sequence that encodes TREM2 (i.e., a TREM2-encoding transgene), where the second nucleotide sequence has at least about 90% (e.g., at least about 90%, 91%, 92, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100%) sequence identity to any one of SEQ ID NOS:3 to 6. As above, the TREM2-encoding transgene can be codon-optimized. Alternatively or additionally, the TREM2-encoding transgene can be CpG minimized or CpG depleted. In certain instances, the nucleotide sequence for the TREM2-encoding transgene is SEQ ID NO:3, 4, 5 or 6.

[0010] In some instances, the at least one expression control element can be a promoter, an enhancer, a post-transcriptional regulatory element or a polyadenylation signal. In other instances, the promoter can be a chicken P-actin (CBA) promoter or a CD68 promoter or F4/80 promoter and can include a nucleotide sequence having at least about 90% (e.g., at least about 90%, 91%, 92, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100%) sequence identity to any one of SEQ ID NOS:7 to 9, respectively. In certain instances, the nucleotide sequence for the promoter is SEQ ID NO: 7, 8 or 9. In certain other instances, the nucleotide sequence for the promoter is SEQ ID NO:8.

[0011] In some instances, the enhancer can be a cytomegalovirus enhancer (CMVe) and can include a nucleotide sequence having at least about 90% (e.g., at least about 90%, 91%, 92, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100%) sequence identity to SEQ ID NO: 10. In certain instances, the nucleotide sequence for the enhancer is SEQ ID NO: 10.

[0012] In some instances, the post-transcriptional regulatory element can be a woodchuck hepatitis virus post-transcriptional regulatory element (WPRE) and can include a nucleotide sequence having at least about 90% (e.g., at least about 90% 91%, 92, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100%) sequence identity to SEQ ID NO: 11. In certain instances, the nucleotide sequence for the post-transcriptional regulatory element is SEQ ID NO: 11.

[0013] In some instances, the polyadenylation signal can be a bovine growth hormone poly A signal tail (BGHpA) and can include a nucleotide sequence having at least about 90% (e.g., at least about 90%, 91%, 92, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100%) sequence identity to SEQ ID NO: 12. In certain instances, the nucleotide sequence for the polyadenylation signal is SEQ ID NO: 12.

[0014] In some instances, the expression construct can include an additional nucleotide sequence that encodes a second transgene or that encodes an inhibitory nucleic acid.

[0015] In another instance, the synthetic nucleic acid can be a vector that includes an expression construct herein. In some instances, the vector can be a plasmid or viral vector. In other instances, the viral vector can be an adeno-associated virus (AAV) vector or a Baculovirus vector. When the vector is an AAV vector, it can include at least one AAV inverted terminal repeat (ITR) sequence flanking the expression construct. In some instances, the AAV ITR can be a wild-type (WT) AAV2 ITR and can have a nucleotide sequence having at least about 90% (e.g., at least about 90%, 91%, 92, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100%) sequence identity to SEQ ID NO: 13 or a reverse complementary sequence thereto. In other instances, the AAV ITR can be a modified AAV2 ITR and can have a nucleotide sequence having at least about 90% (e.g., at least about 90%, 91%, 92, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100%) sequence identity to SEQ ID NO: 14 or a reverse complementary sequence thereto. In certain instances, the nucleotide sequence for the AAV ITR is SEQ ID NO: 13 or 14 or a reverse complementary sequence thereto.

[0016] In some instances, the vector also can include a TRY region and can have a nucleotide sequence having at least about 90% (e.g., at least about 90%, 91%, 92, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100%) sequence identity to SEQ ID NO: 15. In certain instances, the nucleotide sequence for the TRY region is SEQ ID NO: 15.

[0017] In some instances, the vector also can include a nucleotide sequence that assists in packaging the vector with capsid protein (i.e., a sequence that optimizes the size of the vector for packaging within capsid protein, also called a stuffer sequence). In some instances, the stuffer sequence has at least about 90% (e.g, at least about 90%, 91%, 92, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100%) sequence identity to any one of SEQ ID NOS: 16 to 18. In certain instances, the nucleotide sequence is SEQ ID NO: 16, 17 or 18.

[0018] The disclosure also describes a rAAV that includes (i) a rAAV vector including a nucleotide sequence having at least about 90% (e.g, at least about 90%, 91%, 92, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100%) sequence identity to any one of SEQ ID NOS:3 to 6 (i.e., TREM2-encoding transgene) and (2) an AAV6 capsid protein.

[0019] In some instances, the rAAV includes:

(a) a rAAV vector having, in 5' to 3' order, a nucleotide sequence including:

(i) a first AAV ITR or a reverse complementary sequence thereto,

(ii) a promoter,

(iii) a TREM2-encoding transgene,

(iv) a post-transcriptional regulatory element,

(v) a polyadenylation signal, and

(vii) a second AAV ITR or a reverse complementary sequence thereto; and

(b) encapsidated in an AAV6 capsid protein.

[0020] In some instances, the AAV6 capsid protein is a modified AAV6 capsid protein (e.g., an AAV6TM capsid protein) including an amino acid sequence having at least about 95% (e.g., at least about 95%, 96%, 97%, 98%, 99% or 100%) sequence identity to SEQ ID NO: 19. In certain instances, the AAV6TM capsid protein amino acid sequence is SEQ ID NO: 19.

[0021] In some instances, the rAAV vector includes a nucleotide sequence having at least about 90% (e.g., at least about 90%, 91%, 92, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100%) sequence identity to any one of SEQ ID NOS:21 to 26. In certain instances, the nucleotide sequence of the rAAV vector is any one of SEQ ID NOS:21, 22, 23, 24, 25 or 26.

[0022] In some instances, the rAAV vector further includes a nucleotide sequence that assists in packaging the vector with capsid protein (z.e., a stuffer sequence) having at least about 90% (e.g., at least about 90%, 91%, 92, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100%) sequence identity to any one of SEQ ID NOS: 16 to 18. In certain instances, the nucleotide sequence for the stuffer sequence is SEQ ID NO: 16, 17 or 18.

[0023] In some instances, the rAAV vector further includes a second nucleic acid having a nucleotide sequence having reverse complementary of any one of SEQ ID NOS:21, 22, 23, 24, 25 or 26.

[0024] This disclosure also describes a pharmaceutical composition that includes a synthetic nucleic acid, expression construct, vector or rAAV herein and a pharmaceutically acceptable carrier.

[0025] In some instances, the pharmaceutical composition can be a formulation including a rAAV herein and one or more of:

(a) about 20 mM TRIS (pH 8.0),

(b) about 1 mM MgCh,

(c) about 200 mM NaCl, and

(d) about 0.005% (w/v) Poloxamer 188.

[0026] In some instances, the rAAV can be present in the formulation at a concentration from about 1 x 10 ¹³ vector genomes (vg) to about 7 x 10 ¹⁴ vg. In other instances, the rAAV can be present in the formulation at a concentration of about 3.5 x 10 ¹³ vg, about 7.0 x 10 ¹³ vg or about 1.4 x 10 ¹⁴ vg.

[0027] This disclosure also describes a method of treating diseases and disorders associated with TREM2 in an individual. The method can include at least a step of administering to the individual an effective amount of a synthetic nucleic acid, expression construct, vector or rAAV herein. [0028] In some instances, the disease or disorder associated with TREM2 can be AD, ALSP or NHD.

[0029] In some instances, the method also can include a step of measuring TREM2, CSF1R, neurofilament light chain (NfL), chitotriosidase (Chitl) and/or granulocyte-macrophage colony stimulating factor (GM-CSF) in plasma and/or cerebrospinal fluid (CSF) and/or urine from the individual and comparing an obtained value to an earlier-obtained comparable value or to a control value to assess effectiveness of the method.

[0030] In some instances, the method also can include a step of administering to the individual an effective amount of at least one additional therapeutic agent.

[0031] The disclosure further describes a composition including the rAAV for use as a medicament for treatment of diseases and disorders associated with TREM2, such as AD, ALSP or NHD.

[0032] The disclosure further describes a composition including the rAAV for use in the treatment of diseases and disorders associated with TREM2, such as AD, ALSP or NHD.

[0033] The disclosure further describes uses of the rAAV in manufacturing a medicament for treating diseases and disorders associated with TREM2, such as AD, ALSP or NHD, where the medicament optionally may include an additional therapeutic agent.

[0034] An advantage of the synthetic nucleic acids described herein is that they can increase and/or improve microglia function (z.e., development, maintenance and/or activation) in AD, ALSP or NHD by increasing both membrane-bound and secreted TREM2 levels and/or by augmenting or replacing CSF1R signaling with TREM2 signaling to activate downstream CSF 1R effects.

[0035] An advantage of the rAAV described herein is that it shows broader central nervous system (CNS) biodistribution and increased microglial transduction as compared to a rAAV having AAV9 capsid protein.

BRIEF DESCRIPTION OF THE DRAWINGS

[0036] The advantages, effects, features, and objects other than those set forth above will become more readily apparent when consideration is given to the detailed description below. Such detailed description refers to the following drawing(s), where:

[0037] FIG. 1 is a schematic depicting a first exemplary rAAV vector for expressing TREM2. [0038] FIG. 2 is a schematic depicting a second exemplary rAAV vector for expressing TREM2.

[0039] FIG. 3 is a schematic depicting a third exemplary rAAV vector for expressing TREM2.

[0040] FIG. 4 is a schematic depicting a fourth exemplary rAAV vector for expressing TREM2.

[0041] FIGS. 5A-5B show TREM2 mRNA (FIG. 5 A) and protein expression (FIG. 5B) in a HMC3 cell line when transduced with an exemplary rAAV vector encapsidated in AAV6TM capsid protein as compared to the same rAAV vector encapsidated in AAV9 capsid protein (n=4/group) (AAV6TM = filled circles; AAV9 = open circles).

[0042] FIGS. 6A-6B show TREM2 expression in a HMC3 cell line when transduced with one of two exemplary rAAV vectors, both encapsidated in AAV6TM capsid protein (FIG. 6 A = first rAAV vector; FIG. 6B = second rAAV vector).

[0043] FIGS. 7A-D show TREM2 biodistribution in somatosensory cortex (FIG. 7A), hippocampus (FIG. 7B), cervical spinal cord (FIG. 7C) and liver (FIG. 7D) of mice administered a low or high dose of an exemplary rAAV vector encapsidated in AAV6TM capsid protein or the same rAAV vector encapsidated in AAV9 capsid protein (n=8-12/group). [0044] FIGS. 8A-C show TREM2 protein levels in CSF (FIG. 8A), liver (FIG. 8B) and serum (FIG. 8C) of mice administered a low or high dose of an exemplary rAAV vector encapsidated in AAV6TM capsid protein or the same rAAV vector encapsidated in AAV9 capsid protein (n=8-12/group; a value of 20.48 pg/mL was set as the detection threshold (dotted line). For all graphs: statistics determined using ANOVA followed by Dunnett’s test comparing to the 5xF AD + excipient group. (*^<0.1; *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001).

[0045] FIGS. 9A-9C show amyloid-P levels in the hippocampus (FIGS. 9A-9B) and X34 ⁺ plaques in the cortex (FIG. 9C) of mice administered a low or high dose of an exemplary rAAV vector encapsidated in AAV6TM capsid protein or the same rAAV vector encapsidated in AAV9 capsid protein (n=8-12/group; statistics determined using ANOVA followed by Dunnett’s test comparing to the 5xFAD + excipient group. ^(:|:) p<0.1; *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001).

[0046] FIGS. 10A-10B show inflammation (FIG. 10A for Iba and FIG. 10B for IL-10) in the cortex of mice administered a low or high dose of an exemplary rAAV vector encapsidated in AAV6TM capsid protein or the same rAAV vector encapsidated in AAV9 capsid protein (n=8- 12/group; statistics determined using ANOVA followed by Dunnett’s test comparing to the 5xF AD + excipient group. (*^<0.1; *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001).

[0047] FIG. 11 shows TREM2 biodistribution in non-human primates (NHPs) administered a low or high dose of an exemplary rAAV vector encapsidated in AAV6TM capsid protein (n=2/group; vector copies were measured using ddPCR).

[0048] FIG. 12A-12C show TREM2 protein levels in liver, spinal cord and dorsal root ganglion (DRG) (FIG. 12A), CSF (FIG. 12B) and serum (FIG. 12C) of NHPs administered a low or high dose of an exemplary rAAV vector encapsidated in AAV6TM capsid protein (n=2/group; a value of 20.48 pg/mL was set as the detection threshold (dotted line)).

DETAILED DESCRIPTION

[0049] Overview

[0050] TREM2 is a cell surface transmembrane glycoprotein expressed primarily in cells of myeloid lineage, including microglia. TREM2 directly binds to amyloid-beta, facilitating microglial congregation around plaques, limiting plaque accumulation and promoting phagocytosis. TREM2 mutations associated with increased risk of AD decrease microglial response to amyloid plaques.

[0051] Additionally, CRL is a microgliopathy that can be caused by mutations in the kinase domain of CSF1R and represents the most common genetic forms of ALSP. Interestingly, CSF1R and TREM2 share a common, convergent signaling pathway to maintain and activate microglia. As such, TREM2 expression may rescue or compensate for CSF 1R loss-of-function. [0052] There are two TREM2 isoforms, where isoform 1 is 230 amino acids (aa) in length (SEQ ID NO: 1; see also, NCBI Ref. Seq. No. NP 061838.1) and where isoform 2 is 219 aa in length (SEQ ID NO:2; see also, NCBI Ref. Seq. No. NP_001258750.1). Exemplary nucleic acid sequences for can be found in NCBI Ref. Seq. Nos. NM_018965.4 and NM_001271821.2 (human), NM_031254.3 and NM_001272078.1 (mouse), XP_006244486.1 and XP_006244487.1 (rat), and XP_001117305.2 and XP_001174118.2 (non-human primate). One of skill in the art, however, understands that additional examples of TREM2 mRNA sequences are readily available using publicly available databases such as, for example, GenBank and UniProt. [0053] The disclosure describes synthetic nucleic acids for use as a gene therapy in treating TREM2-associated diseases and disorders, where the gene therapy delivers a functional copy of TREAD, which encodes TREM2, to an individual in need thereof.

[0054] Abbreviations and Definitions

[0055] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of skill in the art to which the disclosure pertains. Although any methods and materials similar to or equivalent to those described herein can be used in the practice or testing of the methods herein, the preferred methods and materials are described herein.

[0056] Additionally, reference to an element by the indefinite article “a” or “an” does not exclude the possibility that more than one element is present, unless the context clearly requires that there be one and only one element. The indefinite article “a” or “an” thus usually means “at least one.”

[0057] Moreover, use of “including,” as well as other forms, such as “including but not limited, “include,” “includes” and “included,” is not limiting.

[0058] Certain abbreviations used herein are as follows:

[0059] “aa” refers to amino acid(s); “AAV” refers to adeno-associated virus, “AcNPV” refers to Autographa californica nuclear polyhedrosis; “AD” refers to Alzheimer’s Disease; “ALS” refers to amyotrophic lateral sclerosis; “ALSP” refers to adult-onset leukoencephalopathy with axonal spheroids and pigmented glia; “BAC” refers to bacterial artificial chromosome; “BEYS” refers to baculovirus vector expression system; “BBB” refers to blood-brain-barrier; “BGHpA” refers to bovine growth hormone polyA signal tail; “bp” refers to base pair(s); “CBA” refers to chicken-P actin; “Chit 1” refers to chitotriosidase; “CNS” refers to central nervous system; “CSF” refers to cerebrospinal fluid; “CMVe” refers to cytomegalovirus enhancer; “CpG” refers to cytosine-phosphate-guanine; “CRL” refers to Colony Stimulating Factor 1 Receptor-Related Adult-Onset Leukoencephalopathy; “CSF1R” refers to colony stimulating factor 1 receptor gene; “CSF1R” refers to colony stimulating factor 1 receptor protein “sCSFIR” refers to soluble colony stimulating factor 1 receptor protein; “DAM” refers to disease associated microglia; “ddPCR” refers to droplet digital polymerase chain reaction; “DEA” refers to diethylamine; “DNA” refers to deoxyribonucleic acid; “DRG” refers to dorsal root ganglion; “ds” refers to double-stranded; “EDTA” refers to ethylenediaminetetraacetic acid; “EGTA” refers to ethylene glycol tetraacetic acid; “ELISA” refers to enzyme-linked immunosorbent assay; “FA” refers to formic acid; “FAD” refers to familiar Alzheimer Disease; “FTD” means frontotemporal dementia; “GC” refers to genome copy(ies); “g” refers to gram(s); “gDNA” refers to genomic DNA; “GM-CSF” refers to granulocyte-macrophage colony stimulating factor; “HMC3” refers to human microglial clone 3; “hr” refers hour(s); “ICM” refers to intra-cistema magna; “ICV” refers to intracerebroventricular; “iPSC” refers to induced pluripotent stem cell(s); “IRES” refers to internal ribosome entry site; “ITR” refers to inverted terminal repeat; “IV” refers to intravenous; “kg” refers to kilogram(s); “min” refers to minute(s); “mL” refers to milliliter; “MSD” refers to MesoScale Discovery; “Nab” refers to neutralizing antibody(ies); “NfL” refers to neurofilament light chain; “NHD” refers to Nasu-Hakola Disease; “NHP” refers to non-human primate(s); “nt” refers to nucleotide(s); “pg” refers to picogram(s); “PLX” refers to PLX3397 (also known as pexidartinib; 5-((5-chloro-lH-pyrrolo[2,3-b]pyridin-3-yl)methyl)-N-((6- (trifluoromethyl)pyri din-3 -yl)methyl)pyridin-2-amine or C20H15CIF3N5); “qRT-PCR” refers real-time quantitative reverse transcription polymerase chain reaction; “rAAV” refers to recombinant adeno-associated virus; “RBS” refers to Rep binding site; “RNA” refers to ribonucleic acid; “SC” refers to subcutaneous; “sec” refers to second(s); “SEM” refers to standard error of mean; “ss” refers to single-stranded; “THB” refers to triethylammonium bicarbonate buffer; “sTREM2” refers to soluble triggering receptor expressed on myeloid cells 2 protein; “TREM2” refers to triggering receptor expressed on myeloid cells 2 gene; “TREM2” refers to triggering receptor expressed on myeloid cells 2 protein; “trs” refers to terminal resolution site; “pL” refers to microliter; “VC” refers to vector copy(ies); “vg” refers to vector genome(s); “WPRE” refers to woodchuck hepatitis virus post -transcriptional regulatory element; “WT” refers to wild-type; and “YAC” refers to yeast artificial chromosome.

[0060] Certain definitions used herein are defined as follows:

[0061] As used herein, “AAV6TM” means an AAV6TM capsid protein having at least the following 3 mutations (z.e., triple mutant) in the amino acid sequence as compared to a wildtype (WT; see, NCBI Ref. Seq. No. AAB95450.1) AAV6 capsid protein amino acid sequence: T492V, Y705F and Y731F (see, e.g., SEQ ID NO: 19).

[0062] As used herein, “about” means within a statistically meaningful range of a value or values such as, for example, a stated concentration, length, molecular weight, pH, sequence similarity, time frame, temperature, volume, etc. Such a value or range can be within an order of magnitude typically within 20%, more typically within 10%, and even more typically within 5% of a given value or range. The allowable variation encompassed by “about” will depend upon the particular system under study, and can be readily appreciated by one of skill in the art.

[0063] As used herein, “administer,” “administering,” “administration” and the like mean providing a substance (e.g., a synthetic nucleic acid, expression construct, vector or rAAV herein) to an individual in a manner that is pharmacologically useful (e.g., to treat a disease, disorder, condition or symptom in the individual).

[0064] As used herein, “codon-optimized” means, with respect to a nucleotide sequence such as a gene of interest (e.g., TREM2), an alteration of codons or sequences in the gene or coding regions therein to reflect typical codon usage of a host organism (e.g., a mammal such as a human) or cell thereof without altering the polypeptide encoded by the nucleotide sequence. A codon-optimized transgene therefore is optimized for expression in a particular organism, organ, tissue or cell type, especially a mammal or mammalian organ, tissue or cell type. Alternatively, “codon-optimized” means an alteration of codons or sequences in a gene to improve protein expression as compared to a sequence that lacks the alteration by, for example, eliminating or changing sites that may be latent splice sites, stop codons, miRNA recognition sequences and the like. An entire nucleotide sequence may be codon-optimized or only one or more parts, portions or regions of a nucleotide sequence may be codon-optimized.

[0065] As used herein, “comparison window” means a contiguous and specified segment of a nucleotide sequence or amino acid sequence, where the sequence in the comparison window may include additions and/or deletions (z.e., gaps) compared to a reference sequence (which does not include the additions and/or deletions) for optimal alignment of the two sequences. Generally, the comparison window is at least 10 contiguous nucleotides/amino acids in length, and optionally can be 20, 30, 40, 50, 60, 70, 80, 90 or 100 nucleotides/amino acids, or longer. [0066] As used herein, “complementary” means a structural relationship between two nucleotides (e.g., on two opposing nucleic acids or on opposing regions of a single nucleic acid strand) that permits the two nucleotides to form base pairs (bp) with one another. For example, a purine nucleotide of one nucleic acid that is complementary to a pyrimidine nucleotide of an opposing nucleic acid may base pair together by forming hydrogen bonds with one another. Complementary nucleotides can base pair in the Watson-Crick manner or in any other manner that allows for the formation of stable duplexes. Likewise, two nucleic acids may have regions of multiple nucleotides that are complementary with each other to form regions of complementarity, as described herein.

[0067] As used herein, “CpG depleted,” with regard to a nucleotide sequence, means that all (z.e., 100%) known CpG sites are eliminated/removed in the nucleotide sequence.

[0068] As used herein, “CpG minimized,” with regard to a nucleotide sequence, means that some (z.e., < 100%) CpG sites, but not all, are eliminated/removed in the nucleotide sequence. [0069] As used herein, “CpG site” and the like means where a cytosine (C) nucleotide is followed by a guanine (G) nucleotide in the linear sequence of bases along a 5' to 3' direction in a nucleotide sequence.

[0070] As used herein, “TREM2-associated disease or disorder associated” means a disease or disorder resulting from a mutation in TREM2 causing changes in TREM2 expression, TREM2 amount and/or TREM2 activity/function as compared to its expected physiology. Examples of such diseases or disorders include, but are not limited to, AD, amyotrophic lateral sclerosis (ALS), ALSP, cognitive deficit, frontotemporal dementia (FTD), NHD, memory loss, multiple sclerosis, spinal cord injury and traumatic brain injury.

[0071] As used herein, “effective amount” means an amount, concentration or dose of a therapeutic agent (e.g., a nucleic acid, expression construct, vector or rAAV herein), or a pharmaceutical composition thereof, upon single or multiple dose administration to an individual in need thereof, provides a desired effect in such an individual under diagnosis or treatment (z.e., may produce a clinically measurable difference in a condition of the individual or may prevent a worsening in the individual). An effective amount can be readily determined by one of skill in the art by using known techniques and by observing results obtained under analogous circumstances. In determining the effective amount for an individual, a number of factors are considered, including, but not limited to, the species of mammal, its size, age and general health, the specific disease, disorder, condition or symptom involved, the degree of or involvement or the severity of the disease, disorder, condition or symptom, the response of the individual, the therapeutic agent administered, the mode of administration, the bioavailability characteristics of the preparation administered, the dose regimen selected, the use of concomitant medication, and other relevant circumstances.

[0072] As used herein, “expression construct” means a nucleotide sequence capable of expressing a nucleotide sequence of interest (e.g., a transgene or an inhibitory nucleic acid) when transformed, transfected or transduced into a target cell, tissue, organ or individual. An exemplary expression construct is a vector, such as a viral vector, especially an AAV vector or a baculovirus vector. Here, an expression construct can include at least one expression control element operably linked to the nucleotide sequence of interest such as a transgene (and/or an inhibitory nucleic acid). In this manner, the expression construct can be the expression control element, such as a promoter, in operable interaction with the transgene (and/or inhibitory nucleic acid), which is capable of directing the expression of the transgene (and/or inhibitory nucleic acid) in a cell, tissue, organ or individual, especially brain tissue.

[0073] As used herein, “expression control element” means a nucleotide sequence for a promoter, polyadenylation signal, transcription or translation termination sequence, upstream regulatory domain, origin of replication, internal ribosome entry site (IRES), enhancer and the like, which collectively provide for replication, transcription and/or translation of a desired nucleic acid (e.g., a transgene or an inhibitory nucleic acid) in a cell, tissue, organ or individual. Not all of these control sequences need always be present so long as the desired nucleotide sequence is capable of being replicated, transcribed and translated in the appropriate cell, tissue, organ or individual.

[0074] As used herein, “in combination with” means administering a therapeutic agent (e.g., nucleic acid, vector, rAAV or composition herein) either simultaneously, sequentially or in a single combined formulation with one or more additional therapeutic agents.

[0075] As used herein, “individual” means any mammal, including cats, dogs, mice, rats, and primates, especially humans. Moreover, “subject” or “patient” may be used interchangeably with “individual.”

[0076] As used herein, “individual in need thereof’ means a mammal, such as a human, with a condition, disease, disorder or symptom requiring treatment or therapy, including for example, those listed herein. In particular, the preferred individual to be treated is a human.

[0077] As used herein, “inhibitory nucleic acid” means a nucleic acid molecule capable of attenuating, reducing or preventing expression of a gene or mRNA. Exemplary inhibitory nucleic acids include, but are not limited to, shRNA, siRNA, miRNA, amiRNA, etc. Here, an inhibitory nucleic acid may be a nucleotide sequence encoding for an antisense sequence to a nucleotide sequence of interest.

[0078] As used herein, “nucleic acid” means a polymer of nucleotides. Although it may comprise any type of nucleotide units, the term generally applies to nucleotide polymers of deoxyribonucleic acid (DNA) or ribonucleic acid (RNA). Polynucleotide is used to include single-stranded (ss) nucleic acids, double-stranded (ds) nucleic acids, and DNA and RNA made from nucleotide or nucleoside analogues that may be identified by their sequences, which are generally presented in the 5' to 3' direction (as the coding strand), where the 5' and 3' indicate the linkages formed between the 5' hydroxyl group of one nucleotide and the 3'-hydroxyl group of the next nucleotide. For a coding strand presented in the 5'-3' direction, its complement (or non-coding strand) is the strand that hybridizes to that sequence according to Watson-Crick base pairing. Thus, as used herein, the complement of a nucleic acid such as a polynucleotide is the same as the “reverse complement” and describes the nucleic acid that in its natural form, would be based paired with the nucleic acid in question.

[0079] As used herein, “nucleoside” means a nucleobase-sugar combination, where the nucleobase portion is normally a heterocyclic base. The two most common classes of such heterocyclic bases are purines and pyrimidines. The sugar is normally a pentose sugar such as a ribose or a deoxyribose (e.g., 2'-deoxyribose).

[0080] As used herein, “nucleotide” means an organic molecule having a nucleoside (a nucleobase such as, for example, adenine, cytosine, guanine, thymine or uracil; and a pentose sugar such as, e.g., ribose or 2'-deoxyribose) and a phosphate group, which can serve as a monomeric unit of nucleic acid polymers such as DNA and RNA.

[0081 ] As used herein, “oligonucleotide” means a short nucleic acid molecule (e.g. , less than about 100 nucleotides in length). An oligonucleotide may be ss or ds.

[0082] As used herein, “operably linked” and the like means that the elements of an expression construct (or other nucleic acid construct) are configured to perform their usual function (z.e., under the influence of an expression control element). Thus, an expression control element (e.g., a promoter) operably linked to a desired nucleotide sequence (e.g., a transgene or an inhibitory nucleic acid) is capable of effecting expression of the desired nucleic acid. The control element need not be contiguous with the desired nucleotide sequence, so long as it functions to direct the expression thereof (z.e., maintain proper reading frame). Thus, for example, intervening untranslated, yet transcribed, sequence can be present between a promoter and the desired nucleotide sequence, and the promoter still can be considered “operably linked” to the desired nucleotide sequence.

[0083] As used herein, “pharmaceutically acceptable,” when referring to a material such as a carrier or diluent, means that it does not abrogate the biological activity or properties of a therapeutic agent (e.g., a nucleic acid, expression construct, vector, rAAV or composition herein) and is relatively non-toxic (z.e., the material may be administered to an individual without causing undesirable biological effects or interacting in a deleterious manner with any of the components of the composition in which it is contained.

[0084] As used herein, “pharmaceutically acceptable carrier” means a pharmaceutically acceptable material, composition or carrier, such as a liquid or solid filler, stabilizer, dispersing agent, suspending agent, diluent, excipient, thickening agent, solvent or encapsulating material, involved in carrying or transporting a therapeutic agent within or to an individual such that it may perform its intended function. Additional ingredients that may be included in the pharmaceutical compositions used in the practice of the invention are known in the art and described, for example in Remington’s Pharmaceutical Sciences, 21 ^st Edition, University of the Sciences in Philadelphia, PA (2006).

[0085] As used herein, “pharmaceutical composition” means a composition or therapeutic agent (e.g., a nucleic acid, expression construct, vector, rAAV or composition herein), mixed with at least one pharmaceutically acceptable chemical component, such as, but not limited to carriers, stabilizers, diluents, dispersing agents, suspending agents, thickening agents, excipients and the like.

[0086] As used herein, “recombinant adeno-associated virus,” “recombinant AAV” and “rAAV” mean viral particles comprising a rAAV vector encapsidated by AAV capsid protein. [0087] As used herein, “recombinant adeno-associated virus vector,” “recombinant AAV vector” and “rAAV vector” mean a synthetic polynucleotide vector comprising one or more heterologous sequences (z.e., nucleic acid sequence not of an AAV origin) that are flanked by at least one AAV ITR sequence. Such rAAV vectors can be replicated and packaged into infectious viral particles when present in a host cell that has been infected with a suitable helper virus (or that is expressing suitable helper functions) that expresses AAV rep and cap gene products (z.e., AAV Rep and Cap proteins).

[0088] As used herein, “sequence identity,” in the context of two nucleotide sequences or two amino acid sequences, means that residues in the two sequences that are the same when aligned for maximum correspondence over a specified comparison window.

[0089] As used herein, “synthetic” means a nucleic acid or other molecule or compound that is artificially synthesized (e.g., using a machine such as, for example, a solid phase nucleic acid synthesizer) or that is recombinantly produced (z.e., not naturally derived from a natural source that normally produces the nucleic acid or other compound). [0090] As used herein, “transgene” means a nucleotide sequence that is introduced into a cell and is capable of being transcribed into RNA and optionally, translated and/or expressed under appropriate conditions. The transgene confers a desired property to a cell into which it was introduced, or otherwise leads to a desired therapeutic or diagnostic outcome. Here, the transgene can be a nucleotide sequence encoding for a polypeptide of interest such as, for example, TREM2.

[0091] As used herein, “treat,” “to treat,” “treatment” or “treating” mean a process where there may be a slowing, controlling, delaying or stopping of the progression of the diseases or disorders disclosed herein, or ameliorating disease or disorder symptoms, but does not necessarily indicate a total elimination of all disease or disorder symptoms. Treatment and the like includes administration of a nucleic acid, expression construct, vector, rAAV or composition herein for treatment of a disease or disorder in an individual, particularly in a human.

[0092] As used herein, “TREM2” means human triggering receptor expressed on myeloid cells-2 (also known as PLOSL2, TREM-2, Trem2a, Trem2b or Trem2c).

[0093] As used herein, “vector” means a recombinant plasmid or recombinant virus that includes an oligonucleotide or polynucleotide to be delivered into a host cell, either in vitro or in vivo. Examples of vectors include, but are not limited to, bacterial artificial chromosome (BAC), cosmid, phagemid, plasmid, viral vector and yeast artificial chromosome (YAC).

[0094] As used herein, “viral vector” means a vector that is derived from a naturally occurring or modified virus, especially a rAAV vector or a Baculovirus vector (e.g., Autographa californica nuclear polyhedrosis (AcNPV) vector)).

[0095] Compositions

[0096] Synthetic Nucleic Acids

[0097] TREM2 -Encoding Transgenes: The synthetic nucleic acid herein can be a transgene that includes a nucleotide sequence encoding TREM2 (z.e., TREM2-encoding transgene). The TREM2-encoding transgene can be codon-optimized and/or CpG minimized and/or CpG depleted. While a CpG depleted nucleotide sequence has 100% of CpG sites eliminated/removed, a CpG minimized nucleotide sequence has < 100% of CpG sites eliminated/removed. For example, a CpG minimized nucleotide sequence can have from about 1% to about 99%, from about 10% to about 90%, from about 20% to about 80%, from about 30% to about 70%, from about 40% to about 60% or about 50% CpG sites removed. Alternatively, a CpG minimized nucleotide sequence can have from about 5% to about 10%, about 10% to about 20%, about 20% to about 30%, about 40% to about 50%, about 50% to about 60%, about 60% to about 70%, about 70% to about 80%, about 80% to about 90%, or about 90% to about 99% CpG eliminated/removed. Alternatively, a CpG minimized nucleotide sequence can have about 1%, about 5%, about 10% about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95% or about 99% CpG sites eliminated/removed.

[0098] In some instances, the TREM2-encoding transgene includes a nucleotide sequence having at least about 90% sequence identity to SEQ ID NO:3. Alternatively, the nucleotide sequence has at least about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or about 100% sequence identity to SEQ ID NO:3. In certain instances, the nucleotide sequence for the TREM2-encoding transgene is SEQ ID NO:3, which is a codon-optimized sequence.

[0099] In other instances, the TREM2-encoding transgene includes a nucleotide sequence having at least about 90% sequence identity to SEQ ID NO:4. Alternatively, the nucleotide sequence has at least about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or about 100% sequence identity to SEQ ID NO:4. In certain instances, the nucleotide sequence for the TREM2-encoding transgene is SEQ ID NO:4, which is not only a codon-optimized but also a CpG-depleted sequence.

[00100] In some instances, the TREM2-encoding transgene includes a nucleotide sequence having at least about 90% sequence identity to SEQ ID NO:5. Alternatively, the nucleotide sequence has at least about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or about 100% sequence identity to SEQ ID NO:5. In certain instances, the nucleotide sequence for the TREM2-encoding transgene is SEQ ID NO:5, which is not only a codon-optimized but also a 10% CpG-minimized sequence.

[00101] In some instances, the TREM2-encoding transgene includes a nucleotide sequence having at least about 90% sequence identity to SEQ ID NO:6. Alternatively, the nucleotide sequence has at least about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or about 100% sequence identity to SEQ ID N0:6. In certain instances, the nucleotide sequence for the TREM2-encoding transgene is SEQ ID NO:6, which is not only a codon-optimized but also a 25% CpG-minimized sequence.

[00102] The synthetic nucleic acids herein may exist on their own or may exist as part of an expression construct, a vector or a rAAV as further described herein.

[00103] Expression Constructs: As noted above, synthetic nucleic acids such as a TREM2- encoding transgenes can be incorporated in an expression construct. In some instances, the expression construct can include at least one expression control element operably linked to a TREM2-encoding transgene having a nucleotide sequence of any one of SEQ ID NOS:3 to 6 (or a nucleotide sequence with at least about 90% to about 99% sequence identity thereto). In certain instances, the nucleotide sequence for the TREM2-encoding transgene is SEQ ID NO:3, 4, 5 or 6.

[00104] The at least one expression control element can be at least one transcription factor binding site, at least one repressor binding site, at least one promoter, at least one enhancer, at least one intron splice site, at least one post-transcriptional regulatory element, at least one polyadenylation signal, or combinations thereof.

[00105] When the at least one expression control element is a promoter, the promoter can be a CBA promoter or a CD68 promoter or a F4/80 promoter. In some instances, the promoter is the CBA promoter and has a nucleotide sequence having at least about 90% sequence identity to SEQ ID NO:7. In other instances, the promoter is the CD68 promoter and has a nucleotide sequence having at least about 90% sequence identity to SEQ ID NO:8. In other instances, the promoter is the F4/80 promoter and has a nucleotide sequence having at least about 90% sequence identity to SEQ ID NO:9 (see also, Inti. Patent Application Publication No. WO 2006/122141). In certain instances, the nucleotide sequence for the promoter is SEQ ID NO:8. [00106] When the at least one expression control element is an enhancer, the enhancer can be a CMVe and has a nucleotide sequence having at least about 90% sequence identity to SEQ ID NO: 10. In certain instances, the nucleotide sequence for the enhancer is SEQ ID NO: 10 [00107] When the at least one expression control element is a post -transcriptional regulatory element, the post-transcriptional regulatory element can be a WPRE and has a nucleotide sequence having at least about 90% sequence identity to SEQ ID NO: 11. In certain instances, the nucleotide sequence for the post-transcriptional regulatory element is SEQ ID NO: 11 [00108] When the at least one expression control element is a polyadenylation signal, the polyadenylation signal can be BGHpA tail and has a nucleotide sequence having at least about 90% sequence identity to SEQ ID NO: 12. In certain instances, the nucleotide sequence for the polyadenylation signal is SEQ ID NO: 12.

[00109] In some instances, the expression construct also can include a nucleotide sequence for one or more of an internal ribosomal entry site (IRES), a self-cleaving peptide coding sequence such as a T2A peptide.

[00110] In some instances, the expression construct includes at least one additional nucleotide sequence for another transgene and/or an inhibitory nucleic acid.

[00111] The expression constructs herein may exist on their own or may exist as part of a vector or even a rAAV as further described herein.

[00112] Vectors: As noted above, the synthetic nucleic acids or expression constructs can be incorporated into a vector, especially a viral vector such as an rAAV vector. A rAAV vector may comprise either the “plus strand” or the “minus strand” of the rAAV vector. In some instances, the rAAV vector is ss (e.g., ss DNA or ss RNA). In other instances, the rAAV vector is ds (e.g., ds DNA or ds RNA).

[00113] To aid in expression, rAAV vectors include ITRs that flank the nucleotide sequence of interest (e.g., a TREM2-encoding transgene). In some instances, the ITR sequences are full- length (i.e., are about 145 nt in length and contain a functional Rep binding site (RBS) and a terminal resolution site (trs)) and is the WT AAV2 ITR including a nucleotide sequence having at least about 90% sequence identity to SEQ ID NO: 13 or a reverse complementary sequence thereto. In certain instances, the nucleotide sequence for WT AAV2 ITR is SEQ ID NO: 13 or a reverse complementary sequence thereto. In other instances, the ITR is a modified ITR (i.e., includes an addition, deletion, substitution, etc.) and is a modified AAV2 ITR including a nucleotide sequence having at least about 90% sequence identity to SEQ ID NO: 14 a reverse complementary sequence thereto. In certain instances, the nucleotide sequence for the modified AAV2 ITR is SEQ ID NO: 13 or 14 or a reverse complementary sequence thereto.

[00114] The rAAV vectors also can include a TRY region as described in Francois et al. (2005) J. Virol. 79: 11082-11094, which can be located between an ITR (e.g., a 5' ITR) and the TREM2-encoding transgene. In some instances, the TRY region includes a nucleotide sequence having at least about 90% sequence identity to SEQ ID NO: 15. In certain instances, the nucleotide sequence for the TRY region is SEQ ID NO: 15.

[00115] Additional sequences can be included in the rAAV vector to assist in packaging the rAAV vector into capsid protein (i.e., a sequence that optimizes the size of the vector for packaging within capsid protein; a “stuffer sequence”). In some instances, such a stuffer sequence includes a nucleotide sequence having at least about 90% sequence identity to SEQ ID NO: 16 to 18. In certain instances, the nucleotide sequence for the stuffer sequence is SEQ ID NO: 16, 17 or 18.

[00116] The vectors herein may exist on their own or may exist as part of a rAAV as further described herein.

[00117] rAAV

[00118] As noted above, the vectors can be incorporated into a rAAV (z.e., rAAV vector encapsidated in AAV capsid protein). Here, the rAAV include a capsid protein that readily spreads through the CNS, particularly when introduced into the CSF space or directly into the brain parenchyma. Examples of capsid proteins that can cross the blood-brain barrier include, but are not limited to, a capsid protein having an AAV6, AAV9 or AAVrh.10 serotype.

[00119] Of particular interest herein is rAAV that can infect microglia via AAV6-based capsid proteins, especially AAV6TM capsid protein. In some instances, the AAV6TM capsid protein includes an amino acid sequence having at least about 95% sequence identity to SEQ ID NO: 19. In certain instances, the amino acid sequence for the AAV6TM capsid protein is SEQ ID NO: 19.

[00120] In this manner, an rAAV herein includes (a) an AAV vector including a TREM2- encoding transgene and (b) an AAV6 capsid protein.

[00121] In some instances, the rAAV includes:

(a) a rAAV vector having, in 5' to 3' order, a nucleotide sequence of:

(i) a first AAV ITR or a reverse complementary sequence thereto,

(ii) a promoter,

(iii) a TREM2-encoding transgene,

(iv) a post-transcriptional regulatory element,

(v) a polyadenylation signal, and

(vi) a second AAV ITR a reverse complementary sequence thereto; and

(b) encapsidated in AAV6 capsid protein.

[00122] In other instances, the rAAV includes:

(a) a rAAV vector having, in 5' to 3' order, a nucleotide sequence of:

(i) a first AAV2 ITR having a nucleotide sequence of SEQ ID NO: 13 or 14, or a reverse complementary sequence thereto, (ii) a promoter having a nucleotide sequence of SEQ ID NO:8,

(iii) a TREM2-encoding transgene having a nucleotide sequence of SEQ ID NO:3,

(iv) a post-transcriptional regulatory element having a nucleotide sequence of SEQ ID NO: 11,

(v) a polyadenylation signal having a nucleotide sequence of SEQ ID NO: 12, and

(vi) a second AAV2 ITR having a nucleotide sequence of SEQ ID NO: 13 or 14, or a reverse complementary sequence thereto; and

(b) encapsidated in AAV6 capsid protein, especially AAV6TM capsid protein having an amino acid sequence of SEQ ID NO: 19.

[00123] In yet other instances, the rAAV includes:

(a) a rAAV vector having, in 5' to 3' order, a nucleotide sequence of:

(ii) a first AAV2 ITR having a nucleotide sequence of SEQ ID NO: 13 or 14, or a reverse complementary sequence thereto,

(ii) a promoter having a nucleotide sequence of SEQ ID NO:8,

(iii) a TREM2-encoding transgene having a nucleotide sequence of SEQ ID NO:4,

(iv) a post-transcriptional regulatory element having a nucleotide sequence of SEQ ID NO: 11,

(v) a polyadenylation signal having a nucleotide sequence of SEQ ID NO: 12, and

(vi) a second AAV2 ITR having a nucleotide sequence of SEQ ID NO: 13 or 14, or a reverse complementary sequence thereto; and

(b) encapsidated in AAV6 capsid protein, especially AAV6TM capsid protein having an amino acid sequence of SEQ ID NO: 19.

[00124] In yet other instances, the rAAV includes:

(a) a rAAV vector having, in 5' to 3' order, a nucleotide sequence of:

(i) a first AAV2 ITR having a nucleotide sequence of SEQ ID NO: 13 or 14, or a reverse complementary sequence thereto,

(ii) a promoter having a nucleotide sequence of SEQ ID NO:8, (iii) a TREM2-encoding transgene having a nucleotide sequence of SEQ ID NO:5,

(iv) a post-transcriptional regulatory element having a nucleotide sequence of SEQ ID NO: 11,

(v) a polyadenylation signal having a nucleotide sequence of SEQ ID NO: 12, and

(vi) a second AAV2 ITR having a nucleotide sequence of SEQ ID NO: 13 or 14, or a reverse complementary sequence thereto; and

(b) encapsidated in AAV6 capsid protein, especially AAV6TM capsid protein having an amino acid sequence of SEQ ID NO: 19.

[00125] In yet other instances, the rAAV includes:

(a) a rAAV vector having, in 5' to 3' order, a nucleotide sequence of:

(i) a first AAV2 ITR having a nucleotide sequence of SEQ ID NO: 13 or 14, or a reverse complementary sequence thereto,

(ii) a promoter having a nucleotide sequence of SEQ ID NO:8,

(iii) a TREM2-encoding transgene having a nucleotide sequence of SEQ ID NO: 6,

(iv) a post-transcriptional regulatory element having a nucleotide sequence of SEQ ID NO: 11,

(v) a polyadenylation signal having a nucleotide sequence of SEQ ID NO: 12, and

(vi) a second AAV2 ITR having a nucleotide sequence of SEQ ID NO: 13 or 14, or a reverse complementary sequence thereto; and

(b) encapsidated in AAV6 capsid protein, especially AAV6TM capsid protein having an amino acid sequence of SEQ ID NO: 19.

[00126] Any of the rAAV above further can include a stuffer sequence having a nucleotide sequence of SEQ ID NO: 16 to 18. In some instances, the stuffer sequence is positioned between the polyadenylation signal and the second AAV2 ITR.

[00127] In certain instances, the rAAV includes a rAAV vector having a nucleotide sequence of SEQ ID NO:21 encapsidated in AAV6 capsid protein, especially AAV6TM capsid protein having an amino acid sequence of SEQ ID NO: 19. In some instances, the rAAV includes a reverse complementary nucleotide sequence to SEQ ID NO:21. [00128] In certain other instances, the rAAV includes a rAAV vector having a nucleotide sequence of SEQ ID NO:22 encapsidated in AAV6 capsid protein, especially AAV6TM capsid protein (e.g., VP1) having an amino acid sequence of SEQ ID NO: 19. In some instances, the rAAV includes a reverse complementary nucleotide sequence to SEQ ID NO:22.

[00129] In certain other instances, the rAAV includes a rAAV vector having a nucleotide sequence of SEQ ID NO:23 encapsidated in AAV6 capsid protein, especially AAV6TM capsid protein having an amino acid sequence of SEQ ID NO: 19. In some instances, the rAAV includes a reverse complementary nucleotide sequence to SEQ ID NO:23.

[00130] In certain other instances, the rAAV includes a rAAV vector having a nucleotide sequence of SEQ ID NO:24 encapsidated in AAV6 capsid protein, especially AAV6TM capsid protein having an amino acid sequence of SEQ ID NO: 19. In some instances, the rAAV includes a reverse complementary nucleotide sequence to SEQ ID NO:24.

[00131] In certain other instances, the rAAV includes a rAAV vector having a nucleotide sequence of SEQ ID NO:25 encapsidated in AAV6 capsid protein, especially AAV6TM capsid protein having an amino acid sequence of SEQ ID NO: 19. In some instances, the rAAV includes a reverse complementary nucleotide sequence to SEQ ID NO:25.

[00132] In certain other instances, the rAAV includes a rAAV vector having a nucleotide sequence of SEQ ID NO:26 encapsidated in AAV6 capsid protein, especially AAV6TM capsid protein having an amino acid sequence of SEQ ID NO: 19. In some instances, the rAAV includes a reverse complementary nucleotide sequence to SEQ ID NO:26.

[00133] Pharmaceutical Compositions

[00134] The synthetic nucleic acids described herein (z.e., an expression construct or a vector) or rAAVs described herein can be formulated as a pharmaceutical composition including the synthetic nucleic acid or rAAV and a pharmaceutically acceptable carrier. Pharmaceutical compositions can be prepared by methods well known in the art (e.g. , Remington: The Science and Practice of Pharmacy, 22 ^nd ed. (Pharmaceutical Press, 2013).

[00135] In some instances, the pharmaceutical composition can be in the form of a formulation that includes not only a rAAV herein but also one or more of (a) about 10 mM to about 30 mM TRIS buffer, (b) about 0.5 mM to about 1.5 mM MgCh, (c) about 100 mM to about 300 mM NaCl and (d) about 0.001% (w/v) to about 0.01% (w/v) Poloxamer 188.

[00136] In some instances, the rAAV includes a rAAV vector having a nucleotide sequence of SEQ ID NO:21 encapsidated in AAV6 capsid protein, especially AAV6TM capsid protein (e.g., VP1) having an amino acid sequence of SEQ ID NO: 19. In some instances, the rAAV includes a reverse complementary nucleotide sequence to SEQ ID NO:21. In other instances, the rAAV includes a rAAV vector having a nucleotide sequence of SEQ ID NO:22 encapsidated in AAV6 capsid protein, especially AAV6TM capsid protein having an amino acid sequence of SEQ ID NO: 19. In some instances, the rAAV includes a reverse complementary nucleotide sequence to SEQ ID NO:22. In other instances, the rAAV includes a rAAV vector having a nucleotide sequence of SEQ ID NO:23 encapsidated in AAV6 capsid protein, especially AAV6TM capsid protein having an amino acid sequence of SEQ ID NO: 19. In some instances, the rAAV includes a reverse complementary nucleotide sequence to SEQ ID NO:23. In other instances, the rAAV includes a rAAV vector having a nucleotide sequence of SEQ ID NO:24 encapsidated in AAV6 capsid protein, especially AAV6TM capsid protein having an amino acid sequence of SEQ ID NO: 19. In some instances, the rAAV includes a reverse complementary nucleotide sequence to SEQ ID NO:24. In other instances, the rAAV includes a rAAV vector having a nucleotide sequence of SEQ ID NO:25 encapsidated in AAV6 capsid protein, especially AAV6TM capsid protein having an amino acid sequence of SEQ ID NO: 19. In some instances, the rAAV includes a reverse complementary nucleotide sequence to SEQ ID NO:25. In other instances, the rAAV includes a rAAV vector having a nucleotide sequence of SEQ ID NO:26 encapsidated in AAV6 capsid protein, especially AAV6TM capsid protein having an amino acid sequence of SEQ ID NO: 19. In some instances, the rAAV includes a reverse complementary nucleotide sequence to SEQ ID NO:26.

[00137] In some instances, the TRIS buffer can be at a concentration from about 10 mM to about 30 mM. In other instances, the TRIS buffer can be at a concentration from about 15 mM to about 25 mM, or about 20 mM. In yet other instances, the TRIS buffer can be at a concentration of about 10 mM, about 11 mM, about 12 mM, about 13 mM, about 14 mM, about 15 mM, about 16 mM, about 17 mM, about 18 mM, about 19 mM, about 20 mM, about 21 mM, about 22 mM, about 23 mM, about 24 mM, about 25 mM, about 26 mM, about 27 mM, about 28 mM, about 29 mM or about 30 mM.

[00138] In some instances, MgCh can be at a concentration from about 0.5 mM to about 1.5 mM. In other instances, MgCh can be at a concentration from about 0.6 mM to about 1.4 mM, from about 0.7 mM to about 1.3 mM, from about 0.8 mM to about 1.2 mM, from about 0.9 mM to about 1.1 mM, or about 1.0 mM. In yet other instances, MgCE can be at a concentration of about 0.5 mM, about 0.6 mM, about 0.7 mM, about 0.8 mM, about 0.9 mM, about 1.0 mM, about 1.1 mM, about 1.2 mM, about 1.3 mM, about 1.4 mM or about 1.5 mM.

[00139] In some instances, NaCl can be at a concentration from about 100 mM to about 300 mM. In other instances, NaCl can be at a concentration from about 125 mM to about 275 mM, from about 150 mM to about 250 mM, from about 175 mM to about 225 mM, or about 200 mM. In yet other instances, NaCl can be at a concentration of about 100 mM, about 110 mM, about 120 mM, about 130 mM, about 140 mM, about 150 mM, about 160 mM, about 170 mM, about 180 mM, about 190 mM, about 200 mM, about 210 mM, about 220 mM, about 230 mM, about 240 mM, about 250 mM, about 260 mM, about 270 mM, about 280 mM, about 290 mM or about 300 mM.

[00140] In some instances, Poloxamer 188 can be at a concentration from about 0.001% (w/v) to about 0.01% (w/v). In other instances, Poloxamer 188 can be at a concentration from about 0.002% (w/v) to about 0.009% (w/v), from about 0.003% (w/v) to about 0.008% (w/v), from about 0.004% (w/v) to about 0.007% (w/v), or from about 0.005% (w/v) to about 0.006% (w/v). In yet other instances, the Poloxamer 188 can be at a concentration of about 0.001% (w/v), about 0.002% (w/v), about 0.003% (w/v), about 0.004% (w/v), about 0.005% (w/v), about 0.006% (w/v), about 0.007% (w/v), about 0.008% (w/v), about 0.009% (w/v) or about 0.01% (w/v).

[00141] In certain instances, the formulation can include a rAAV herein and (a) about 20 mM TRIS (pH 8.0), (b) about 1 mM MgCh, (c) about 200 mM NaCl and (d) about 0.005% (w/v) Poloxamer 188.

[00142] In some instances, the effective amount can be a titer between about 10 ⁹ genome copies (GC)/kg to about 10 ¹⁴ GC/kg (e.g., about 10 ⁹ GC/kg, about 10 ¹⁰ GC/kg, about 10 ¹¹ GC/kg, about 10 ¹² GC/kg, about 10 ¹³ GC/kg, or about 10 ¹⁴ GC/kg). In some instances, the individual is administered a high titer (e.g., > 10 ¹² GC/kg of rAAV) by injection to the CSF space, especially via ICM.

[00143] In other instances, the effective amount can be a dose ranging from about l x 10 ¹² vg to about 1 x 10 ¹⁵ vg or about 1 x 10 ¹³ vg to about 7 x 10 ¹⁴ vg. In other instances, the dose can be about 3.5 x 10 ¹³ vg, about 7.0 x 10 ¹³ vg or about 1.4 x 10 ¹⁴ vg. In yet other instances, the dose can be about 1 x 10 ¹⁴ vg, about 2.0 x 10 ¹⁴ vg, or about 4.0 x 10 ¹⁴ vg. Alternatively, the dose can be about 2 x 10 ¹³ vg, about 3 x 10 ¹³ vg, about 4 x 10 ¹³ vg, about 5 x 10 ¹³ vg, about 6 x IO ¹³ vg, about 7 x 10 ¹³ vg, about 8 x 10 ¹³ vg, about 9 x 10 ¹³ vg, about 1 x 10 ¹⁴ vg, or about 2 x 10 ¹⁴ vg. In certain instances, the dose is 7.0 x 10 ¹³ vg or 1.4 x 10 ¹⁴ vg.

[00144] The pharmaceutical composition can be administered by any route including, for example, intra-arterial, intradermal, intramuscular, intrathecal, intravenous (IV), intraventricular, parenteral, subcutaneous (SC) or transdermal. Specifically contemplated routes are IV administration (e.g., systemic intravenous injection), and/or direct administration to an affected site (e.g., intracistemal magna (ICM) injection, intracerebroventricular (ICV) injection) or combinations thereof.

[00145] Generally, the most appropriate route of administration will depend upon a variety of factors including, but not limited to, the nature of the agent (e.g., its stability in the environment of its administration and/or intended target) and/or the condition of the individual (e.g. , whether the subject is able to tolerate oral administration). In some instances, the synthetic nucleic acids, rAAV or pharmaceutical compositions are suitable for administration to the CNS of an individual by, for example, intrathecal, ICM, ICV or combinations thereof.

[00146] Kits

[00147] In some instances, synthetic nucleic acids (z.e., an expression construct or a vector) or rAAVs herein can be included in a kit that includes the synthetic nucleic acid or rAAV and instructions for its use. In other instances, the kit includes the synthetic nucleic acid or rAAV and a package insert containing instructions for use of the kit and/or any component thereof. In yet other instances, the kit comprises, in a suitable container or other means for containing, the synthetic nucleic acid or rAAV, one or more controls, and various buffers, reagents, enzymes and other standard ingredients well known in the art. In some instances, the container comprises at least one vial, well, test tube, flask, bottle, syringe, or other container means, into which the synthetic nucleic acid, rAAV or other therapeutic oligonucleotide is placed, and in some instances, suitably aliquoted. In those instances where an additional component is provided, the kit includes additional containers into which this component is placed. The kits can also include a means for containing the synthetic nucleic acid or rAAV and any other reagent in close confinement for commercial sale. Such containers may include injection or blow-molded plastic containers into which the desired vials are retained. Containers and/or kits can include labeling with instructions for use and/or warnings.

[00148] In some instances, the kit includes the synthetic nucleic acid or rAAV and a pharmaceutically acceptable carrier, or a pharmaceutical composition including the synthetic nucleic acid, rAAV or other therapeutic oligonucleotide and instructions for treating or delaying progression of a neurodegenerative disease in an individual in need thereof.

[00149] In some instances, the kit includes the synthetic nucleic acid or rAAV and a pharmaceutically acceptable carrier or a pharmaceutical composition comprising the synthetic nucleic acid or rAAV and instructions for administering the synthetic nucleic acid or rAAV or pharmaceutical composition.

[00150] Methods

[00151] Methods of Making

[00152] Methods of making rAAVs are described, for example, in Samulski et al. (1989) J. Virol. 63:3822-3828 and Wright (2009) Hum. Gene Ther. 20:698-706. In some instances, the rAAV can be produced in a Baculovirus vector expression system (BEVS). Production of rAAVs using BEVS are described, for example, in Urabe et al. (2002) Hum. Gene Ther. 13: 1935-1943, Smith et al. (2009) Mol. Ther. 17: 1888-1896, as well as US Patent Nos. 8,945,918 and 9,879,282, and Inti. Patent Application Publication Nos. WO 2017/184879 and WO 2022/082017. Alternatively, the rAAV can be produced in human embryonic kidney (e.g., HEK293) cells (see, e.g., Inti. Patent Application Publication Nos. WO 2020/210689 and WO 2022/035900). However, the rAAV can be produced using any suitable method (e.g., using recombinant rep and cap genes).

[00153] Methods of Treatment and Uses

[00154] The nucleic acids, expression constructs, vectors, rAAV or pharmaceutical compositions described herein can be used for treating a neurological disease such as, for example, AD, ALS, ALSP, cognitive deficit, FTD, memory loss, NHD, spinal cord injury, traumatic brain injury, or multiple sclerosis.

[00155] The methods can include the steps described herein, and these maybe be, but not necessarily, carried out in the sequence as described. Other sequences, however, also are conceivable. Moreover, individual or multiple steps bay be carried out either in parallel and/or overlapping in time and/or individually or in multiply repeated steps. Furthermore, the methods may include additional, unspecified steps.

[00156] Here, the synthetic nucleic acids, rAAV or pharmaceutical compositions including the same may be used in a method to treat TREM2 -associated diseases and disorders, where such method includes at least a step of administering to an individual in need of such treatment an effective amount of a synthetic nucleic acid, rAAV or a pharmaceutical composition including the same.

[00157] In some instances, the synthetic nucleic acid, rAVV or pharmaceutical composition is administered via an IV injection. In other instances, the synthetic nucleic acid, rAVV or pharmaceutical composition is administered via an ICM injection of the individual.

[00158] When a rAAV, the effective amount can be a titer between about 10 ⁹ genome copies (GC)/kg to about 10 ¹⁴ GC/kg (e.g., about 10 ⁹ GC/kg, about IO ¹⁰ GC/kg, about 10 ¹¹ GC/kg, about 10 ¹² GC/kg, about 10 ¹³ GC/kg, or about 10 ¹⁴ GC/kg). In some instances, the individual is administered a high titer (e.g., > 10 ¹² GC/kg of rAAV) by injection to the CSF space, especially via ICM.

[00159] In other instances, the effective amount can be a dose ranging from about 1 x 10 ¹² vg to about 1 x 10 ¹⁵ vg or about 1 x 10 ¹³ vg to about 7 x 10 ¹⁴ vg. In other instances, the dose can be about 3.5 x 10 ¹³ vg, about 7.0 x 10 ¹³ vg or about 1.4 x 10 ¹⁴ vg. In yet other instances, the dose can be about 1 x 10 ¹⁴ vg, about 2.0 x 10 ¹⁴ vg, or about 4.0 x 10 ¹⁴ vg. Alternatively, the dose can be about 2 x 10 ¹³ vg, about 3 x 10 ¹³ vg, about 4 x 10 ¹³ vg, about 5 x 10 ¹³ vg, about 6 x 10 ¹³ vg, about 7 x 10 ¹³ vg, about 8 x 10 ¹³ vg, about 9 x 10 ¹³ vg, about 1 x 10 ¹⁴ vg, or about 2 x 10 ¹⁴ vg. In certain instances, the dose is 7.0 x 10 ¹³ vg or 1.4 x 10 ¹⁴ vg.

[00160] In some instances, the rAAV or composition including the same can be administered to a subject once or multiple times (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 20 or more).

[00161] In some instances, the individual has or is suspected of having a disease or disorder associated with TREM2, especially AD, ALSP or NHD.

[00162] In some instances, the individual is between the ages of about 1 month old to about 10 years old (e.g., about 1 month, 2 months, 3 months, 4, months, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months, 11 months, 12 months, 13 months, 14 months, 15 months, 16 months, 17 months, 18 months, 19 months, 20 months, 21 months, 22 months, 23 months, 24 months, 3 years, 4 years, 5 years, 6 years, 7 years, 8 years, 9 years, 10 years, or any age therebetween). In other instances, the individual is between about 10 years old to about 20 years old (e.g., about 10 years, 11 years, 12 years, 13 years, 14 years, 15 years, 16 years, 17 years, 18 years, 19 years, 20 years, or any age therebetween). In other instances, the individual is older than 20 years old (e.g., about 21 years, 22 years, 23 years, 24 years, 25 years, 26 years, 27 years, 28 years, 29 years, 30 years, or any age therebetween), older than 30 years old (e.g., about 31 years, 32 years, 33 years, 34 years, 35 years, 36 years, 37 years, 38 years, 39 years, 40 years, or any age therebetween), older than 40 years old (e.g., about 41 years, 42 years, 43 years, 44 years, 45 years, 46 years, 47 years, 48 years, 49 years, 50 years, or any age therebetween), or even older than 50 years old (e.g., about 51 years, 52 years, 53 years, 54 years, 55 years, 56 years, 57 years, 58 years, 59 years, 60 years, 70 years, 80 years, 90 years, or any age therebetween).

[00163] In some instances, the individual has a pathogenic CSF1R mutation.

[00164] In some instances, the method also can include a step of measuring TREM2 in plasma and/or CSF and/or urine from the individual and comparing an obtained value to an earlier- obtained comparable value or to a control value to assess effectiveness of the method. In some instances, the TREM2 is soluble TREM2 (sTREM2).

[00165] In other instances, the method also can include a step of measuring CSF1R in plasma and/or CSF and/or urine from the individual and comparing an obtained value to an earlier- obtained comparable value or to a control value to assess effectiveness of the method. In some instances, the CSF1R is soluble CSF1R (sCSFIR).

[00166] In other instances, the method also can include a step of measuring NfL in plasma and/or CSF and/or urine from the individual and comparing an obtained value to an earlier- obtained comparable value or to a control value to assess effectiveness of the method.

[00167] In other instances, the method also can include a step of measuring Chitl in plasma and/or CSF and/or urine from the individual and comparing an obtained value to an earlier- obtained comparable value or to a control value to assess effectiveness of the method.

[00168] In other instances, the method also can include a step of measuring GM-CSF in plasma and/or CSF and/or urine from the individual and comparing an obtained value to an earlier-obtained comparable value or to a control value to assess effectiveness of the method.

[00169] Uses

[00170] A rAAV herein or pharmaceutical composition including the same can be used, or adapted for use, to treat an individual (e.g., a human) having or suspected of having a disease or disorder associated with TREM2, such as AD, ALSP or NHD. As such, the rAAV or pharmaceutical composition including the same is provided for use, or adapted for use, to treat an individual having or suspected of having a disease or disorder associated with TREM2, such as AD, ALSP or NDH. Also, the rAAV or pharmaceutical composition including the same is provided for use, or adaptable for use, in the manufacture of a medicament or a pharmaceutical composition for treating a disease or disorder associated with TREM2, such as AD, ALSP or NHD.

[00171] Also described are nucleic acids, vectors, rAAVs or pharmaceutical compositions for use in therapy. Furthermore, nucleic acids, vectors, rAAVs or pharmaceutical compositions are described herein for use in the treatment of a neurological disease such as, for example, AD, ALS, ALSP, cognitive deficit, FTD, memory loss, NHD, spinal cord injury, traumatic brain injury or multiple sclerosis.

[00172] Also described are use of nucleic acids, vectors, rAAVs or pharmaceutical compositions in the manufacture of a medicament for the treatment of a neurological disease such as, for example, AD, ALS, ALSP, cognitive deficit, FTD, memory loss, NHD, spinal cord injury, traumatic brain injury or multiple sclerosis.

EXAMPLES

[00173] The following non-limiting examples are offered for purposes of illustration, not limitation.

[00174] Example 1 : Generating rAAV

[00175] Methods: rAAV vectors expressing various TREM2-encoding transgenes were generated using cells, such as HEK293 cells or Sf9 insect cells see, e.g., Inti. Patent Application Nos. WO 2008/024988, 2017/184879 and WO 2022/082017). The ITR sequences flank an expression construct having a promoter/enhancer element for the transgene, a 3' poly A signal, and posttranslational signals such as the WPRE element.

[00176] The rAAV vectors were encapsidated in either AAV6TM capsid protein (SEQ ID NO: 19) or AAV9 capsid protein (SEQ ID NO: 20).

[00177] IN VITRO STUDIES

[00178] Example 2: In Vitro Transducing of a Human Microglial Cell Line with TREM-2 rAAV

[00179] Methods: A HMC3 microglial cell line was transduced with a rAAV vector having a TREM2 transgene (SEQ ID NO:3) encapsidated in AAV6TM capsid protein (SEQ ID NO: 19) or in AAV9 capsid protein (SEQ ID NO:20) at a range of MOIs. 72 hr post-transduction, supernatant was collected for protein and cells were lysed for RNA. [00180] TREM2 mRNA was quantified using qRT-PCR and normalized to GAPDH. TREM2 protein was quantified using a MSD assay.

[00181] Results: FIGS. 5A-5B show that TREM2 mRNA expression (FIG. 5 A) and protein expression (FIG. 5B) were observed with both rAAV; however, rAAV vector encapsidated in AAV6TM capsid protein performed better than the same rAAV vector encapsidated in AAV9 capsid protein.

[00182] Example 3: In Vitro Transducing of a Human Microglial Cell Line with Alternative TREM-2 rAAV

[00183] Methods: HMC3 cells were transduced with a first rAAV vector having a TREM2 transgene (SEQ ID NO:25) or a second rAAV vector having a TREM2 transgene (SEQ ID NO:26) encapsidated in AAV6TM capsid protein (SEQ ID NO: 19) over a range of MOI from 1.09x l0 ⁵ to 3.50* 10 ⁶ vg/cell. 3 days post-infection, supernatant was collected for ELISA.

[00184] Results: Both rAAV effectively transduced HMC3 cells in vitro, resulting in a robust, dose-dependent expression and secretion of TREM2 (FIGS 6A-6B, where FIG 6A is the first rAAV vector and FIG. 6B is the second rAAV vector). In contrast, a minimal level of TREM2 (78 pg/mL) was detected in the negative control group treated with excipient alone.

[00185] Example 4: Determining In Vitro Potency of TREM2in Human Induced Pluripotent Stem Cell (iPSC)-Derived Microglia Cells

[00186] Methods: Human iPSC-derived microglia either were treated with excipient (negative control) or were transduced with a rAAV vector having a TREM2 transgene (SEQ ID NO:26) encapsidated in AAV6TM capsid protein (SEQ ID NO: 19) at a MOI ranging from 2.0* 10 ³ to 2.0* 10 ⁶ vg/cell. 7 days post-transduction, cells were lysed for ELISA or were processed for RNA. All iPSC-derived cells were obtained from FujiFilm Cellular Dynamics, Inc. (Madison, WI).

[00187] Results: By Day 7, rAAV transduction resulted in robust, dose-dependent expression of TREM2 that was detected in groups treated at 2.0* 10 ⁵ and 2.0* 10 ⁶ vg/cell of about 1000 pg/mL and 1250 pg/mL, respectively. In contrast, TREM2 was detected from excipient-treated iPSC-derived microglia at a concentration of about 625 pg/mL. Furthermore, rAAV transduction resulted in a dose-dependent increase in mRNA expression of TREM2 at all dose groups, while no TREM2 mRNA was detected from excipient -treated microglia. [00188] Example 5: Improving Cell Viability with a TREM2 rAAV in a Pharmacological CSFIR-Inhibition Assay in Human iPSC-Derived Microglia Cells

[00189] Methods: The iPSC-derived microglia as described in Example 4 either were treated with excipient (negative control) or were transduced with a rAAV vector having a TREM2 transgene (SEQ ID NO:26) encapsidated in AAV6TM capsid protein (SEQ ID NO: 19) at a MOI of 2.0 ^x 10 ³ vg/cell. 7 days post-transduction, 150 nM or 200 nM of PLX3397 in 0.1% DMSO was added to the cell culture media and incubated for additional 8 hr at 37°C and 5% CO2. After this period of incubation, cells were assayed for viability using a colorimetric kit (Abeam).

[00190] Results: PLX treatment caused cytotoxicity and reduced the total number of microglial cells (z.e., about 80% cell death); whereas rAAV transduction significantly reduced the PLX-induced toxicity to 50% as compared to control levels.

[00191] IN VIVO STUDIES

[00192] Example 6: In Vivo Rodent Studies of AAV Capsid Proteins

[00193] Methods: 36 one-month old 5xFAD mice were allocated into 3 groups (n=12/group) and treated with the same rAAVs as in Example 2 or excipient (20 mM Tris pH 8.0, 200 mM NaCl and 1 mM MgCh + 0.001 % Pluronic F68)) via ICV injection (10 pl - 5 pl bilateral per mouse) to determine efficacy. A group (n=12) of non-transgenic, age-matched littermates (i.e., WT) were similar injected with excipient as a control. Treatment doses were 5.68 x 10 ¹⁰ vg/animal or 1.8 x 10 ¹¹ vg/animal. 5 months post-ICV injection, the presence of vg was assessed by ddPCR.

[00194] In addition, efficacy endpoints including protein levels, amyloid-P levels (both biochemical and immunohistochemical) and inflammation were examined. TREM2 protein and amyloid-P levels were measured using a MSD assay. Iba ⁺ cells were measured by immunohi stochemi stry .

[00195] In vivo blood collection: In vivo blood was collected by mandibular bleeding from the facial vein/artery plexus without anesthesia before treatment start, and at 2, 3, 4 and 5 months of age (5 time points). Collected blood was then transferred into serum gel clotting activator microtube. After incubation for at least 20 min (60 min maximum) at room temperature, serum was prepared from the samples by centrifugation (10000 ^x g, 5 min, room temperature). After centrifugation, serum was frozen on dry ice and stored at -80 °C.

[00196] Tissue sampling: At 6 months, the mice were terminally anesthetized by i.p. injection of Pentobarbital (600 mg/kg) and CSF, blood, brain, several organs (gonads, kidney, heart, liver, lung and spleen) and spinal cord were collected for biochemical, immunochemical and/or histological analysis.

[00197] Immunohistology: For each incubation a uniform systematic random set of 5 sections per mouse from all animals per group was selected (one section each from levels 2, 4, 6, 8, 10). All sections were counterstained with the nuclear dye DAPI. Binding of primary antibodies was visualized using highly cross-absorbed secondary antibodies.

[00198] Imaging: Whole slide scans of the stained sections were recorded on a Zeiss automatic microscope AxioScan Z1 with high aperture lenses, equipped with a Zeiss Axiocam 506 mono and a Hitachi 3CCD HV-F202SCL camera and Zeiss ZEN 3.3 software.

[00199] Sample Preparation - Homogenization: Hippocampus and somatosensory cortex from all animals were homogenized in 14 volumes PBS at 55 Hz for 50 sec using UPHO bead mill (Geneye) and 3 aliquots were generated (30 pL for RNA/DNA isolation, 50 pL for soluble insoluble protein isolation and rest).

[00200] Sample Preparation - DNA and RNA Isolation: DNA and RNA were isolated from hippocampus and somatosensory cortex from all animals using the Ambion TriZOL kit from a 30 pL PBS aliquot. The amount and quality of total extracted RNA was evaluated by UV-VIS spectrometry using a NanoDrop 1000 spectrophotometer. RNA (1 pg of each sample) was reverse transcribed by using the iScript gDNA Clear cDNA Synthesis Kit.

[00201] Amyloid-P40 and Amyloid-P42 Sample Preparation and Measurement (soluble and insoluble): A second aliquot of 50 pl of hippocampus and cortex samples was substituted with the same volume of 2x THB buffer (2x THB; 500 mM Sucrose, 2 mM EDTA, 2 mM EGTA, 40 mM Tris pH 7.4) including lx protease inhibitor (Calbiochem) vortexed and incubated for 15 min on ice. The THB homogenate was then processed for extraction of soluble and deposited proteins from brain homogenates for analysis of amyloid-P40 and amyloid-P42. For extraction of non-plaque associated proteins, 50 pl THB homogenate were mixed with 1-part DEA solution (0.4 % DEA, 100 mM NaCl). The mixture was centrifuged for 120 min at 20,000x g, 4°C. The supernatant was neutralized with 1/10 of the volume 0.5 M Tris-HCl, pH 6.8 and vortexed briefly. Aliquots were stored at -80°C. For extraction of deposited proteins, 30 pL THB homogenate were mixed with 2.2 parts cold FA, sonicated for 30 sec on ice and centrifuged for 120 min at 20,000 x g, 4°C. The supernatant was mixed with 19 parts FA Neutralization Solution (IM Tris, 0.5 M Na2HPO4, 0.05 % NaNs). Aliquots were stored at - 80°C. Soluble and insoluble fraction of hippocampus and somatosensory cortex samples from all animals were then analyzed by immunosorbent assays from Mesoscale Discovery (Amyloid-P40 Peptide (6E10) Kit and Amyloid-P42 Peptide (6E10) Kit) according to the instructions of the manufacturer. Amyloid-P levels in study samples were evaluated in comparison to calibration curves provided in the kit and are expressed as pg/mg brain.

[00202] Statistics: Statistical analysis was performed in GraphPad Prism 9. Data was tested for normality using the Kolmogorow-Smirnow-Test. If normal distribution was confirmed, differences between groups were tested with the one or two-way ANOVA / Mixed-effects analysis followed by Bonferroni, Dunnett’s or Sidak’s multiple comparisons test. If data was not normally distributed, differences between groups were tested with Kruskal -Wallis-Test, followed by Dunn’s post hoc test for multiple comparisons. The 5xFAD-excipient group was used as reference group. Data are presented as mean + or +/- SEM.

[00203] Results: FIGS. 7A-7D show both capsid proteins resulted in broad biodistribution in the cortex, hippocampus, cervical spinal cord and liver.

[00204] FIGS. 8A-8C show both capsid proteins resulted in TREM2 protein levels in CSF, liver and serum, where the AAV6TM capsid protein showed higher TREM2 protein levels in the CSF than the AAV9 capsid protein.

[00205] FIGS. 9A-9C show that the rAAV vector encapsidated in AAV6TM capsid protein significantly decreased amyloid-P in the hippocampus and cortex whereas the rAAV vector encapsidated in AAV9 capsid protein did not decrease amyloid-P in the hippocampus and cortex.

[00206] FIGS. 10A-10B show that rAAV vector decreased inflammatory markers in cortex regardless of capsid protein.

[00207] Overall, rAAV vector dosing resulted in broad biodistribution and dose-dependent increases in TREM2 protein levels for both capsids; however, significant reduction of cortical disease burden was observed only in mice treated with AAV6TM capsid protein.

[00208] Example 7: In Vivo Rodent Studies of TREM2 rAAV [00209] Methods: Excipient or 2 different doses of one or the other of the TREM2 rAAVs of Example 3 were administered to 1 -month old WT C57BL/6 mice by ICV injection (n=10/group). The rAAV were injected at a dose of either 5.68* IO ¹⁰ vg (1.42xlO ⁿ vg/g brain) or 1.8* 10 ¹¹ vg (4.49* 10 ¹¹ vg/g brain), respectively. Vector particles per gram brain weight were calculated based on an adult mouse brain weight of 400 mg.

[00210] CNS tissues and liver were collected to analyze biodistribution by ddPCR. CSF, liver, and serum were collected to analyze TREM2 expression.

[00211] Biodistribution was determined by measuring vg presence using ddPCR (with >50 vg/1 pg gDNA defined as positive).

[00212] Results: Mice that received rAAV were positive for vg in the cortex indicating that ICV administration successfully results in comparable transduction in the brain. There was no significant effect of vector design on biodistribution (all ps > 0.05). ICV administration also resulted in vg presence in the spinal cord and liver.

[00213] TREM2 expression was measured in the CSF and serum using MSD. ICV injection of rAAV increased TREM2 in the CSF at both doses. These data indicate that there is no difference in TREM2 expression levels generated by either rAAV, consistent with the biodistribution data. Additionally, ICV injection of rAAV significantly increased levels of TREM2 in the blood at 1.8X10 ¹¹ vg. A dose-dependent effect in serum was observed when comparing the doses.

[00214] Example 8: Assessing In Vivo Efficacy of TREM2 rAAV in a CSFIR-Inhibition Mouse Model for CRL

[00215] Methods: Excipient or 2 different doses of a TREM2 rAAV of Example 3 (i.e., a rAAV vector of SEQ ID NO:26 encapsidated in AAV6TM capsid protein (SEQ ID NO: 19)) were administered to 1-month old WT C57BL/6 mice by ICV injection (n=10). The rAAV was injected at a dose of either 1.8x l0 ¹⁰ vg (4.49x lO ¹⁰ vg/g brain) or 1.8xl0 ⁿ vg (4.49x lO ⁿ vg/g brain), respectively. Vector particles per g brain weight was based on an adult mouse brain weight of 400 mg. At 8 weeks of age (i.e., 4 weeks post-ICV injection), 1 excipient-treated group and both dose-treated groups were fed with PLX-containing mouse chow. Mice were fed with PLX-chow at 185 mg/kg for 1 week and then sacrificed at 9 weeks of age (i.e., 5 weeks post-ICV injection). [00216] Biodistribution was determined by measuring vg presence using ddPCR (with > 50 vg/1 pg gDNA defined as positive).

[00217] Results: Mice that received rAAV were positive for vg in all the brain tissues tested, indicating that ICV administration successfully resulted in transduction in the brain. ICV administration also resulted in vg presence in the spinal cord, with lower levels of transduction observed in the liver. Reduced biodistribution in the liver was reflected in the lower TREM2 levels in this tissue.

[00218] Inhibition of CSF1R by PLX treatment was apparent at both the mRNA and protein level in mice. There was a statistically significant improvement in CSF1R expression at the mRNA level in mice treated with both doses of rAAV + PLX as compared to mice treated with excipient + PLX. The level of sCSFIR fragment in the CSF of animals treated with both doses of rAAV + PLX exhibited a positive trending pattern toward excipient levels. Thus, rAAV treatment ameliorates the negative effect of PLX in the CSFIR-inhibition model.

[00219] Moreover, dosing of mice with excipient + PLX resulted in almost complete depletion of microglia throughout the brain. In fact, there was a statistically significant increase in the number of IBA1 ⁺ microglia in mice that were treated with either dose of rAAV + PLX as compared to mice treated with excipient + PLX. This observation was further confirmed by analysis of AIF1 mRNA expression level, a gene that encodes IB Al, using qRT-PCR. Furthermore, a significant therapeutic benefit of rAAV was evident in higher mRNA levels of AIF1, in mice treated with either dose + PLX as compared to treatment group with excipient + PLX.

[00220] Furthermore, PLX treatment decreased expression of genes associated with homeostatic microglia, such as P2RY12 and HEXB, which are known to be stably expressed in the microglia in the homeostatic state. In mice treated with rAAV + PLX, a significantly higher expression of both P2RY12 and HEXB was observed.

[00221 ] Consistent with the PLX model-effect that was observed in universal and homeostatic genes, disease associated microglia (DAM) genes (e.g., CCE12, CD48, ('1)68, LYZ1 and PTPRC) also showed a significant decrease in expression in the excipient + PLX treatment group. In contrast, expression of these genes was fully, or in some cases partially, elevated by rAAV, in most cases to normal levels as compared to excipient-only treated animals. [00222] Example 9: TREM2 rAAV Dose-Ranging in a CSFIR-Inhibition Mouse Model for CRL

[00223] Methods: Excipient or a TREM2 rAAV of Example 3 (z.e., a rAAV vector of SEQ ID NO:26 encapsidated in AAV6TM capsid protein (SEQ ID NO: 19)) were administered to 4- week-old WT C57BL/6 mice by ICV injection (n=16/group). TREM2 rAAV was injected at the following 3 doses: 1.33 MO ¹¹ vg (3.33 MO ¹¹ vg/g brain), 1.8 x lO ¹⁰ vg (4.49x lO ¹⁰ vg/g brain) or 2.44x 10 ⁹ vg (6.10x 10 ⁹ vg/g brain). Vector particles per g brain weight was based on an adult mouse brain weight of 400 mg.

[00224] At 8 weeks of age (z.e., 4 weeks post-ICV injection), 1 excipient-treated group and both 1.33X 10 ¹¹ vg- and 2.44x l0 ⁹ -treated groups were treated with PLX3397. Mice were fed with PLX3397-chow at 185 mg/kg for 1 week and then sacrificed at 9 weeks of age (z.e., 5 weeks post-ICV injection).

[00225] Biodistribution, TREM2 expression levels, microglial panel gene expression, and safety through histopathology was assessed. Biodistribution was determined using ddPCR, which was developed in alignment with the FDA’s guidance “Long-Term Follow-up After Administration of Human Gene Therapy Products” (2020; with limit of quantification of < 50 copies/pg of gDNA). TREM2 expression was measured in the CSF using an MSD assay (as above, sTREM2 was used as a proxy for protein production within the tissues).

[00226] Results: All mice that received TREM2 rAAV were positive for vg in all brain regions tested, indicating that ICV administration successfully resulted in widespread transduction in the brain. Additionally, ICV administration of TREM2 rAAV resulted in reduced vector transduction in the liver as compared to brain.

[00227] ICV administration of TREM2 rAAV robustly increased levels of TREM2 in the CSF at all 3 doses. Lower biodistribution of TREM2 rAAV in peripheral tissues was reflected in lower TREM2 levels in the liver, as well as lower TREM2 levels in serum. A dose -dependent effect was observed in the serum when comparing the 1.33X 10 ¹¹ vg and 2.44x l0 ⁹ vg doses.

[00228] Out of the 3 doses tested in this study, the 2.44x 10 ⁹ vg dose exhibited no efficacious benefit in almost any of the microglial genes tested (z.e., a subtherapeutic dose). The other two doses exhibited significant improvements in microglial gene expression profile in TREM2 rAAV + PLX-treated mice as compared to excipient +PLX-treated mice, in almost 90% of genes with model effect. [00229] Taken together, the 1.33xl0 ⁿ vg and 1.8* IO ¹⁰ vg doses suppressed PLX-induced microglial damage in the brainstem of the CSFIR-inhibition mouse model. This effect also was observed in different brain regions, including the hippocampus, cortex and cerebellum.

[00230] Example 10: In Vivo Non-Human Primate (NHP) Studies of TREM2 rAAV

[00231] Methods: 2-4-year-old female cynomolgus monkeys were administered a rAAV vector having a TREM2 transgene of SEQ ID NO:3 encapsidated in AAV6TM capsid protein (SEQ ID NO: 19) or excipient via ICM injection. One-month post-ICM injection, the presence of vg, TREM2 and safety were assessed.

[00232] Results: FIG. 11 shows that AAV6TM capsid protein resulted in broad biodistribution in NHPs.

[00233] FIGS. 12A-12C show that the AAV6TM capsid protein resulted in increased TREM2 protein levels in liver, spinal cord, CSF and serum.

[00234] Overall, in the NHP safety studies, all animals survived and there was no test article- related variations in mortality, clinical signs, body weights/body weight gains, body temperature and neurological parameters in either group dosed via a single injection into the ICM. No findings from examination of general attitude, behavior, motor function, proprioception and postural reaction. Clinical pathology revealed minimal changes in hematology, coagulation and clinical chemistry. Histopathology findings were limited and comparable to AAV9 capsid protein in previous NHP studies. In the high dose, microscopic findings of minimal to mild mononuclear cell infiltration in the brain and DRG were noted.

[00235] Both AAV9 capsid protein and AAV6TM capsid proteins transduced microglial cells in vitro with the rAAV vector encoding TREM2; however, AAV6TM capsid protein drove higher TREM2 mRNA and protein expression as compared to AAV9 capsid protein. In vivo, both rAAV demonstrated broad biodistribution and dose-dependent increases in TREM2 protein levels. Additionally, AAV6TM capsid protein exhibited comparable biodistribution to AAV9 capsid protein in key brain regions in the rodent studies while showing substantially less liver tropism.

[00236] Example 11 : In Fzvo NHP Studies of TREM2 rAAV

[00237] Methods: Excipient or a TREM2 rAAV of Example 3 (a rAAV vector of SEQ ID NO:25 encapsidated in AAV6TM capsid protein (SEQ ID NO: 19)) or vehicle were administered to female cynomolgus monkeys via ICM (n=2/group). TREM2 rAAV was injected at the following 2 doses: 3.32>< 10 ¹² vg (4.49>< 1O ¹⁰ vg/g brain) or 1.05>< 10 ¹³ vg (1.42x 1011 vg/g brain). The NHPs were sacrificed 29 days after injection to detect potential early toxicity of TREM2 rAAV, with the expectation of capturing potential toxicity at a time in which biodistribution throughout the brain and peripheral organs was expected to be near its maximum.

[00238] Biodistribution, TREM2 expression levels, microglial panel gene expression, and safety through histopathology was assessed. Biodistribution was determined using ddPCR as described in Example 9. TREM2 expression was measured in the CSF using an MSD assay (as above, sTREM2 was used as a proxy for protein production within the tissues).

[00239] Results: All NHPs survived to Day 29. There were no test article -related variations observed in mortality, clinical signs, body weights/body weight gains, body temperature or neurological signs in either group dosed with a single injection of TREM2 rAAV into the ICM at the 3.32x l0 ¹² dose or 1.05x l0 ¹³ total vg dose.

[00240] Minimal changes in hematology and coagulation parameters were observed in NHPs treated with the 1.05x l0 ¹³ vg dose. These changes consisted of an increased neutrophil count at Days 15 and 29, with increased fibrinogen levels at Day 15 as compared to baseline values, indicating a potential acute phase response.

[00241] There were no TREM2 rAAV-related macroscopic observations. Test article-related microscopic observations included minimal to mild mononuclear cell infiltration in the brain in all NHPS administered > 3.32x l0 ¹² vg. The minimal to mild mononuclear cell infiltration was perivascular in distribution and also present in the pia. The mononuclear cells were primarily lymphocytes, and the distribution and severity were greater in the animals receiving the 3.32x l0 ¹² vg dose.

[00242] Given that the only treatment-related findings were minimal increases in neutrophils and fibrinogen and minimal to mild mononuclear infiltration of the brain and ganglia, ICM injection of either dose of TREM2 rAAV was judged to be well -tolerated in NHPs.

[00243] As for biodistribution, all tissues tested were positive in NHPs administered TREM2 rAAV, indicating widespread distribution throughout the CNS and periphery.

[00244] In the CSF, TREM2 rAAV administration increased TREM2 in the 1.05x l0 ¹³ vg dose. In contrast, a dose-dependent effect of the 2 doses on TREM2 was observed in serum and liver. [00245] Taken together, the results showed broad distribution throughout the brain, comparable to the levels shown to be efficacious in mouse models. This transduction leads to the elevation of TREM2 in the brain. Furthermore, all NHPs survived, and postmortem pathology analysis indicated minimal test article-related increases in neutrophils and fibrinogen, as well as minimal to mild mononuclear infiltration of the brain and ganglia. Consequently, the TREM2 rAAV demonstrated a generally favorable safety profile in NHPs.

[00246] Example 12: In Ezvo NHP Studies of TREM2 rAAV

[00247] Methods: Excipient or a TREM2 rAAV of Example 3 (a rAAV vector of SEQ ID NO:26 encapsidated in AAV6TM capsid protein (SEQ ID NO: 19)) or vehicle will be administered to cynomolgus monkeys (male and female) via ICM (n=6/group). TREM2 rAAV will be injected at one of the following 3 doses: 3.32* 10 ¹² vg (4.49* 10 ¹⁰ vg/g brain), 1.05x 10° vg (1.42x lO ⁿ vg/g brain) or 3.32x 10° vg (4.49x 10° vg/g brain). The NHPs will be followed for 29 days or 26 weeks after injection to assess tolerability and safety.

[00248] This study also will assess peak vector distribution in the brain at about 4 weeks postadministration, as well as post-peak assessments at 183 days post-administration. Control NHPs will receive an ICM administration of the same volume of formulation buffer excipient including 20 mM Tris (pH 8.0), 1 mM MgCE, and 200 mM NaCl containing 0.005% (w/v) Poloxamer 188. All NHPs will be screened for AAV6 neutralizing antibodies (NAb) with a 1 :20 titer cut-off.

SEQUENCE LISTING

[00249] The following nucleic and/or amino acid sequences are referred to in the disclosure above and are provided below for reference.

[00250] SEQ ID NO: 1 - human TREM2 protein isoform 1 (230 aa)

MEPLRLLILLFVTELSGAHNTTVFQGVAGQSLQVSCPYDSMKHWGRRKAWCRQLGE KGPCQRWSTHNLWLLSFLRRWNGSTAITDDTLGGTLTITLRNLQPHDAGLYQCQSL HGSEADTLRKVLVEVLADPLDHRDAGDLWFPGESESFEDAHVEHSISRSLLEGEIPFP

PTSILLLLACIFLIKILAASALWAAAWHGQKPGTHPPSELDCGHDPGYQLQTLPGLR D T

[00251] SEQ ID NO:2 - human TREM2 protein isoform 2 (219 aa)

MEPLRLLILLFVTELSGAHNTTVFQGVAGQSLQVSCPYDSMKHWGRRKAWCRQLGE KGPCQRWSTHNLWLLSFLRRWNGSTAITDDTLGGTLTITLRNLQPHDAGLYQCQSL HGSEADTLRKVLVEVLADPLDHRDAGDLWFPGESESFEDAHVEHSISRAERHVKED

DGRKSPGEVPPGTSPACILATWPPGLLVLLWQETTLPEHCFSWTLEAGTG

[00252] SEQ ID NO:3 - Artificial sequence (TREM2 transgene 1; 690 nt) atggagcccctgcgcctgctgatcctgctgttcgtgaccgagctgagcggcgcccacaac accaccgtgttccagggcgtggccgg ccagagcctgcaggtgagctgcccctacgacagcatgaagcactggggccgccgcaaggc ctggtgccgccagctgggcgagaa gggcccctgccagcgcgtggtgagcacccacaacctgtggctgctgagcttcctgcgccg ctggaacggcagcaccgccatcacc gacgacaccctgggcggcaccctgaccatcaccctgcgcaacctgcagccccacgacgcc ggcctgtaccagtgccagagcctgc acggcagcgaggccgacaccctgcgcaaggtgctggtggaggtgctggccgaccccctgg accaccgcgacgccggcgacctgt ggttccccggcgagagcgagagcttcgaggacgcccacgtggagcacagcatcagccgca gcctgctggagggcgagatcccctt cccccccaccagcatcctgctgctgctggcctgcatcttcctgatcaagatcctggccgc cagcgccctgtgggccgccgcctggca cggccagaagcccggcacccacccccccagcgagctggactgcggccacgaccccggcta ccagctgcagaccctgcccggcct gcgcgacacc

[00253] SEQ ID NO:4 - Artificial sequence (TREM2 transgene 2; 690 nt) atggagcccctgaggctgctcatcctgctgtttgtgacagaactgtctggagcccacaac accacagtgttccagggagttgctggcca gtctctgcaagtgtcttgcccctatgacagcatgaagcactggggaaggaggaaggcttg gtgtaggcagctgggagagaaaggac cttgccagagggtggtgagcacacacaacctgtggctgctgagcttcctcagaaggtgga atggctctacagccatcacagatgaca ccctgggtggcaccctcaccatcaccttaaggaacctgcagcctcatgatgctggcctgt accaatgccagagcctgcatggctctga ggctgataccctcaggaaggtgttggtggaggtgctggctgatcctctggatcacaggga tgctggagacctgtggttcccaggaga gtctgagagctttgaggatgcccatgtggagcacagcatcagcaggtctcttctggaggg agagatccccttccctcccacaagcatc ctgttgctgcttgcctgcatcttcctgatcaagatccttgctgcttctgctctttgggct gctgcctggcatggccagaaacctggaacaca tcctccctctgaactggactgtggccatgaccctggctaccagttgcaaaccttgcctgg cttgagggacacc

[00254] SEQ ID NO: 5 - Artificial sequence (TREM2 transgene 3; 690 nt) atggagcccctgaggctgctgatcctgctgtttgtgacagaactgtctggagcccacaac accacagtgttccagggagttgctggcc agtctctgcaagtgtcttgcccctatgacagcatgaagcactggggaaggaggaaggctt ggtgtaggcagctgggagagaaagga ccttgccagagggtggtgagcacacacaacctgtggctgctgagcttcctcagaaggtgg aatggctctacagccatcacagatgac accctgggtggcaccctcaccatcaccctgaggaacctgcagcctcatgatgctggcctg taccaatgccagagcctgcatggctctg aggctgataccctcaggaaggtgttggtggaggtgctggctgatcctctggatcacaggg atgctggagacctgtggttcccaggaga gtctgagagctttgaggatgcccatgtggagcacagcatcagcaggtctcttctggaggg agagatccccttccctcccacaagcatc ctgttgctgcttgcctgcatcttcctcatcaagatccttgctgcttctgctctttgggct gctgcctggcatggccagaaacctggaacaca tcctccctctgaactggactgtggccatgaccctggctaccagttgcaaaccttgcctgg cttgagggacacc

[00255] SEQ ID N0:6 - Artificial sequence (TREM 2 transgene 4; 690 nt) atggagcccctgagactgctcatcctgctgtttgtgacagaactgtctggggcccacaac accacagtgttccagggggtggctggcc agtccctccaggtgtcctgcccctatgactccatgaagcactgggggagaagaaaggctt ggtgtagacagctgggggagaaaggg ccttgccagagagtggtgtccacacacaacctgtggctgctgtccttcctgagaagatgg aatggctccacagccatcacagatgaca ccctggggggcaccctcaccatcaccctgagaaacctgcagcctcatgatgctggcctgt accagtgccagtccctgcatggctctga ggctgataccctgagaaaggtgctggtggaggtgctggctgatcctctggatcacagaga tgctggggacctgtggttcccagggga gtctgagtcctttgaggatgcccatgtggagcactccatctccagatccctgctggaggg ggagatccccttccctcccacatccatcct gctgctgctggcctgcatcttcctcatcaagatcctggctgcttctgctctgtgggctgc tgcctggcatggccagaaacctgggacaca tcctccctctgaactggactgtggccatgaccctggctaccagctgcagaccctgcctgg cctgagagacacc

[00256] SEQ ID N0:7 - Chicken [3-actin promoter (283 nt) catggtcgaggtgagccccacgttctgcttcactctccccatctcccccccctccccacc cccaattttgtatttatttattttttaattattttgt gcagcgatgggggcggggggggggggggggcgcgcgccaggcggggcggggcggggcgag gggcggggcggggcgag gcggagaggtgcggcggcagccaatcagagcggcgcgctccgaaagtttccttttatggc gaggcggcggcggcggcggccctat aaaaagcgaagcgcgcggcgggcg [00257] SEQ ID NO:8 - CD68 promoter (715 nt) gatatcaaactgcctgtttgggcttctcatttcttacctccccttccctctcccacctgc tactgggtgcatctctgctccccccttccccagc agatggttacctttgggctgttgctttcttgtcaccatctgagttctcagacgctggaaa gccatgttctcggctctgtgaatgacaatgctg actggagtgctgcccctctgtaaagggctgggtgtggatggtcacaagcccctcacatgc ctcagccaagaggaagtagtacagggg tcagcccagaggtccaggggaaaggagtggaaaccgatttccccaccaagggaggggcct gtacctcagctgttcccatagcttact tgccacaactgccaagcaagtttcgctgagtttgacacatggatccctgtggatcaactg ccctaggactccgtttgcacccatgtgaca ctgttgactttgccctgacgaagcagggccaacagtcccctaacttaattacaaaaacta atgactaagagagaggtggctagagctga ggcccctgagtcaggctgtgggtgggatcatctccagtacaggaagtgagactttcattt cctcctttccaagagagggctgagggag cagggttgagcaactggtgcagacagcctagctggactttgggtgaggcggttcagccat atcgaattctgctggggctactggcag

[00258] SEQ ID NO: 9 - F4/80 promoter (2068 nt) gaattctttgtttaggtctgtatgccatatttaatagggttatttggttctctggagtct aacttcctgagttctttgtatattttggatatcagtcct ctatcagatgttgggttggtaaagatttttttcccccaatctgtgagttgctgttttgtc atattgacagtctcctttgctttacagaagctttttaa tttcatgaggtcccatttgtcaattgttgatcttagagcctgagccattggtgttctgta caggacaaattttcccctgtgccaatatgttcga ggttcttccccattttcttttctattagactcagtgtatctggttttatgtggaggtcct tgatccactcggacttgagctttgtacaaggagata agaatggatcagtttgtatttttctacatcctaactgccagttgaactagcaacatttgt tgaaaatgtttttttttcttccctcactggatgacttt gacttctttgtcaaagatcaagtgaccataggtgtgtaggttcacttttgggtcttcaat tttattccattgatctacctttctgtcattgtatcaa taccatgaaggttttttaaaaaaaaatttttttaatcatattgctctgcagtacagcttg aggtcagggatggtgattcccccagaggctcttt gattgttgagaacggtttttgctatcctgggttttttgttattccagatgaatttgagtt gggattttgatagggaatacactgaatttgtagatt gcttttggcaagatggccatctttccatcttctgaggtctttgatttctttcttcagaga cttgaagttcttgtcatacagatctttcaattgcttg gttagagtcacaccaagatattttatattatttgtgactattgtgaagggtgtcatttct ctaatttatttctcagcccgtttatcctttgagtaga ggaaaactactgatttgtttgaattaattttatatctagccactaagctgaagttgttta tcagctgtaggagttctctggtggattattttttagg gttatttatgtattctaccacatcatctgcaaatagtgatatcttgacttcctcctttct aatttgtatccctttgacctccttttgttgcctaattgtt ctgggtagaactttgagtactatattaaatgataggggaaaagtggacagccttgggtgt ggtgtgtgtggtatggtgtggtgtgtgtatg gtgatgatgcatgtggtgtgtatggtgtggtgtatgtggtgtggtgtgtggtggtggtga taatgatgatggtggtctctctgtctctgtctgt ctctttctatctctgcctctctctctctgtctctctctgcccccctctgtctccctctgt ctgtctgtctgtctctttggtgtgatgtatgtggtgtgt gtgtattctacaaggttgacatgatgacagaatttaattttcttagcagcaagctcatgg atcctggtgataaatgcagcatgactttactga aaaggctttgtgatcttgaagagtggattgacttcactgtcggcagcacatgcaatctca cttgtttggtgtaatgaaagaagagaatgag aggtggaagggggatggtaatgttgaaaaaaagaatggtacagaggaaactgaggttgga gagagatggggtagatggtaagagat ggagaaagagggaaggaaatggagagaaagacagagagacagagagagacacacagagag acacacagagacagagaggaa gggaaagggaaagagaaaggaagaggaagagggggaggggaaggggaaggggaaggggaa gggagagggagaaatgtgg acactagccagatttaagggagaaattagggggttgccagtctgtccacctctgatggtg gcaactcagcagaaagctgctgggctca gtctggctttgttgagcaaccctgactccaccccttttcttccccacaaagcaagctttt aaagggaaggctttcttcattgaatgactgcca cagtacgatg

[00259] SEQ ID NO: 10 - Cytomegalovirus enhancer (352 nt) aatagtaatcaattacggggtcattagttcatagcccatatatggagttccgcgttacat aacttacggtaaatggcccgcctggctgacc gcccaacgacccccgcccattgacgtcaataatgacgtatgttcccatagtaacgccaat agggactttccattgacgtcaatgggtgg agtatttacggtaaactgcccacttggcagtacatcaagtgtatcatatgccaagtacgc cccctattgacgtcaatgacggtaaatggc ccgcctggcattatgcccagtacatgaccttatgggactttcctacttggcagtacatct acgtattagtcatcgctattac

[00260] SEQ ID NO: 11 - Woodchuck hepatitis virus post-transcriptional regulatory element (594 nt) ttatcgataatcaacctctggattacaaaatttgtgaaagattgactggtattcttaact atgttgctccttttacgctatgtggatacgctgctt taatgcctttgtatcatgctattgcttcccgtatggctttcattttctcctccttgtata aatcctggttgctgtctctttatgaggagttgtggccc gttgtcaggcaacgtggcgtggtgtgcactgtgtttgctgacgcaacccccactggttgg ggcattgccaccacctgtcagctcctttcc gggactttcgctttccccctccctattgccacggcggaactcatcgccgcctgccttgcc cgctgctggacaggggctcggctgttggg cactgacaattccgtggtgttgtcggggaaatcatcgtcctttccttggctgctcgcctg tgttgccacctggattctgcgcgggacgtcc ttctgctacgtcccttcggccctcaatccagcggaccttccttcccgcggcctgctgccg gctctgcggcctcttccgcgtcttcgccttc gccctcagacgagtcggatctccctttgggccgcctccc

[00261] SEQ ID NO: 12 - Bovine Growth Hormone PolyA Signal Tail (235 nt) cgactagagctcgctgatcagcctcgactgtgccttctagttgccagccatctgttgttt gcccctcccccgtgccttccttgaccctgga aggtgccactcccactgtcctttcctaataaaatgaggaaattgcatcgcattgtctgag taggtgtcattctattctggggggtggggtg gggcaggacagcaagggggaggattgggaagacaatagcaggcatgctgggga

[00262] SEQ ID NO: 13 - wild-type AAV2 ITR (145 nt) aggaacccctagtgatggagttggccactccctctctgcgcgctcgctcgctcactgagg ccgggcgaccaaaggtcgcccgacgc ccgggctttgcccgggcggcctcagtgagcgagcgagcgcgcagagagggagtggccaa

[00263] SEQ ID NO: 14 - modified AAV2 ITR (141 nt) cctgcaggcagctgcgcgctcgctcgctcactgaggccgcccgggcaaagcccgggcgtc gggcgacctttggtcgcccggcctc agtgagcgagcgagcgcgcagagagggagtggccaactccatcactaggggttcct [00264] SEQ ID NO: 15 - Artificial sequence (TRY region; 60 nt) agctctgggtatttaagcccgagtgagcacgcagggtctccattttgaagcgggaggtta

[00265] SEQ ID NO: 16 - Artificial sequence (Stuffer sequence 1; 1229 nt) gcagtcctcgcatgcctgagtgccctgttctgcactcctctcctccccaccctgcccata ttcttcacaccctccttccctccccagtatca aaacctaccctcagactctttctgcatgtggacatgtaaggggctggtggactctggttg cattggaagggaaggagtgctaacagtgg catcccaggcactagctggcagttgggggaagctttggggggcactggcactggtaatag cctctgaaattataagccactaattatga gcccctacagttataaaggaggaaagaacctgaggatgttcatctgcatctttggggcac tctctcccctgctctgaaggtccccttgtcc tcagctcttgatgggaggtgaggagcgtagagatttccgttgctgagttgaggggaattg acacaccaaatttttgtacacaattgcttctt cccgctgaggaaagcgcaccttttgctcccccagagcactggacaaacctgggcaagagg agagacggtgccaaatggagtcttgtt cctgcagcttcttagatggtggtcagggggaaggcggggttctgaggcatggatgggagg tggtcagatgggagagggccacggc ccatctggtctctctagttccaaaaggcaagccaccaacttcccaatcctatctttcaag cctctgtcacagtaatggttgaagatggcag gcaagggaggaccaagagagaagtcaaagcagggggctctgggggctgccccagcaacag gctggtgctctaagcccatctccc ccgccctaaggagggttcccaaaatagcagcctcatgtctcccccaaaatatctccgaga cgggtccttcctgaaaggggaacaaag ccacagaaatagggaagctggaagttaaaggtcaggaaagtcggtaccaatgtgggcggc tgcagagcaagcaagagtggcggg gcagggagagccagccccaggccgagaggagaggtcccagtcccaaatgatggcaaagag atgtgcagaacagaactggaggg ggttttaagaccaagtgcctccagaatagacccagagaaggtcagttacttctccaagga ggcccagcaagagcaacagtgagaaaa ccaaacccaggtcccaggtttcctggttcccaatttcccacaaacacatgctgtgccatc cgctcccaacttgtataagaacctaagg

[00266] SEQ ID NO: 17 - Artificial sequence (Stuffer sequence 2; 1229 nt) gcagtcctcacttacctgagtgccctgttctgcactcctctcctccccaccctgcccata ttcttcacaccctccttccctccccagtatcaa aacctaccctcagactctttctgctgtgagactgataaggggctggtggactctggttgc attggaagggaaggagtgcttacagtggc atcccaggcactagctggcagttgggggaagctttggggggcactggcactggtaatagc ctctgaaattataagcctctcatttgaag cccctacagttataaaggaggaaagaacctgaggtgattcatctgcatctttggggcact ctctcccctgctctgaaggtccccttgtcct cagctcttgtgaggaggtcaggagcatagagatttccattgctgagttgaggggaactta cacaccaaatttttgtacacaattgcttcttc ccactgaggaaagcacaccttttgctcccccagagcactggacaaacctgggcaagagga gagacagtgcctgaaagagtcttgttc ctgcagcttcttagtgagtggtcagggggaaggcagggttctgaggctgagtgaggaggt ggtcagtgaggagagggccacagcc catctggtctctctagttccaaaaggcaagccaccaacttcccaatcctatctttcaagc ctctgtcacagttgaagttgaagtgagcagg caagggaggaccaagagagaagtcaaagcagggggctctgggggctgccccagcaacagg ctggtgctctaagcccatctcccc caccctaaggagggttcccaaaatagcagcctctcatctcccccaaaatatctccaagac aggtccttcctgaaaggggaacaaagcc acagaaatagggaagctggaagttaaaggtcaggaaagtcagtacctgtgaaggcagctg cagagcaagcaagagtggcagggca gggagagccagccccaggccaagaggagaggtcccagtccctgtgaaaagcaaagagtgt gacagaacagaactggagggggtt ttaagaccaagtgcctccagaatagacccagagaaggtcagttacttctccaaggaggcc cagcaagagcaacagtgagaaaacca aacccaggtcccaggtttcctggttcccaatttcccacaaacacttactgtgccatccac tcccaacttgtataagaacctaagg

[00267] SEQ ID NO: 18 - Artificial sequence (Stuff er sequence 3; 1254 nt) attgactgaattcccgtgcggaccggcagtcctcacttacctgagtgccctgttctgcac tcctctcctccccaccctgcccatattcttca caccctccttccctccccagtatcaaaacctaccctcagactctttctgctgtgagactg ataaggggctggtggactctggttgcattgg aagggaaggagtgcttacagtggcatcccaggcactagctggcagttgggggaagctttg gggggcactggcactggtaatagcct ctgaaattataagcctctcatttgaagcccctacagttataaaggaggaaagaacctgag gtgattcatctgcatctttggggcactctctc ccctgctctgaaggtccccttgtcctcagctcttgtgaggaggtcaggagcatagagatt tccattgctgagttgaggggaacttacaca ccaaatttttgtacacaattgcttcttcccactgaggaaagcacaccttttgctccccca gagcactggacaaacctgggcaagaggag agacagtgcctgaaagagtcttgttcctgcagcttcttagtgagtggtcagggggaaggc agggttctgaggctgagtgaggaggtgg tcagtgaggagagggccacagcccatctggtctctctagttccaaaaggcaagccaccaa cttcccaatcctatctttcaagcctctgtc acagttgaagttgaagtgagcaggcaagggaggaccaagagagaagtcaaagcagggggc tctgggggctgccccagcaacag gctggtgctctaagcccatctcccccaccctaaggagggttcccaaaatagcagcctctc atctcccccaaaatatctccaagacaggt ccttcctgaaaggggaacaaagccacagaaatagggaagctggaagttaaaggtcaggaa agtcagtacctgtgaaggcagctgca gagcaagcaagagtggcagggcagggagagccagccccaggccaagaggagaggtcccag tccctgtgaaaagcaaagagtgt gacagaacagaactggagggggttttaagaccaagtgcctccagaatagacccagagaag gtcagttacttctccaaggaggccca gcaagagcaacagtgagaaaaccaaacccaggtcccaggtttcctggttcccaatttccc acaaacacttactgtgccatccactccca acttgtataagaacctaagg

[00268] SEQ ID NO: 19 - AAV6TM capsid protein (736 aa)

MAADGYLPDWLEDNLSEGIREWWDLKPGAPKPKANQQKQDDGRGLVLPGYKYLG PFNGLDKGEPVNAADAAALEHDKAYDQQLKAGDNPYLRYNHADAEFQERLQEDTS FGGNLGRAVFQAKKRVLEPFGLVEEGAKTAPGKKRPVEQSPQEPDSSSGIGKTGQQP AKKRLNFGQTGDSESVPDPQPLGEPPATPAAVGPTTMASGGGAPMADNNEGADGV GNASGNWHCDSTWLGDRVITTSTRTWALPTYNNHLYKQISSASTGASNDNHYFGYS TPWGYFDFNRFHCHFSPRDWQRLINNNWGFRPKRLNFKLFNIQVKEVTTNDGVTTIA NNLTSTVQVFSDSEYQLPYVLGSAHQGCLPPFPADVFMIPQYGYLTLNNGSQAVGRS SFYCLEYFPSQMLRTGNNFTFSYTFEDVPFHSSYAHSQSLDRLMNPLIDQYLYYLNRT QNQSGSAQNKDLLFSRGSPAGMSVQPKNWLPGPCYRQQRVSKVKTDNNNSNFTWT GASKYNLNGRESIINPGTAMASHKDDKDKFFPMSGVMIFGKESAGASNTALDNVMIT DEEEIKATNP VATERFGT VAVNLQS S STDP ATGD VHVMGALPGMVWQDRD VYLQG PIWAKIPHTDGHFHPSPLMGGFGLKHPPPQILIKNTPVPANPPAEFSATKFASFITQYST GQVSVEIEWELQKENSKRWNPEVQYTSNFAKSANVDFTVDNNGLYTEPRPIGTRFLT RPL

[00269] SEQ ID NO:20 - AVV9 capsid protein (736 aa)

MAADGYLPDWLEDNLSEGIREWWALKPGAPQPKANQQHQDNARGLVLPGYKYLG PGNGLDKGEPVNAADAAALEHDKAYDQQLKAGDNPYLKYNHADAEFQERLKEDTS FGGNLGRAVFQAKKRLLEPLGLVEEAAKTAPGKKRPVEQSPQEPDSSAGIGKSGAQP AKKRLNFGQTGDTESVPDPQPIGEPPAAPSGVGSLTMASGGGAPVADNNEGADGVG SSSGNWHCDSQWLGDRVITTSTRTWALPTYNNHLYKQISNSTSGGSSNDNAYFGYST PWGYFDFNRFHCHFSPRDWQRLINNNWGFRPKRLNFKLFNIQVKEVTDNNGVKTIA NNLTSTVQVFTDSDYQLPYVLGSAHEGCLPPFPADVFMIPQYGYLTLNDGSQAVGRS SF YCLEYFPSQMLRTGNNFQF S YEFENVPFHS S YAHSQ SLDRLMNPLIDQYL YYLSKT INGSGQNQQTLKF SV AGP SNMAVQGRNYIPGPS YRQQRVSTTVTQNNNSEF AWPGA SSWALNGRNSLMNPGPAMASHKEGEDRFFPLSGSLIFGKQGTGRDNVDADKVMITN EEEIKTTNPVATESYGQVATNHQSAQAQAQTGWVQNQGILPGMVWQDRDVYLQGP IWAKIPHTDGNFHPSPLMGGFGMKHPPPQILIKNTPVPADPPTAFNKDKLNSFITQYST GQVSVEIEWELQKENSKRWNPEIQYTSNYYKSNNVEFAVNTEGVYSEPRPIGTRYLT RNL

[00270] SEQ ID NO:21 - Artificial sequence (Vector 1; 8305 nt) cctgcaggcagctgcgcgctcgctcgctcactgaggccgcccgggcaaagcccgggcgtc gggcgacctttggtcgcccggcctc agtgagcgagcgagcgcgcagagagggagtggccaactccatcactaggggttcctgcgg ccgcacgcgtgttactaggttctaga accggtgacgtctcccatggtgaagcttggatctgagatatcaaactgcctgtttgggct tctcatttcttacctccccttccctctcccacc tgctactgggtgcatctctgctccccccttccccagcagatggttacctttgggctgttg ctttcttgtcaccatctgagttctcagacgctg gaaagccatgttctcggctctgtgaatgacaatgctgactggagtgctgcccctctgtaa agggctgggtgtggatggtcacaagcccc tcacatgcctcagccaagaggaagtagtacaggggtcagcccagaggtccaggggaaagg agtggaaaccgatttccccaccaag ggaggggcctgtacctcagctgttcccatagcttacttgccacaactgccaagcaagttt cgctgagtttgacacatggatccctgtgga tcaactgccctaggactccgtttgcacccatgtgacactgttgactttgccctgacgaag cagggccaacagtcccctaacttaattaca aaaactaatgactaagagagaggtggctagagctgaggcccctgagtcaggctgtgggtg ggatcatctccagtacaggaagtgag actttcatttcctcctttccaagagagggctgagggagcagggttgagcaactggtgcag acagcctagctggactttgggtgaggcg gttcagccatatcgaattctgctggggctactggcaggtaaggaggaaggaggctgaggg gagggggcccctgggagggagcctg ccctgggttgctaaccatctcctctctgccaaaagtccggaaagccaccatggagcccct gcgcctgctgatcctgctgttcgtgaccg agctgagcggcgcccacaacaccaccgtgttccagggcgtggccggccagagcctgcagg tgagctgcccctacgacagcatgaa gcactggggccgccgcaaggcctggtgccgccagctgggcgagaagggcccctgccagcg cgtggtgagcacccacaacctgtg gctgctgagcttcctgcgccgctggaacggcagcaccgccatcaccgacgacaccctggg cggcaccctgaccatcaccctgcgc aacctgcagccccacgacgccggcctgtaccagtgccagagcctgcacggcagcgaggcc gacaccctgcgcaaggtgctggtg gaggtgctggccgaccccctggaccaccgcgacgccggcgacctgtggttccccggcgag agcgagagcttcgaggacgcccac gtggagcacagcatcagccgcagcctgctggagggcgagatccccttcccccccaccagc atcctgctgctgctggcctgcatcttc ctgatcaagatcctggccgccagcgccctgtgggccgccgcctggcacggccagaagccc ggcacccacccccccagcgagctg gactgcggccacgaccccggctaccagctgcagaccctgcccggcctgcgcgacacctga caattgttaattaagtttaaaccctcga ggccgcaagcttatcgataatcaacctctggattacaaaatttgtgaaagattgactggt attcttaactatgttgctccttttacgctatgtg gatacgctgctttaatgcctttgtatcatgctattgcttcccgtatggctttcattttct cctccttgtataaatcctggttgctgtctctttatgag gagttgtggcccgttgtcaggcaacgtggcgtggtgtgcactgtgtttgctgacgcaacc cccactggttggggcattgccaccacctg tcagctcctttccgggactttcgctttccccctccctattgccacggcggaactcatcgc cgcctgccttgcccgctgctggacaggggc tcggctgttgggcactgacaattccgtggtgttgtcggggaaatcatcgtcctttccttg gctgctcgcctgtgttgccacctggattctgc gcgggacgtccttctgctacgtcccttcggccctcaatccagcggaccttccttcccgcg gcctgctgccggctctgcggcctcttccg cgtcttcgccttcgccctcagacgagtcggatctccctttgggccgcctccccgcatcga taccgtcgactagagctcgctgatcagcc tcgactgtgccttctagttgccagccatctgttgtttgcccctcccccgtgccttccttg accctggaaggtgccactcccactgtcctttcc taataaaatgaggaaattgcatcgcattgtctgagtaggtgtcattctattctggggggt ggggtggggcaggacagcaagggggagg attgggaagacaatagcaggcatgctggggagagatccacgataacaaacagcttttttg gggtgaacatattgactgaattcccgtgc ggaccggcagtcctcacttacctgagtgccctgttctgcactcctctcctccccaccctg cccatattcttcacaccctccttccctcccca gtatcaaaacctaccctcagactctttctgctgtgagactgataaggggctggtggactc tggttgcattggaagggaaggagtgcttac agtggcatcccaggcactagctggcagttgggggaagctttggggggcactggcactggt aatagcctctgaaattataagcctctcat ttgaagcccctacagttataaaggaggaaagaacctgaggtgattcatctgcatctttgg ggcactctctcccctgctctgaaggtcccct tgtcctcagctcttgtgaggaggtcaggagcatagagatttccattgctgagttgagggg aacttacacaccaaatttttgtacacaattg cttcttcccactgaggaaagcacaccttttgctcccccagagcactggacaaacctgggc aagaggagagacagtgcctgaaagagt cttgttcctgcagcttcttagtgagtggtcagggggaaggcagggttctgaggctgagtg aggaggtggtcagtgaggagagggcca cagcccatctggtctctctagttccaaaaggcaagccaccaacttcccaatcctatcttt caagcctctgtcacagttgaagttgaagtga gcaggcaagggaggaccaagagagaagtcaaagcagggggctctgggggctgccccagca acaggctggtgctctaagcccatc tcccccaccctaaggagggttcccaaaatagcagcctctcatctcccccaaaatatctcc aagacaggtccttcctgaaaggggaaca aagccacagaaatagggaagctggaagttaaaggtcaggaaagtcagtacctgtgaaggc agctgcagagcaagcaagagtggca gggcagggagagccagccccaggccaagaggagaggtcccagtccctgtgaaaagcaaag agtgtgacagaacagaactggag ggggttttaagaccaagtgcctccagaatagacccagagaaggtcagttacttctccaag gaggcccagcaagagcaacagtgaga aaaccaaacccaggtcccaggtttcctggttcccaatttcccacaaacacttactgtgcc atccactcccaacttgtataagaacctaagg cggaccgagcggccgcaggaacccctagtgatggagttggccactccctctctgcgcgct cgctcgctcactgaggccgggcgacc aaaggtcgcccgacgcccgggctttgcccgggcggcctcagtgagcgagcgagcgcgcag ctgcctgcaggcatgcaagctgta gccaaccactagaactatagctagagtcctgggcgaacaaacgatgctcgccttccagaa aaccgaggatgcgaaccacttcatccg gggtcagcaccaccggcaagcgccgcgacggccgaggtcttccgatctcctgaagccagg gcagatccgtgcacagcaccttgcc gtagaagaacagcaaggccgccaatgcctgacgatgcgtggagaccgaaaccttgcgctc gttcgccagccaggacagaaatgcct cgacttcgctgctgcccaaggttgccgggtgacgcacaccgtggaaacggatgaaggcac gaacccagttgacataagcctgttcg gttcgtaaactgtaatgcaagtagcgtatgcgctcacgcaactggtccagaaccttgacc gaacgcagcggtggtaacggcgcagtg gcggttttcatggcttgttatgactgtttttttgtacagtctatgcctcgggcatccaag cagcaagcgcgttacgccgtgggtcgatgtttg atgttatggagcagcaacgatgttacgcagcagcaacgatgttacgcagcagggcagtcg ccctaaaacaaagttaggtggctcaag tatgggcatcattcgcacatgtaggctcggacctgaccaagtcaaatccatgcgggctgc tcttgatcttttcggtcgtgagttcggaga cgtagccacctactcccaacatcagccggactccgattacctcgggaacttgctccgtag taagacattcatcgcgcttgctgccttcga ccaagaagcggttgttggcgctctcgcggcttacgttctgcccaggtttgagcagccgcg tagtgagatctatatctatgatctcgcagt ctccggcgagcaccggaggcagggcattgccaccgcgctcatcaatctcctcaagcatga ggccaacgcgcttggtgcttatgtgat ctacgtgcaagcagattacggtgacgatcccgcagtggctctctatacaaagttgggcat acgggaagaagtgatgcactttgatatcg acccaagtaccgccacctaacaattcgttcaagccgagatcggcttcccggccgcggagt tgttcggtaaattgtcacaacgccgcga atatagtctttaccatgcccttggccacgcccctctttaatacgacgggcaatttgcact tcagaaaatgaagagtttgctttagccataac aaaagtccagtatgctttttcacagcataactggactgatttcagtttacaactattctg tctagtttaagactttattgtcatagtttagatctat tttgttcagtttaagactttattgtccgcccacacccgcttacgcagggcatccatttat tactcaaccgtaaccgattttgccaggttacgc ggctggtctgcggtgtgaaataccgcacagatgcgtaaggagaaaataccgcatcaggcg ctcttccgcttcctcgctcactgactcg ctgcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaatacgg ttatccacagaatcaggggataacgca ggaaagaacatgtgagcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttg ctggcgtttttccataggctccgccc ccctgacgagcatcacaaaaatcgacgctcaagtcagaggtggcgaaacccgacaggact ataaagataccaggcgtttccccctg gaagctccctcgtgcgctctcctgttccgaccctgccgcttaccggatacctgtccgcct ttctcccttcgggaagcgtggcgctttctca atgctcacgctgtaggtatctcagttcggtgtaggtcgttcgctccaagctgggctgtgt gcacgaaccccccgttcagcccgaccgct gcgccttatccggtaactatcgtcttgagtccaacccggtaagacacgacttatcgccac tggcagcagccactggtaacaggattagc agagcgaggtatgtaggcggtgctacagagttcttgaagtggtggcctaactacggctac actagaaggacagtatttggtatctgcgc tctgctgaagccagttaccttcggaaaaagagttggtagctcttgatccggcaaacaaac caccgctggtagcggtggtttttttgtttgc aagcagcagattacgcgcagaaaaaaaggatctcaagaagatcctttgatcttttctacg gggtctgacgctcagtggaacgaaaactc acgttaagggattttggtcatgagattatcaaaaaggatcttcacctagatccttttaaa ttaaaaatgaagttttaaatcaatctaaagtatat atgagtaaacttggtctgacagttaccaatgcttaatcagtgaggcacctatctcagcga tctgtctatttcgttcatccatagttgcctgact ccccgtcgtgtagataactacgatacgggagggcttaccatctggccccagtgctgcaat gataccgcgagacccacgctcaccggc tccagatttatcagcaataaaccagccagccggaagggccgagcgcagaagtggtcctgc aactttatccgcctccatccagtctatta attgttgccgggaagctagagtaagtagttcgccagttaatagtttgcgcaacgttgttg ccattgctacaggcatcgtggtgtcacgctc gtcgtttggtatggcttcattcagctccggttcccaacgatcaaggcgagttacatgatc ccccatgttgtgcaaaaaagcggttagctcc ttcggtcctccgatcgttgtcagaagtaagttggccgcagtgttatcactcatggttatg gcagcactgcataattctcttactgtcatgcca tccgtaagatgcttttctgtgactggtgagtactcaaccaagtcattctgagaatagtgt atgcggcgaccgagttgctcttgcccggcgt caatacgggataataccgcgccacatagcagaactttaaaagtgctcatcattggaaaac gttcttcggggcgaaaactctcaaggatc ttaccgctgttgagatccagttcgatgtaacccactcgtgcacccaactgatcttcagca tcttttactttcaccagcgtttctgggtgagca aaaacaggaaggcaaaatgccgcaaaaaagggaataagggcgacacggaaatgttgaata ctcatactcttcctttttcaatattattga agcatttatcagggttattgtctcatgagcggatacatatttgaatgtatttagaaaaat aaacaaataggggttccgcgcacatttccccg aaaagtgccacctgaaattgtaaacgttaatattttgttaaaattcgcgttaaatttttg ttaaatcagctcattttttaaccaataggccgaaat cggcaaaatcccttataaatcaaaagaatagaccgagatagggttgagtgttgttccagt ttggaacaagagtccactattaaagaacgt ggactccaacgtcaaagggcgaaaaaccgtctatcagggcgatggcccactacgtgaacc atcaccctaatcaagttttttggggtcg aggtgccgtaaagcactaaatcggaaccctaaagggagcccccgatttagagcttgacgg ggaaagccggcgaacgtggcgagaa aggaagggaagaaagcgaaaggagcgggcgctagggcgctggcaagtgtagcggtcacgc tgcgcgtaaccaccacacccgcc gcgcttaatgcgccgctacagggcgcgtccattcgccattcaggctgcaaataagcgttg atattcagtcaattacaaacattaataacg aagagatgacagaaaaattttcattctgtgacagagaaaaagtagccgaagatgacggtt tgtcacatggagttggcaggatgtttgatt aaaaacataacaggaagaaaaatgccccgctgtgggcggacaaaatagttgggaactggg aggggtggaaatggagtttttaaggat tatttagggaagagtgacaaaatagatgggaactgggtgtagcgtcgtaagctaatacga aaattaaaaatgacaaaatagtttggaact agatttcacttatctggttcggatctcctagagcttacagctt

[00271] SEQ ID NO:22 - Artificial sequence (Vector 2; 8305 nt) cattcgccattcaggctgcaaataagcgttgatattcagtcaattacaaacattaataac gaagagatgacagaaaaattttcattctgtga cagagaaaaagtagccgaagatgacggtttgtcacatggagttggcaggatgtttgatta aaaacataacaggaagaaaaatgccccg ctgtgggcggacaaaatagttgggaactgggaggggtggaaatggagtttttaaggatta tttagggaagagtgacaaaatagatggg aactgggtgtagcgtcgtaagctaatacgaaaattaaaaatgacaaaatagtttggaact agatttcacttatctggttcggatctcctaga gcttacagcttcctgcaggcagctgcgcgctcgctcgctcactgaggccgcccgggcaaa gcccgggcgtcgggcgacctttggtc gcccggcctcagtgagcgagcgagcgcgcagagagggagtggccaactccatcactaggg gttcctgcggccgcacgcgtgttac taggttctagaaccggtgacgtctcccatggtgaagcttggatctgagatatcaaactgc ctgtttgggcttctcatttcttacctccccttc cctctcccacctgctactgggtgcatctctgctccccccttccccagcagatggttacct ttgggctgttgctttcttgtcaccatctgagttc tcagacgctggaaagccatgttctcggctctgtgaatgacaatgctgactggagtgctgc ccctctgtaaagggctgggtgtggatggt cacaagcccctcacatgcctcagccaagaggaagtagtacaggggtcagcccagaggtcc aggggaaaggagtggaaaccgattt ccccaccaagggaggggcctgtacctcagctgttcccatagcttacttgccacaactgcc aagcaagtttcgctgagtttgacacatgg atccctgtggatcaactgccctaggactccgtttgcacccatgtgacactgttgactttg ccctgacgaagcagggccaacagtccccta acttaattacaaaaactaatgactaagagagaggtggctagagctgaggcccctgagtca ggctgtgggtgggatcatctccagtaca ggaagtgagactttcatttcctcctttccaagagagggctgagggagcagggttgagcaa ctggtgcagacagcctagctggactttg ggtgaggcggttcagccatatcgaattctgctggggctactggcaggtaaggaggaagga ggctgaggggagggggcccctggga gggagcctgccctgggttgctaaccatctcctctctgccaaaagtccggaaagccaccat ggagcccctgaggctgctcatcctgctg tttgtgacagaactgtctggagcccacaacaccacagtgttccagggagttgctggccag tctctgcaagtgtcttgcccctatgacagc atgaagcactggggaaggaggaaggcttggtgtaggcagctgggagagaaaggaccttgc cagagggtggtgagcacacacaac ctgtggctgctgagcttcctcagaaggtggaatggctctacagccatcacagatgacacc ctgggtggcaccctcaccatcaccttaag gaacctgcagcctcatgatgctggcctgtaccaatgccagagcctgcatggctctgaggc tgataccctcaggaaggtgttggtggag gtgctggctgatcctctggatcacagggatgctggagacctgtggttcccaggagagtct gagagctttgaggatgcccatgtggagc acagcatcagcaggtctcttctggagggagagatccccttccctcccacaagcatcctgt tgctgcttgcctgcatcttcctgatcaagat ccttgctgcttctgctctttgggctgctgcctggcatggccagaaacctggaacacatcc tccctctgaactggactgtggccatgaccc tggctaccagttgcaaaccttgcctggcttgagggacacctgacaattgttaattaagtt taaaccctcgaggccgcaagcttatcgataa tcaacctctggattacaaaatttgtgaaagattgactggtattcttaactatgttgctcc ttttacgctatgtggatacgctgctttaatgccttt gtatcatgctattgcttcccgtatggctttcattttctcctccttgtataaatcctggtt gctgtctctttatgaggagttgtggcccgttgtcag gcaacgtggcgtggtgtgcactgtgtttgctgacgcaacccccactggttggggcattgc caccacctgtcagctcctttccgggacttt cgctttccccctccctattgccacggcggaactcatcgccgcctgccttgcccgctgctg gacaggggctcggctgttgggcactgac aattccgtggtgttgtcggggaaatcatcgtcctttccttggctgctcgcctgtgttgcc acctggattctgcgcgggacgtccttctgcta cgtcccttcggccctcaatccagcggaccttccttcccgcggcctgctgccggctctgcg gcctcttccgcgtcttcgccttcgccctca gacgagtcggatctccctttgggccgcctccccgcatcgataccgtcgactagagctcgc tgatcagcctcgactgtgccttctagttgc cagccatctgttgtttgcccctcccccgtgccttccttgaccctggaaggtgccactccc actgtcctttcctaataaaatgaggaaattgc atcgcattgtctgagtaggtgtcattctattctggggggtggggtggggcaggacagcaa gggggaggattgggaagacaatagcag gcatgctggggagagatccacgataacaaacagcttttttggggtgaacatattgactga attcccgtgcggaccggcagtcctcactt acctgagtgccctgttctgcactcctctcctccccaccctgcccatattcttcacaccct ccttccctccccagtatcaaaacctaccctca gactctttctgctgtgagactgataaggggctggtggactctggttgcattggaagggaa ggagtgcttacagtggcatcccaggcact agctggcagttgggggaagctttggggggcactggcactggtaatagcctctgaaattat aagcctctcatttgaagcccctacagttat aaaggaggaaagaacctgaggtgattcatctgcatctttggggcactctctcccctgctc tgaaggtccccttgtcctcagctcttgtgag gaggtcaggagcatagagatttccattgctgagttgaggggaacttacacaccaaatttt tgtacacaattgcttcttcccactgaggaaa gcacaccttttgctcccccagagcactggacaaacctgggcaagaggagagacagtgcct gaaagagtcttgttcctgcagcttctta gtgagtggtcagggggaaggcagggttctgaggctgagtgaggaggtggtcagtgaggag agggccacagcccatctggtctctct agttccaaaaggcaagccaccaacttcccaatcctatctttcaagcctctgtcacagttg aagttgaagtgagcaggcaagggaggac caagagagaagtcaaagcagggggctctgggggctgccccagcaacaggctggtgctcta agcccatctcccccaccctaaggag ggttcccaaaatagcagcctctcatctcccccaaaatatctccaagacaggtccttcctg aaaggggaacaaagccacagaaataggg aagctggaagttaaaggtcaggaaagtcagtacctgtgaaggcagctgcagagcaagcaa gagtggcagggcagggagagccag ccccaggccaagaggagaggtcccagtccctgtgaaaagcaaagagtgtgacagaacaga actggagggggttttaagaccaagt gcctccagaatagacccagagaaggtcagttacttctccaaggaggcccagcaagagcaa cagtgagaaaaccaaacccaggtcc caggtttcctggttcccaatttcccacaaacacttactgtgccatccactcccaacttgt ataagaacctaaggcggaccgagcggccgc aggaacccctagtgatggagttggccactccctctctgcgcgctcgctcgctcactgagg ccgggcgaccaaaggtcgcccgacgc ccgggctttgcccgggcggcctcagtgagcgagcgagcgcgcagctgcctgcaggcatgc aagctgtagccaaccactagaactat agctagagtcctgggcgaacaaacgatgctcgccttccagaaaaccgaggatgcgaacca cttcatccggggtcagcaccaccggc aagcgccgcgacggccgaggtcttccgatctcctgaagccagggcagatccgtgcacagc accttgccgtagaagaacagcaagg ccgccaatgcctgacgatgcgtggagaccgaaaccttgcgctcgttcgccagccaggaca gaaatgcctcgacttcgctgctgccca aggttgccgggtgacgcacaccgtggaaacggatgaaggcacgaacccagttgacataag cctgttcggttcgtaaactgtaatgca agtagcgtatgcgctcacgcaactggtccagaaccttgaccgaacgcagcggtggtaacg gcgcagtggcggttttcatggcttgttat gactgtttttttgtacagtctatgcctcgggcatccaagcagcaagcgcgttacgccgtg ggtcgatgtttgatgttatggagcagcaacg atgttacgcagcagcaacgatgttacgcagcagggcagtcgccctaaaacaaagttaggt ggctcaagtatgggcatcattcgcacat gtaggctcggccctgaccaagtcaaatccatgcgggctgctcttgatcttttcggtcgtg agttcggagacgtagccacctactcccaac atcagccggactccgattacctcgggaacttgctccgtagtaagacattcatcgcgcttg ctgccttcgaccaagaagcggttgttggc gctctcgcggcttacgttctgcccaggtttgagcagccgcgtagtgagatctatatctat gatctcgcagtctccggcgagcaccggag gcagggcattgccaccgcgctcatcaatctcctcaagcatgaggccaacgcgcttggtgc ttatgtgatctacgtgcaagcagattacg gtgacgatcccgcagtggctctctatacaaagttgggcatacgggaagaagtgatgcact ttgatatcgacccaagtaccgccacctaa caattcgttcaagccgagatcggcttcccggccgcggagttgttcggtaaattgtcacaa cgccgcgaatatagtctttaccatgccctt ggccacgcccctctttaatacgacgggcaatttgcacttcagaaaatgaagagtttgctt tagccataacaaaagtccagtatgctttttca cagcataactggactgatttcagtttacaactattctgtctagtttaagactttattgtc atagtttagatctattttgttcagtttaagactttattg tccgcccacacccgcttacgcagggcatccatttattactcaaccgtaaccgattttgcc aggttacgcggctggtctgcggtgtgaaat accgcacagatgcgtaaggagaaaataccgcatcaggcgctcttccgcttcctcgctcac tgactcgctgcgctcggtcgttcggctg cggcgagcggtatcagctcactcaaaggcggtaatacggttatccacagaatcaggggat aacgcaggaaagaacatgtgagcaaa aggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctggcgtttttccataggct ccgcccccctgacgagcatcacaaaa atcgacgctcaagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttc cccctggaagctccctcgtgcgctctc ctgttccgaccctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgtgg cgctttctcaatgctcacgctgtaggtatctc agttcggtgtaggtcgttcgctccaagctgggctgtgtgcacgaaccccccgttcagccc gaccgctgcgccttatccggtaactatcg tcttgagtccaacccggtaagacacgacttatcgccactggcagcagccactggtaacag gattagcagagcgaggtatgtaggcgg tgctacagagttcttgaagtggtggcctaactacggctacactagaaggacagtatttgg tatctgcgctctgctgaagccagttaccttc ggaaaaagagttggtagctcttgatccggcaaacaaaccaccgctggtagcggtggtttt tttgtttgcaagcagcagattacgcgcag aaaaaaaggatctcaagaagatcctttgatcttttctacggggtctgacgctcagtggaa cgaaaactcacgttaagggattttggtcatg agattatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaatca atctaaagtatatatgagtaaacttggtctgaca gttaccaatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgttcatcca tagttgcctgactccccgtcgtgtagataacta cgatacgggagggcttaccatctggccccagtgctgcaatgataccgcgagacccacgct caccggctccagatttatcagcaataaa ccagccagccggaagggccgagcgcagaagtggtcctgcaactttatccgcctccatcca gtctattaattgttgccgggaagctaga gtaagtagttcgccagttaatagtttgcgcaacgttgttgccattgctacaggcatcgtg gtgtcacgctcgtcgtttggtatggcttcattc agctccggttcccaacgatcaaggcgagttacatgatcccccatgttgtgcaaaaaagcg gttagctccttcggtcctccgatcgttgtc agaagtaagttggccgcagtgttatcactcatggttatggcagcactgcataattctctt actgtcatgccatccgtaagatgcttttctgtg actggtgagtactcaaccaagtcattctgagaatagtgtatgcggcgaccgagttgctct tgcccggcgtcaatacgggataataccgc gccacatagcagaactttaaaagtgctcatcattggaaaacgttcttcggggcgaaaact ctcaaggatcttaccgctgttgagatccag ttcgatgtaacccactcgtgcacccaactgatcttcagcatcttttactttcaccagcgt ttctgggtgagcaaaaacaggaaggcaaaat gccgcaaaaaagggaataagggcgacacggaaatgttgaatactcatactcttccttttt caatattattgaagcatttatcagggttattgt ctcatgagcggatacatatttgaatgtatttagaaaaataaacaaataggggttccgcgc acatttccccgaaaagtgccacctgaaattg taaacgttaatattttgttaaaattcgcgttaaatttttgttaaatcagctcatttttta accaataggccgaaatcggcaaaatcccttataaat caaaagaatagaccgagatagggttgagtgttgttccagtttggaacaagagtccactat taaagaacgtggactccaacgtcaaagg gcgaaaaaccgtctatcagggcgatggcccactacgtgaaccatcaccctaatcaagttt tttggggtcgaggtgccgtaaagcactaa atcggaaccctaaagggagcccccgatttagagcttgacggggaaagccggcgaacgtgg cgagaaaggaagggaagaaagcg aaaggagcgggcgctagggcgctggcaagtgtagcggtcacgctgcgcgtaaccaccaca cccgccgcgcttaatgcgccgctac agggcgcgtc

[00272] SEQ ID NO:23 - Artificial sequence (Vector 3; 8305 nt) cattcgccattcaggctgcaaataagcgttgatattcagtcaattacaaacattaataac gaagagatgacagaaaaattttcattctgtga cagagaaaaagtagccgaagatgacggtttgtcacatggagttggcaggatgtttgatta aaaacataacaggaagaaaaatgccccg ctgtgggcggacaaaatagttgggaactgggaggggtggaaatggagtttttaaggatta tttagggaagagtgacaaaatagatggg aactgggtgtagcgtcgtaagctaatacgaaaattaaaaatgacaaaatagtttggaact agatttcacttatctggttcggatctcctaga gcttacagcttcctgcaggcagctgcgcgctcgctcgctcactgaggccgcccgggcaaa gcccgggcgtcgggcgacctttggtc gcccggcctcagtgagcgagcgagcgcgcagagagggagtggccaactccatcactaggg gttcctgcggccgcacgcgtgttac taggttctagaaccggtgacgtctcccatggtgaagcttggatctgagatatcaaactgc ctgtttgggcttctcatttcttacctccccttc cctctcccacctgctactgggtgcatctctgctccccccttccccagcagatggttacct ttgggctgttgctttcttgtcaccatctgagttc tcagacgctggaaagccatgttctcggctctgtgaatgacaatgctgactggagtgctgc ccctctgtaaagggctgggtgtggatggt cacaagcccctcacatgcctcagccaagaggaagtagtacaggggtcagcccagaggtcc aggggaaaggagtggaaaccgattt ccccaccaagggaggggcctgtacctcagctgttcccatagcttacttgccacaactgcc aagcaagtttcgctgagtttgacacatgg atccctgtggatcaactgccctaggactccgtttgcacccatgtgacactgttgactttg ccctgacgaagcagggccaacagtccccta acttaattacaaaaactaatgactaagagagaggtggctagagctgaggcccctgagtca ggctgtgggtgggatcatctccagtaca ggaagtgagactttcatttcctcctttccaagagagggctgagggagcagggttgagcaa ctggtgcagacagcctagctggactttg ggtgaggcggttcagccatatcgaattctgctggggctactggcaggtaaggaggaagga ggctgaggggagggggcccctggga gggagcctgccctgggttgctaaccatctcctctctgccaaaagtccggaaagccaccat ggagcccctgaggctgctgatcctgctg tttgtgacagaactgtctggagcccacaacaccacagtgttccagggagttgctggccag tctctgcaagtgtcttgcccctatgacagc atgaagcactggggaaggaggaaggcttggtgtaggcagctgggagagaaaggaccttgc cagagggtggtgagcacacacaac ctgtggctgctgagcttcctcagaaggtggaatggctctacagccatcacagatgacacc ctgggtggcaccctcaccatcaccctga ggaacctgcagcctcatgatgctggcctgtaccaatgccagagcctgcatggctctgagg ctgataccctcaggaaggtgttggtgga ggtgctggctgatcctctggatcacagggatgctggagacctgtggttcccaggagagtc tgagagctttgaggatgcccatgtggag cacagcatcagcaggtctcttctggagggagagatccccttccctcccacaagcatcctg ttgctgcttgcctgcatcttcctcatcaaga tccttgctgcttctgctctttgggctgctgcctggcatggccagaaacctggaacacatc ctccctctgaactggactgtggccatgacc ctggctaccagttgcaaaccttgcctggcttgagggacacctgacaattgttaattaagt ttaaaccctcgaggccgcaagcttatcgata atcaacctctggattacaaaatttgtgaaagattgactggtattcttaactatgttgctc cttttacgctatgtggatacgctgctttaatgcctt tgtatcatgctattgcttcccgtatggctttcattttctcctccttgtataaatcctggt tgctgtctctttatgaggagttgtggcccgttgtcag gcaacgtggcgtggtgtgcactgtgtttgctgacgcaacccccactggttggggcattgc caccacctgtcagctcctttccgggacttt cgctttccccctccctattgccacggcggaactcatcgccgcctgccttgcccgctgctg gacaggggctcggctgttgggcactgac aattccgtggtgttgtcggggaaatcatcgtcctttccttggctgctcgcctgtgttgcc acctggattctgcgcgggacgtccttctgcta cgtcccttcggccctcaatccagcggaccttccttcccgcggcctgctgccggctctgcg gcctcttccgcgtcttcgccttcgccctca gacgagtcggatctccctttgggccgcctccccgcatcgataccgtcgactagagctcgc tgatcagcctcgactgtgccttctagttgc cagccatctgttgtttgcccctcccccgtgccttccttgaccctggaaggtgccactccc actgtcctttcctaataaaatgaggaaattgc atcgcattgtctgagtaggtgtcattctattctggggggtggggtggggcaggacagcaa gggggaggattgggaagacaatagcag gcatgctggggagagatccacgataacaaacagcttttttggggtgaacatattgactga attcccgtgcggaccggcagtcctcactt acctgagtgccctgttctgcactcctctcctccccaccctgcccatattcttcacaccct ccttccctccccagtatcaaaacctaccctca gactctttctgctgtgagactgataaggggctggtggactctggttgcattggaagggaa ggagtgcttacagtggcatcccaggcact agctggcagttgggggaagctttggggggcactggcactggtaatagcctctgaaattat aagcctctcatttgaagcccctacagttat aaaggaggaaagaacctgaggtgattcatctgcatctttggggcactctctcccctgctc tgaaggtccccttgtcctcagctcttgtgag gaggtcaggagcatagagatttccattgctgagttgaggggaacttacacaccaaatttt tgtacacaattgcttcttcccactgaggaaa gcacaccttttgctcccccagagcactggacaaacctgggcaagaggagagacagtgcct gaaagagtcttgttcctgcagcttctta gtgagtggtcagggggaaggcagggttctgaggctgagtgaggaggtggtcagtgaggag agggccacagcccatctggtctctct agttccaaaaggcaagccaccaacttcccaatcctatctttcaagcctctgtcacagttg aagttgaagtgagcaggcaagggaggac caagagagaagtcaaagcagggggctctgggggctgccccagcaacaggctggtgctcta agcccatctcccccaccctaaggag ggttcccaaaatagcagcctctcatctcccccaaaatatctccaagacaggtccttcctg aaaggggaacaaagccacagaaataggg aagctggaagttaaaggtcaggaaagtcagtacctgtgaaggcagctgcagagcaagcaa gagtggcagggcagggagagccag ccccaggccaagaggagaggtcccagtccctgtgaaaagcaaagagtgtgacagaacaga actggagggggttttaagaccaagt gcctccagaatagacccagagaaggtcagttacttctccaaggaggcccagcaagagcaa cagtgagaaaaccaaacccaggtcc caggtttcctggttcccaatttcccacaaacacttactgtgccatccactcccaacttgt ataagaacctaaggcggaccgagcggccgc aggaacccctagtgatggagttggccactccctctctgcgcgctcgctcgctcactgagg ccgggcgaccaaaggtcgcccgacgc ccgggctttgcccgggcggcctcagtgagcgagcgagcgcgcagctgcctgcaggcatgc aagctgtagccaaccactagaactat agctagagtcctgggcgaacaaacgatgctcgccttccagaaaaccgaggatgcgaacca cttcatccggggtcagcaccaccggc aagcgccgcgacggccgaggtcttccgatctcctgaagccagggcagatccgtgcacagc accttgccgtagaagaacagcaagg ccgccaatgcctgacgatgcgtggagaccgaaaccttgcgctcgttcgccagccaggaca gaaatgcctcgacttcgctgctgccca aggttgccgggtgacgcacaccgtggaaacggatgaaggcacgaacccagttgacataag cctgttcggttcgtaaactgtaatgca agtagcgtatgcgctcacgcaactggtccagaaccttgaccgaacgcagcggtggtaacg gcgcagtggcggttttcatggcttgttat gactgtttttttgtacagtctatgcctcgggcatccaagcagcaagcgcgttacgccgtg ggtcgatgtttgatgttatggagcagcaacg atgttacgcagcagcaacgatgttacgcagcagggcagtcgccctaaaacaaagttaggt ggctcaagtatgggcatcattcgcacat gtaggctcggccctgaccaagtcaaatccatgcgggctgctcttgatcttttcggtcgtg agttcggagacgtagccacctactcccaac atcagccggactccgattacctcgggaacttgctccgtagtaagacattcatcgcgcttg ctgccttcgaccaagaagcggttgttggc gctctcgcggcttacgttctgcccaggtttgagcagccgcgtagtgagatctatatctat gatctcgcagtctccggcgagcaccggag gcagggcattgccaccgcgctcatcaatctcctcaagcatgaggccaacgcgcttggtgc ttatgtgatctacgtgcaagcagattacg gtgacgatcccgcagtggctctctatacaaagttgggcatacgggaagaagtgatgcact ttgatatcgacccaagtaccgccacctaa caattcgttcaagccgagatcggcttcccggccgcggagttgttcggtaaattgtcacaa cgccgcgaatatagtctttaccatgccctt ggccacgcccctctttaatacgacgggcaatttgcacttcagaaaatgaagagtttgctt tagccataacaaaagtccagtatgctttttca cagcataactggactgatttcagtttacaactattctgtctagtttaagactttattgtc atagtttagatctattttgttcagtttaagactttattg tccgcccacacccgcttacgcagggcatccatttattactcaaccgtaaccgattttgcc aggttacgcggctggtctgcggtgtgaaat accgcacagatgcgtaaggagaaaataccgcatcaggcgctcttccgcttcctcgctcac tgactcgctgcgctcggtcgttcggctg cggcgagcggtatcagctcactcaaaggcggtaatacggttatccacagaatcaggggat aacgcaggaaagaacatgtgagcaaa aggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctggcgtttttccataggct ccgcccccctgacgagcatcacaaaa atcgacgctcaagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttc cccctggaagctccctcgtgcgctctc ctgttccgaccctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgtgg cgctttctcaatgctcacgctgtaggtatctc agttcggtgtaggtcgttcgctccaagctgggctgtgtgcacgaaccccccgttcagccc gaccgctgcgccttatccggtaactatcg tcttgagtccaacccggtaagacacgacttatcgccactggcagcagccactggtaacag gattagcagagcgaggtatgtaggcgg tgctacagagttcttgaagtggtggcctaactacggctacactagaaggacagtatttgg tatctgcgctctgctgaagccagttaccttc ggaaaaagagttggtagctcttgatccggcaaacaaaccaccgctggtagcggtggtttt tttgtttgcaagcagcagattacgcgcag aaaaaaaggatctcaagaagatcctttgatcttttctacggggtctgacgctcagtggaa cgaaaactcacgttaagggattttggtcatg agattatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaatca atctaaagtatatatgagtaaacttggtctgaca gttaccaatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgttcatcca tagttgcctgactccccgtcgtgtagataacta cgatacgggagggcttaccatctggccccagtgctgcaatgataccgcgagacccacgct caccggctccagatttatcagcaataaa ccagccagccggaagggccgagcgcagaagtggtcctgcaactttatccgcctccatcca gtctattaattgttgccgggaagctaga gtaagtagttcgccagttaatagtttgcgcaacgttgttgccattgctacaggcatcgtg gtgtcacgctcgtcgtttggtatggcttcattc agctccggttcccaacgatcaaggcgagttacatgatcccccatgttgtgcaaaaaagcg gttagctccttcggtcctccgatcgttgtc agaagtaagttggccgcagtgttatcactcatggttatggcagcactgcataattctctt actgtcatgccatccgtaagatgcttttctgtg actggtgagtactcaaccaagtcattctgagaatagtgtatgcggcgaccgagttgctct tgcccggcgtcaatacgggataataccgc gccacatagcagaactttaaaagtgctcatcattggaaaacgttcttcggggcgaaaact ctcaaggatcttaccgctgttgagatccag ttcgatgtaacccactcgtgcacccaactgatcttcagcatcttttactttcaccagcgt ttctgggtgagcaaaaacaggaaggcaaaat gccgcaaaaaagggaataagggcgacacggaaatgttgaatactcatactcttccttttt caatattattgaagcatttatcagggttattgt ctcatgagcggatacatatttgaatgtatttagaaaaataaacaaataggggttccgcgc acatttccccgaaaagtgccacctgaaattg taaacgttaatattttgttaaaattcgcgttaaatttttgttaaatcagctcatttttta accaataggccgaaatcggcaaaatcccttataaat caaaagaatagaccgagatagggttgagtgttgttccagtttggaacaagagtccactat taaagaacgtggactccaacgtcaaagg gcgaaaaaccgtctatcagggcgatggcccactacgtgaaccatcaccctaatcaagttt tttggggtcgaggtgccgtaaagcactaa atcggaaccctaaagggagcccccgatttagagcttgacggggaaagccggcgaacgtgg cgagaaaggaagggaagaaagcg aaaggagcgggcgctagggcgctggcaagtgtagcggtcacgctgcgcgtaaccaccaca cccgccgcgcttaatgcgccgctac agggcgcgtc

[00273] SEQ ID NO:24 - Artificial sequence (Vector 4; 8305 nt) cctgcaggcagctgcgcgctcgctcgctcactgaggccgcccgggcaaagcccgggcgtc gggcgacctttggtcgcccggcctc agtgagcgagcgagcgcgcagagagggagtggccaactccatcactaggggttcctgcgg ccgcacgcgtgttactaggttctaga accggtgacgtctcccatggtgaagcttggatctgagatatcaaactgcctgtttgggct tctcatttcttacctccccttccctctcccacc tgctactgggtgcatctctgctccccccttccccagcagatggttacctttgggctgttg ctttcttgtcaccatctgagttctcagacgctg gaaagccatgttctcggctctgtgaatgacaatgctgactggagtgctgcccctctgtaa agggctgggtgtggatggtcacaagcccc tcacatgcctcagccaagaggaagtagtacaggggtcagcccagaggtccaggggaaagg agtggaaaccgatttccccaccaag ggaggggcctgtacctcagctgttcccatagcttacttgccacaactgccaagcaagttt cgctgagtttgacacatggatccctgtgga tcaactgccctaggactccgtttgcacccatgtgacactgttgactttgccctgacgaag cagggccaacagtcccctaacttaattaca aaaactaatgactaagagagaggtggctagagctgaggcccctgagtcaggctgtgggtg ggatcatctccagtacaggaagtgag actttcatttcctcctttccaagagagggctgagggagcagggttgagcaactggtgcag acagcctagctggactttgggtgaggcg gttcagccatatcgaattctgctggggctactggcaggtaaggaggaaggaggctgaggg gagggggcccctgggagggagcctg ccctgggttgctaaccatctcctctctgccaaaagtccggaaagccaccatggagcccct gagactgctcatcctgctgtttgtgacaga actgtctggggcccacaacaccacagtgttccagggggtggctggccagtccctccaggt gtcctgcccctatgactccatgaagcac tgggggagaagaaaggcttggtgtagacagctgggggagaaagggccttgccagagagtg gtgtccacacacaacctgtggctgct gtccttcctgagaagatggaatggctccacagccatcacagatgacaccctggggggcac cctcaccatcaccctgagaaacctgca gcctcatgatgctggcctgtaccagtgccagtccctgcatggctctgaggctgataccct gagaaaggtgctggtggaggtgctggct gatcctctggatcacagagatgctggggacctgtggttcccaggggagtctgagtccttt gaggatgcccatgtggagcactccatctc cagatccctgctggagggggagatccccttccctcccacatccatcctgctgctgctggc ctgcatcttcctcatcaagatcctggctgc ttctgctctgtgggctgctgcctggcatggccagaaacctgggacacatcctccctctga actggactgtggccatgaccctggctacc agctgcagaccctgcctggcctgagagacacctgacaattgttaattaagtttaaaccct cgaggccgcaagcttatcgataatcaacct ctggattacaaaatttgtgaaagattgactggtattcttaactatgttgctccttttacg ctatgtggatacgctgctttaatgcctttgtatcat gctattgcttcccgtatggctttcattttctcctccttgtataaatcctggttgctgtct ctttatgaggagttgtggcccgttgtcaggcaacgt ggcgtggtgtgcactgtgtttgctgacgcaacccccactggttggggcattgccaccacc tgtcagctcctttccgggactttcgctttcc ccctccctattgccacggcggaactcatcgccgcctgccttgcccgctgctggacagggg ctcggctgttgggcactgacaattccgt ggtgttgtcggggaaatcatcgtcctttccttggctgctcgcctgtgttgccacctggat tctgcgcgggacgtccttctgctacgtccctt cggccctcaatccagcggaccttccttcccgcggcctgctgccggctctgcggcctcttc cgcgtcttcgccttcgccctcagacgagt cggatctccctttgggccgcctccccgcatcgataccgtcgactagagctcgctgatcag cctcgactgtgccttctagttgccagccat ctgttgtttgcccctcccccgtgccttccttgaccctggaaggtgccactcccactgtcc tttcctaataaaatgaggaaattgcatcgcatt gtctgagtaggtgtcattctattctggggggtggggtggggcaggacagcaagggggagg attgggaagacaatagcaggcatgct ggggagagatccacgataacaaacagcttttttggggtgaacatattgactgaattcccg tgcggaccggcagtcctcacttacctgag tgccctgttctgcactcctctcctccccaccctgcccatattcttcacaccctccttccc tccccagtatcaaaacctaccctcagactctttc tgctgtgagactgataaggggctggtggactctggttgcattggaagggaaggagtgctt acagtggcatcccaggcactagctggca gttgggggaagctttggggggcactggcactggtaatagcctctgaaattataagcctct catttgaagcccctacagttataaaggag gaaagaacctgaggtgattcatctgcatctttggggcactctctcccctgctctgaaggt ccccttgtcctcagctcttgtgaggaggtca ggagcatagagatttccattgctgagttgaggggaacttacacaccaaatttttgtacac aattgcttcttcccactgaggaaagcacacc ttttgctcccccagagcactggacaaacctgggcaagaggagagacagtgcctgaaagag tcttgttcctgcagcttcttagtgagtgg tcagggggaaggcagggttctgaggctgagtgaggaggtggtcagtgaggagagggccac agcccatctggtctctctagttccaa aaggcaagccaccaacttcccaatcctatctttcaagcctctgtcacagttgaagttgaa gtgagcaggcaagggaggaccaagaga gaagtcaaagcagggggctctgggggctgccccagcaacaggctggtgctctaagcccat ctcccccaccctaaggagggttccca aaatagcagcctctcatctcccccaaaatatctccaagacaggtccttcctgaaagggga acaaagccacagaaatagggaagctgg aagttaaaggtcaggaaagtcagtacctgtgaaggcagctgcagagcaagcaagagtggc agggcagggagagccagccccagg ccaagaggagaggtcccagtccctgtgaaaagcaaagagtgtgacagaacagaactggag ggggttttaagaccaagtgcctccag aatagacccagagaaggtcagttacttctccaaggaggcccagcaagagcaacagtgaga aaaccaaacccaggtcccaggtttcct ggttcccaatttcccacaaacacttactgtgccatccactcccaacttgtataagaacct aaggcggaccgagcggccgcaggaaccc ctagtgatggagttggccactccctctctgcgcgctcgctcgctcactgaggccgggcga ccaaaggtcgcccgacgcccgggcttt gcccgggcggcctcagtgagcgagcgagcgcgcagctgcctgcaggcatgcaagctgtag ccaaccactagaactatagctagag tcctgggcgaacaaacgatgctcgccttccagaaaaccgaggatgcgaaccacttcatcc ggggtcagcaccaccggcaagcgcc gcgacggccgaggtcttccgatctcctgaagccagggcagatccgtgcacagcaccttgc cgtagaagaacagcaaggccgccaat gcctgacgatgcgtggagaccgaaaccttgcgctcgttcgccagccaggacagaaatgcc tcgacttcgctgctgcccaaggttgcc gggtgacgcacaccgtggaaacggatgaaggcacgaacccagttgacataagcctgttcg gttcgtaaactgtaatgcaagtagcgt atgcgctcacgcaactggtccagaaccttgaccgaacgcagcggtggtaacggcgcagtg gcggttttcatggcttgttatgactgtttt tttgtacagtctatgcctcgggcatccaagcagcaagcgcgttacgccgtgggtcgatgt ttgatgttatggagcagcaacgatgttacg cagcagcaacgatgttacgcagcagggcagtcgccctaaaacaaagttaggtggctcaag tatgggcatcattcgcacatgtaggctc ggacctgaccaagtcaaatccatgcgggctgctcttgatcttttcggtcgtgagttcgga gacgtagccacctactcccaacatcagcc ggactccgattacctcgggaacttgctccgtagtaagacattcatcgcgcttgctgcctt cgaccaagaagcggttgttggcgctctcgc ggcttacgttctgcccaggtttgagcagccgcgtagtgagatctatatctatgatctcgc agtctccggcgagcaccggaggcagggc attgccaccgcgctcatcaatctcctcaagcatgaggccaacgcgcttggtgcttatgtg atctacgtgcaagcagattacggtgacgat cccgcagtggctctctatacaaagttgggcatacgggaagaagtgatgcactttgatatc gacccaagtaccgccacctaacaattcgtt caagccgagatcggcttcccggccgcggagttgttcggtaaattgtcacaacgccgcgaa tatagtctttaccatgcccttggccacgc ccctctttaatacgacgggcaatttgcacttcagaaaatgaagagtttgctttagccata acaaaagtccagtatgctttttcacagcataa ctggactgatttcagtttacaactattctgtctagtttaagactttattgtcatagttta gatctattttgttcagtttaagactttattgtccgccca cacccgcttacgcagggcatccatttattactcaaccgtaaccgattttgccaggttacg cggctggtctgcggtgtgaaataccgcaca gatgcgtaaggagaaaataccgcatcaggcgctcttccgcttcctcgctcactgactcgc tgcgctcggtcgttcggctgcggcgagc ggtatcagctcactcaaaggcggtaatacggttatccacagaatcaggggataacgcagg aaagaacatgtgagcaaaaggccagc aaaaggccaggaaccgtaaaaaggccgcgttgctggcgtttttccataggctccgccccc ctgacgagcatcacaaaaatcgacgct caagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctggaa gctccctcgtgcgctctcctgttccgac cctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgctttctca atgctcacgctgtaggtatctcagttcggtgt aggtcgttcgctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgctgcg ccttatccggtaactatcgtcttgagtcc aacccggtaagacacgacttatcgccactggcagcagccactggtaacaggattagcaga gcgaggtatgtaggcggtgctacaga gttcttgaagtggtggcctaactacggctacactagaaggacagtatttggtatctgcgc tctgctgaagccagttaccttcggaaaaag agttggtagctcttgatccggcaaacaaaccaccgctggtagcggtggtttttttgtttg caagcagcagattacgcgcagaaaaaaag gatctcaagaagatcctttgatcttttctacggggtctgacgctcagtggaacgaaaact cacgttaagggattttggtcatgagattatca aaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaatcaatctaaagt atatatgagtaaacttggtctgacagttaccaat gcttaatcagtgaggcacctatctcagcgatctgtctatttcgttcatccatagttgcct gactccccgtcgtgtagataactacgatacgg gagggcttaccatctggccccagtgctgcaatgataccgcgagacccacgctcaccggct ccagatttatcagcaataaaccagcca gccggaagggccgagcgcagaagtggtcctgcaactttatccgcctccatccagtctatt aattgttgccgggaagctagagtaagta gttcgccagttaatagtttgcgcaacgttgttgccattgctacaggcatcgtggtgtcac gctcgtcgtttggtatggcttcattcagctccg gttcccaacgatcaaggcgagttacatgatcccccatgttgtgcaaaaaagcggttagct ccttcggtcctccgatcgttgtcagaagta agttggccgcagtgttatcactcatggttatggcagcactgcataattctcttactgtca tgccatccgtaagatgcttttctgtgactggtg agtactcaaccaagtcattctgagaatagtgtatgcggcgaccgagttgctcttgcccgg cgtcaatacgggataataccgcgccacat agcagaactttaaaagtgctcatcattggaaaacgttcttcggggcgaaaactctcaagg atcttaccgctgttgagatccagttcgatgt aacccactcgtgcacccaactgatcttcagcatcttttactttcaccagcgtttctgggt gagcaaaaacaggaaggcaaaatgccgcaa aaaagggaataagggcgacacggaaatgttgaatactcatactcttcctttttcaatatt attgaagcatttatcagggttattgtctcatga gcggatacatatttgaatgtatttagaaaaataaacaaataggggttccgcgcacatttc cccgaaaagtgccacctgaaattgtaaacgt taatattttgttaaaattcgcgttaaatttttgttaaatcagctcattttttaaccaata ggccgaaatcggcaaaatcccttataaatcaaaaga atagaccgagatagggttgagtgttgttccagtttggaacaagagtccactattaaagaa cgtggactccaacgtcaaagggcgaaaa accgtctatcagggcgatggcccactacgtgaaccatcaccctaatcaagttttttgggg tcgaggtgccgtaaagcactaaatcggaa ccctaaagggagcccccgatttagagcttgacggggaaagccggcgaacgtggcgagaaa ggaagggaagaaagcgaaaggag cgggcgctagggcgctggcaagtgtagcggtcacgctgcgcgtaaccaccacacccgccg cgcttaatgcgccgctacagggcgc gtccattcgccattcaggctgcaaataagcgttgatattcagtcaattacaaacattaat aacgaagagatgacagaaaaattttcattctg tgacagagaaaaagtagccgaagatgacggtttgtcacatggagttggcaggatgtttga ttaaaaacataacaggaagaaaaatgcc ccgctgtgggcggacaaaatagttgggaactgggaggggtggaaatggagtttttaagga ttatttagggaagagtgacaaaatagat gggaactgggtgtagcgtcgtaagctaatacgaaaattaaaaatgacaaaatagtttgga actagatttcacttatctggttcggatctcct agagcttacagctt

[00274] SEQ ID NO:25 - Artificial sequence (Vector 5; 2804 nt) cctgcaggcagctgcgcgctcgctcgctcactgaggccgcccgggcaaagcccgggcgtc gggcgacctttggtcgcccggcctc agtgagcgagcgagcgcgcagagagggagtggccaactccatcactaggggttcctgcgg ccgcacgcgtactaggttctagaac cggtgacgtctcccatggtgaagcttggatctgagatatcaaactgcctgtttgggcttc tcatttcttacctccccttccctctcccacctg ctactgggtgcatctctgctccccccttccccagcagatggttacctttgggctgttgct ttcttgtcaccatctgagttctcagacgctgga aagccatgttctcggctctgtgaatgacaatgctgactggagtgctgcccctctgtaaag ggctgggtgtggatggtcacaagcccctc acatgcctcagccaagaggaagtagtacaggggtcagcccagaggtccaggggaaaggag tggaaaccgatttccccaccaagg gaggggcctgtacctcagctgttcccatagcttacttgccacaactgccaagcaagtttc gctgagtttgacacatggatccctgtggat caactgccctaggactccgtttgcacccatgtgacactgttgactttgccctgacgaagc agggccaacagtcccctaacttaattacaa aaactaatgactaagagagaggtggctagagctgaggcccctgagtcaggctgtgggtgg gatcatctccagtacaggaagtgaga ctttcatttcctcctttccaagagagggctgagggagcagggttgagcaactggtgcaga cagcctagctggactttgggtgaggcggt tcagccatatcgaattctgctggggctactggcaggtaaggaggaaggaggctgagggga gggggcccctgggagggagcctgc cctgggttgctaaccatctcctctctgccaaaagtccggaaagccaccatggagcccctg cgcctgctgatcctgctgttcgtgaccga gctgagcggcgcccacaacaccaccgtgttccagggcgtggccggccagagcctgcaggt gagctgcccctacgacagcatgaa gcactggggccgccgcaaggcctggtgccgccagctgggcgagaagggcccctgccagcg cgtggtgagcacccacaacctgtg gctgctgagcttcctgcgccgctggaacggcagcaccgccatcaccgacgacaccctggg cggcaccctgaccatcaccctgcgc aacctgcagccccacgacgccggcctgtaccagtgccagagcctgcacggcagcgaggcc gacaccctgcgcaaggtgctggtg gaggtgctggccgaccccctggaccaccgcgacgccggcgacctgtggttccccggcgag agcgagagcttcgaggacgcccac gtggagcacagcatcagccgcagcctgctggagggcgagatccccttcccccccaccagc atcctgctgctgctggcctgcatcttc ctgatcaagatcctggccgccagcgccctgtgggccgccgcctggcacggccagaagccc ggcacccacccccccagcgagctg gactgcggccacgaccccggctaccagctgcagaccctgcccggcctgcgcgacacctga caattgttaattaagtttaaaccctcga ggccgcaagcttatcgataatcaacctctggattacaaaatttgtgaaagattgactggt attcttaactatgttgctccttttacgctatgtg gatacgctgctttaatgcctttgtatcatgctattgcttcccgtatggctttcattttct cctccttgtataaatcctggttgctgtctctttatgag gagttgtggcccgttgtcaggcaacgtggcgtggtgtgcactgtgtttgctgacgcaacc cccactggttggggcattgccaccacctg tcagctcctttccgggactttcgctttccccctccctattgccacggcggaactcatcgc cgcctgccttgcccgctgctggacaggggc tcggctgttgggcactgacaattccgtggtgttgtcggggaaatcatcgtcctttccttg gctgctcgcctgtgttgccacctggattctgc gcgggacgtccttctgctacgtcccttcggccctcaatccagcggaccttccttcccgcg gcctgctgccggctctgcggcctcttccg cgtcttcgccttcgccctcagacgagtcggatctccctttgggccgcctccccgcatcga taccgtcgactagagctcgctgatcagcc tcgactgtgccttctagttgccagccatctgttgtttgcccctcccccgtgccttccttg accctggaaggtgccactcccactgtcctttcc taataaaatgaggaaattgcatcgcattgtctgagtaggtgtcattctattctggggggt ggggtggggcaggacagcaagggggagg attgggaagacaatagcaggcatgctggggagagatccacgataacaaacagcttttttg gggtgaacatattgactgaattcccgtgc ggaccgagcggccgcaggaacccctagtgatggagttggccactccctctctgcgcgctc gctcgctcactgaggccgggcgacca aaggtcgcccgacgcccgggctttgcccgggcggcctcagtgagcgagcgagcgcgcagc tgcctgcagg

[00275] SEQ ID NO:26 - Artificial sequence (Vector 6; 4048 nt) ttggccactccctctctgcgcgctcgctcgctcactgaggccgcccgggcaaagcccggg cgtcgggcgacctttggtcgcccggc ctcagtgagcgagcgagcgcgcagagagggagtggccaactccatcactaggggttcctg cggccgcacgcgtactaggttctaga accggtgacgtctcccatggtgaagcttggatctgagatatcaaactgcctgtttgggct tctcatttcttacctccccttccctctcccacc tgctactgggtgcatctctgctccccccttccccagcagatggttacctttgggctgttg ctttcttgtcaccatctgagttctcagacgctg gaaagccatgttctcggctctgtgaatgacaatgctgactggagtgctgcccctctgtaa agggctgggtgtggatggtcacaagcccc tcacatgcctcagccaagaggaagtagtacaggggtcagcccagaggtccaggggaaagg agtggaaaccgatttccccaccaag ggaggggcctgtacctcagctgttcccatagcttacttgccacaactgccaagcaagttt cgctgagtttgacacatggatccctgtgga tcaactgccctaggactccgtttgcacccatgtgacactgttgactttgccctgacgaag cagggccaacagtcccctaacttaattaca aaaactaatgactaagagagaggtggctagagctgaggcccctgagtcaggctgtgggtg ggatcatctccagtacaggaagtgag actttcatttcctcctttccaagagagggctgagggagcagggttgagcaactggtgcag acagcctagctggactttgggtgaggcg gttcagccatatcgaattctgctggggctactggcaggtaaggaggaaggaggctgaggg gagggggcccctgggagggagcctg ccctgggttgctaaccatctcctctctgccaaaagtccggaaagccaccatggagcccct gaggctgctcatcctgctgtttgtgacag aactgtctggagcccacaacaccacagtgttccagggagttgctggccagtctctgcaag tgtcttgcccctatgacagcatgaagcac tggggaaggaggaaggcttggtgtaggcagctgggagagaaaggaccttgccagagggtg gtgagcacacacaacctgtggctgc tgagcttcctcagaaggtggaatggctctacagccatcacagatgacaccctgggtggca ccctcaccatcaccttaaggaacctgca gcctcatgatgctggcctgtaccaatgccagagcctgcatggctctgaggctgataccct caggaaggtgttggtggaggtgctggct gatcctctggatcacagggatgctggagacctgtggttcccaggagagtctgagagcttt gaggatgcccatgtggagcacagcatca gcaggtctcttctggagggagagatccccttccctcccacaagcatcctgttgctgcttg cctgcatcttcctgatcaagatccttgctgct tctgctctttgggctgctgcctggcatggccagaaacctggaacacatcctccctctgaa ctggactgtggccatgaccctggctacca gttgcaaaccttgcctggcttgagggacacctgacaattgttaattaagtttaaaccctc gaggccgcaagcttatcgataatcaacctct ggattacaaaatttgtgaaagattgactggtattcttaactatgttgctccttttacgct atgtggatacgctgctttaatgcctttgtatcatgc tattgcttcccgtatggctttcattttctcctccttgtataaatcctggttgctgtctct ttatgaggagttgtggcccgttgtcaggcaacgtgg cgtggtgtgcactgtgtttgctgacgcaacccccactggttggggcattgccaccacctg tcagctcctttccgggactttcgctttcccc ctccctattgccacggcggaactcatcgccgcctgccttgcccgctgctggacaggggct cggctgttgggcactgacaattccgtgg tgttgtcggggaaatcatcgtcctttccttggctgctcgcctgtgttgccacctggattc tgcgcgggacgtccttctgctacgtcccttcg gccctcaatccagcggaccttccttcccgcggcctgctgccggctctgcggcctcttccg cgtcttcgccttcgccctcagacgagtcg gatctccctttgggccgcctccccgcatcgataccgtcgactagagctcgctgatcagcc tcgactgtgccttctagttgccagccatct gttgtttgcccctcccccgtgccttccttgaccctggaaggtgccactcccactgtcctt tcctaataaaatgaggaaattgcatcgcattg tctgagtaggtgtcattctattctggggggtggggtggggcaggacagcaagggggagga ttgggaagacaatagcaggcatgctg gggagagatccacgataacaaacagcttttttggggtgaacatattgactgaattcccgt gcggaccggcagtcctcacttacctgagt gccctgttctgcactcctctcctccccaccctgcccatattcttcacaccctccttccct ccccagtatcaaaacctaccctcagactctttct gctgtgagactgataaggggctggtggactctggttgcattggaagggaaggagtgctta cagtggcatcccaggcactagctggca gttgggggaagctttggggggcactggcactggtaatagcctctgaaattataagcctct catttgaagcccctacagttataaaggag gaaagaacctgaggtgattcatctgcatctttggggcactctctcccctgctctgaaggt ccccttgtcctcagctcttgtgaggaggtca ggagcatagagatttccattgctgagttgaggggaacttacacaccaaatttttgtacac aattgcttcttcccactgaggaaagcacacc ttttgctcccccagagcactggacaaacctgggcaagaggagagacagtgcctgaaagag tcttgttcctgcagcttcttagtgagtgg tcagggggaaggcagggttctgaggctgagtgaggaggtggtcagtgaggagagggccac agcccatctggtctctctagttccaa aaggcaagccaccaacttcccaatcctatctttcaagcctctgtcacagttgaagttgaa gtgagcaggcaagggaggaccaagaga gaagtcaaagcagggggctctgggggctgccccagcaacaggctggtgctctaagcccat ctcccccaccctaaggagggttccca aaatagcagcctctcatctcccccaaaatatctccaagacaggtccttcctgaaagggga acaaagccacagaaatagggaagctgg aagttaaaggtcaggaaagtcagtacctgtgaaggcagctgcagagcaagcaagagtggc agggcagggagagccagccccagg ccaagaggagaggtcccagtccctgtgaaaagcaaagagtgtgacagaacagaactggag ggggttttaagaccaagtgcctccag aatagacccagagaaggtcagttacttctccaaggaggcccagcaagagcaacagtgaga aaaccaaacccaggtcccaggtttcct ggttcccaatttcccacaaacacttactgtgccatccactcccaacttgtataagaacct aaggcggaccgagcggccgcaggaaccc ctagtgatggagttggccactccctctctgcgcgctcgctcgctcactgaggccgggcga ccaaaggtcgcccgacgcccgggcttt gcccgggcggcctcagtgagcgagcgagcgcgcagagagggagtggccaa