DESCRIPTION

The present invention applies to the medical field, and in particular for the treatment of tumors.

DNA polymerases (Pols) are enzymes involved in important cellular processes such as gene expression, regulation, transcription, and DNA damage repair.

Humans possess at least 16 different DNA polymerases, which are divided into families, each with different specific roles and functions.

In particular, Y-Family Pols is an important group of enzymes capable of conducting the synthesis of translesion DNA (TLS): a biological process that involves the replication of damaged DNA, with good precision.

Such enzymes are capable of bypassing damaged bases, which would otherwise block the normal progression of the replication fork.

Each Y-family polymerase is unique and has different "preferences" for lesions to bypass and for deoxyribonucleoside triphosphate (dNTP) to be incorporated.

In particular, among the Y-family polymerases, DNA polymerase p (Pol) is capable of bypassing UV-induced cyclobutane pyrimidine (CPD) cis-syn dimers, suppressing the mutagenic effect of UV- induced DNA damage (Yang, 2014).

In fact, genetic defects in the gene Pol η result in the development of a variant form of xeroderma pigmentosa (XPV) and the patients are much more sensitive to sunlight and prone to developing skin cancer.

Aurintricarboxylic acid and ellagic acid, known inhibitors of Pol η, show very promising nanomolar IC50 values (Dorjsuren et al., 2009), but are characterized by poor selectivity and with an unspecific action mechanism that therefore limits the use thereof mainly to biological experiments only.

So far, the most potent class of Pol p inhibitors are N- benzoyl indolylbarbituric acid (ITBA) derivatives, which show IC50s in the low micromolar range (Coggins et al., 2013) and act on the allosteric site of the enzyme.

Nowadays, platinum-based drugs and nucleoside analogues are regularly prescribed in the treatment of cancer, and although they are effective, the use thereof is limited by inherent or acquired resistance.

Mechanically, cisplatin and the analogues thereof react with DNA bases to cross-link adjacent purines.

The preferential activation of DNA damage responses prevents replication fork collapse and promotes cell survival during chemotherapy treatment, leading to chemoresistance (Srivastava et al., 2015).

To date, there are no drugs for treating cancer in clinical practice and overcoming drug resistance to platinum-based drugs and nucleoside analogues.

Prior art document US 2010/0035887 describes poxvirus DNA polymerase inhibitor compounds. International patent application WO 2006/076863 describes compounds having the general formula (I) shown in figure 1 having synergistic activity with anticancer agents such as cisplatin and/or 5-fluorouracil.

Prior art document "Synthesis, biological evaluation and SAR analysis of O-alkylated analogs of quercetin for anticancer" (Shi et al. Bioorganic & Medicinal Chemistry Letters 24 (2014) 4424- 4427) describes the synthesis of O-alkylated analogues of quercetin having anti-cancer activity. The general structure of such compounds is shown in figure 1.

Prior art document "A Small-Molecule Inhibitor of Human DNA Polymerase η Potentiates the Effects of Cisplatin in Tumor Cells." (Zafar et al. Biochemistry. 2018 February 20; 57(7): 1262-1273) describes the compound PNR-7-02, shown in figure 1, as a polymerase inhibitor. The same compound is described by WO 2006/076863 (WU YIXIN, China).

Prior art document WO 2019/008537 (Vera Salus Ricerca SRL, Italy) describes quercetin-derived compounds for medical use in the treatment of metastatic tumors and malignancies, depicted in figure 1.

Prior art document WO 2011/099978 (N30 Pharma. INC) describes flavone derivatives for treating cancer, tumors and metastatic neoplasms, depicted in figure 1.

Summary of the invention

The inventors of the present patent application have surprisingly found that the compounds characterized by the general formula (I) can be employed for treating diseases or disorders represented by primary and/or metastatic neoplasms, which have developed resistance after previous treatment with chemotherapeutic compounds.

Such an activity is completely unexpected based on the structure of similar compounds already known to have antiviral activity.

Object of the invention

In a first object, the present patent application describes compounds for medical use in treating diseases or disorders represented by primary and/or metastatic neoplasms, which have developed resistance after previous treatment with chemotherapeutic compounds.

According to an even more particular aspect, such compounds are described for treating tumors in combination with other chemotherapeutic compounds.

In a second object, the present patent application describes compounds per se with the condition given in the following description.

In a third object, the present patent application describes pharmaceutical formulations comprising the compounds of the invention.

In another object, the present patent application describes a method for treating diseases or disorders represented by primary and/or metastatic neoplasms, which have developed resistance after previous treatment with chemotherapeutic compounds comprising the administration of a pharmaceutically effective amount of a compound or formulation according to the invention. According to an even more particular aspect, such a method is carried out in association with other chemotherapeutic compounds. Brief description of the drawings

Figure 1 shows the structure of compounds known in the art.

Figures 2, 3 and 4 show Schemes 1, 2 and 3 mentioned in the present patent application, respectively.

Detailed description of the invention

In accordance with a first object, compounds of formula (I) for medical use are described.

In particular, the compounds have the following formula: wherein

R ¹ , R ² , R ³ , R ⁴ , R ⁵ and R ⁶ are, independently of each other, -H, -OH, -R ⁷ , -x, -OR ⁷ , -R ⁷ (X)n, -NO ₂ , -NH ₂ , -NHR ⁷ , where

R ⁷ is a linear or branched C _1-4 alkyl optionally substituted with one or more groups selected from -OH, -X, -NO ₂ , -NH ₂ , -NHR ⁸ , where R ⁸ is a linear or branched C _1-4 alkyl, or where

R ¹ , R ² , R ³ , R ⁴ and R ⁷ are as described above and R ⁵ and R ⁶ form, together with the benzene ring to which they are bound, a (2,2- difluorobenzo[d][1,3]dioxol cycle: , and X is halogen in any position of the chain, n=1-4 on the entire chain and preferably 1-3 and

A is preferably a single bond or, alternatively, is one of:

According to a preferred aspect of the present invention, the described compounds have the following formula: wherein

R ¹ , R ² , R ³ , R ⁴ , R ⁵ and R ⁶ are, independently of each other, -H, -OH, -R ⁷ , -X, -OR ⁷ , —R ⁷ (X) _n , -NO ₂ , -NH2, -NHR ⁷ , wherein

R ⁷ is a linear or branched C _1-4 alkyl optionally substituted with one or more groups selected from -OH, -X, -NO ₂ , -NH ₂ , -NHR ⁸ , where R ⁸ is a linear or branched C _1-4 alkyl, or wherein

R ¹ , R ² , R ³ , R ⁴ and R ⁷ are as described above and R ⁵ and R ⁶ form, together with the benzene ring to which they are bound, a (2,2- difluorobenzo[d][1,3]dioxol cycle: , and

X is halogen in any position of the chain, n=1-4 on the entire chain and preferably 1-3 and

A is preferably a single bond or alternative

In accordance with an even more preferred aspect, the following compounds for medical use are described:

According to a particularly preferred aspect of the present invention, the following compounds for medical use are described: According to an even more particularly preferred aspect, the following compounds for medical use are described:

For the purposes of the present invention, the compounds above are described for medical use in the treatment of diseases or disorders associated with increased activity and/or expression of DNA polymerase η.

Objects of the present invention are also prodrugs of the above compounds capable of increasing the bioavailability thereof in the body, such as boronates and hyaluronic acid polymers.

In particular, such a medical use is described for treating diseases or disorders associated with increased activity and/or expression of DNA polymerase p represented by neoplasms. More in particular, such neoplasms are represented by primary and/or metastatic neoplasms.

According to a preferred aspect of the present invention, such a medical use is described for treating primary and/or metastatic neoplasms, which have developed resistance after a prior treatment with chemotherapeutic compounds.

In particular, for the purposes of the present invention, such chemotherapeutics are represented by nucleoside analogs or alkylating agents derived from platinum, e.g., Cis platinum or analog compounds.

In particular, such neoplasms are represented by: ovarian cancer, breast cancer, pancreatic cancer, lung cancer, gastric adenocarcinoma, mucosa-derived squamous cell carcinoma of the neck.

In accordance with an aspect of the present invention, the compounds of the present patent application are described for medical use in association with chemotherapeutic compounds.

In particular, such chemotherapeutic compounds are represented by nucleoside analogs or alkylating agents derived from platinum, such as Cis platinum or analog compounds, such as carboplatin or oxaliplatin.

According to an alternative aspect, the compounds of the present patent application are described for medical use in association with other compounds having another mechanism of action, possibly in association with or as an alternative to radiotherapy. In an aspect of the invention, the compounds of the present patent application are described for medical use so as to avoid the occurrence of treatment resistance phenomena conducted against a primary and/or metastatic neoplasia.

For the purposes of the present invention, the term "in association" means a not necessarily simultaneous therapeutic association, where the simultaneous association represents a preferred aspect.

In a second object, the present patent application describes compounds of formula (I): wherein

R ¹ , R ² , R ⁴ , R ⁵ and R ⁶ are, independently of each other, -H, -R ⁷ , -X, -OH, -OR ⁷ , -R ⁷ (X)n, -NO ₂ , -NH ₂ , -NHR ⁷ , wherein

R ⁷ is a linear or branched C _1-4 alkyl optionally substituted with one or more groups selected from -OH, -X, -NO ₂ , -NH ₂ , -NHR ⁸ , where R ⁸ is a linear or branched C _1-4 alkyl,

R ³ is H,

X is halogen in any position of the chain, n=1-4 on the entire chain and preferably 1-3 and

A is a single bond, provided that the compounds of formula (I) do not include:

According to an even more preferred aspect, the compounds of formula (I) are represented by the following compounds:

In accordance with a second object, the present patent application describes formulations comprising the compounds of the invention. Such formulations can be administered orally, nasally, subcutaneously or intramuscularly.

In particular, such formulations comprise one or more pharmaceutically acceptable excipients being suitable for the route of administration.

According to a particular aspect, the formulations of the invention are also described for the medical use and uses reported above.

In accordance with another object, the present patent application describes a method for treating diseases or disorders associated with increased activity and/or expression of DNA polymerase p comprising administering a pharmaceutically effective amount of a compound or formulation according to the invention.

More in particular, such neoplasms are represented by primary and/or metastatic neoplasms.

According to a preferred aspect of the present invention, such a method is described for treating primary and/or metastatic neoplasms, which have developed resistance after a prior treatment with chemotherapeutic compounds.

In particular, for the purposes of the present invention, such chemotherapeutics are represented by nucleoside analogs or alkylating agents derived from platinum, e.g., Cis platinum or analog compounds.

In particular, such neoplasms are represented by: ovarian cancer, breast cancer, pancreatic cancer, lung cancer, gastric adenocarcinoma, mucosa-derived squamous cell carcinoma of the In accordance with an aspect of the present invention, the treatment method is described in association with chemotherapeutic compounds.

In particular, such chemotherapeutic compounds are represented by nucleoside analogs or alkylating agents derived from platinum, such as Cis platinum or analog compounds.

According to an alternative aspect, the treatment method of the present patent application is described in association with other compounds having a different mechanism of action, possibly in association with or as an alternative to radiotherapy.

In an aspect of the invention, the method of the present patent application is described to avoid the occurrence of treatment resistance phenomena conducted against a primary and/or metastatic neoplasia.

For the purposes of the present invention, the term "in association" means a not necessarily simultaneous therapeutic association, where the simultaneous association represents a preferred aspect.

The present invention is further described in the following experimental section.

Experimental section

Synthesis - General considerations

All the commercially available reagents and solvents were used as purchased from vendors without further purification. The intermediates la and lb can be synthesized according to the procedure shown in European Journal of Medicinal Chemistry 180 (2019) 350-366. The intermediate 1c can be synthesized according to the procedure shown in prior art document WO 2017/132928 Al. The anhydrous solvents were purchased from Sigma-Aldrich. Automated column chromatography purifications were done using a Teledyne ISCO apparatus (CombiFlash® Rf) with pre-packed silica gel columns of different sizes (from 4 g up to 120 g) and mixtures of increasing polarity of cyclohexane and ethyl acetate (EtOAc) or dichloromethane (DCM) and methanol (MeOH). NMR experiments were run on a Bruker Avance III 400 system (400.13 MHz for 1H, and 100.62 MHz for 13C), equipped with a BBI probe and Z-gradients. Spectra were acquired at 300 K, using deuterated dimethylsulfoxide (DMSO-d ₆ ) or deuterated chloroform (CDCI ₃ ) as solvents. For ¹ H-NMR, data are shown as follows: chemical shift, multiplicity (s= singlet, d= doublet, dd= double of doublets, ddd= doublet of doublet of doublets, t= triplet, td= triplet of doublets, q= quartet, p= quintet, m= multiplet), coupling constants (Hz) and integration. UPLC/MS analyses were run on a Waters Acquity UPLC/MS system consisting of a SQD (single quadrupole detector) mass spectrometer equipped with an electrospray ionization interface and a photodiode array detector. The PDA range was 210-400 nm. The analyses were performed on an ACQUITY UPLC BEH C18 column (100x2.1 mmID, 1.7 pm particle size) with a VanGuard BEH C18 pre-column (5x2.1 mmID, 1.7 pm particle size). The mobile phase was 10 mM NH ₄ OAC in H ₂ O at pH 5 adjusted with CH ₃ COOH (A) and 10 M NH ₄ O C in CH ₃ CN-H2O (95:5) at pH 5.0. Two types of gradients were applied depending on the analysis, gradient 1 (5% to 100% mobile phase B in 3 min) or gradient 2 (50% to 100% mobile phase B in 3 min). Electrospray ionization in positive and negative mode was applied. ESI was applied in positive and negative mode. All tested compounds showed ≥ 95% purity by UPLC/MS analysis.

General procedure A. Aldol condensation (Scheme 1).

The appropriate ketone of type 1 (1.0 eq.) and benzaldehyde of type 2 (1.1 eq) were added to a solution of potassium hydroxide (20.0 eq.) in MeOH (0.12 M). The reaction mixture was stirred at room temperature. After complete conversion of starting materials, the reaction mixture was acidified to pH=5 with 1 M HC1 and extracted with EtOAc (3x5 mL). Combined organic layers were dried over magnesium sulfate, filtered and concentrated under vacuum. The product was purified by flash chromatography or trituration with EtOH giving the pure intermediate of type 3.

General procedure B. Oxidative cyclization (Scheme 1).

The appropriate chaicone of type 3 (1 eq.) was dissolved in DMSO (0.3 M) and heated at 135°C under argon atmosphere. I2 (0.05 eq.) was added and the reaction mixture was stirred until full conversion of the starting material. After reaction completion, the reaction mixture was cooled to room temperature and sodium thiosulfate 1 N was added to quench iodine. The crude product was filtered, washed with water and purified by flash chromatography giving the pure compound of type 4.

General procedure C. Methyl deprotection (Scheme 1).

Method A.

The appropriate compound of type 4 (1 eq.) was treated with pyridinium chloride (10 eq.) and the reaction mixture was heated at 190°C under argon atmosphere until total conversion of the starting material. After reaction completion, the reaction mixture was cooled down to room temperature and added with water. The crude product was filtered, washed with water and purified by silica.

Method B.

The appropriate compound of type 4 (1 eq.) was dissolved in DCM (0.05 M) and cooled to 0°C. A IM solution of boron tribromide in DCM (1.5 eq for each methoxy group) was added and the reaction mixture was allowed to warm to room temperature and stirred until complete conversion of the starting material under argon atmosphere. After reaction completion, the reaction was quenched with MeOH and concentrated under reduced pressure. The crude product was redissolved in MeOH, concentrated under vacuum and purified by flash chromatography.

General Procedure D. Benzyl deprotection (Scheme 1, 2)

An appropriate benzylated compound was dissolved in a 1:1 mixture MeOH/DCM (0.04 M) under an argon atmosphere. Pd/C (20% w/w) triethylsilane (6 eq. for each benzyl group) were added to the solution. The reaction mixture was stirred at 40°C until complete conversion of the starting material. Then, the reaction mixture was filtered over a bed of celite and concentrated under vacuum. The crude product was taken up in ethyl acetate and the organic phase was washed with water, dried over magnesium sulfate, filtered and concentrated under vacuum, and purified by silica.

Synthesis of 2- (3,4-dihydroxyphenyl)-5,7-dihydroxy-3-propy1-4H- chromen-4-one (compound 5aa, Scheme 1). Step 1. Synthesis of (E)-2-(3,4-dimethoxybenzylidene)-1-(2- hydroxy-4,6-dimethoxyphenyl)pentane-1-one (Intermediate 3aa, Scheme 1)

The title compound was synthesized following the general procedure A described above using 1-(2-hydroxy-4,6-dimethoxyphenyl)pentane-1-one la (105 mg, 0.60 mmol), 3,4-dimethoxybenzaldehyde 2a (101 mg, 0.66 mmol) and potassium hydroxide (675 mg, 12 mmol) in MeOH (5 mL). Purification by silica (elution by gradient from 100 to 70/30 cyclohexane/EtOAc) gave the pure intermediate 3aa (116 mg, 50% yield).

Characterization: UPLC/MS Rt: 2.40 min (gradient 1), MS (ESI) m/z: 387.2 [M+H] ⁺ .[M+H] ⁺ Calculated for C ₂₂ H ₂₇ O ₆ : 387.2. ¹ H-NMR (400 MHz,

DMSO-d ₆ ) δ 9.70 (s, 1H), 7.04 (s, 1H), 7.01 (d, J = 8.1 Hz, 1H),

6.97 - 6.89 (m, 2H), 6.12 (d, J = 2.1 Hz, 1H), 6.07 (d, J = 2.1

Hz, 1H), 3.78 (s, 3H), 3.76 (s, 3H), 3.74 (s, 3H), 3.65 (s, 3H), 1.57 - 1.44 (m, 2H), 1.17 (t, J = 7.1 Hz, 2H), 0.96 (t, J = 7.3

Hz, 3H).

Step 2, Synthesis of 2-(3,4-dimethoxyphenyl)-5,7-dimethoxy-3- propy1-4H-chromen-4-one (Intermediate 4aa, Scheme 1).

The title compound was synthesized following the general procedure B described above using intermediate 3aa (116 mg, 0.3 mmol) and I2 (4 mg, 0.01 mmol) in DMSO (1 mL). Purification by silica (elution by gradient from 15/85 to 50/50 cyclohexane/EtOAc) gave the pure intermediate 4aa (54 mg, 50% yield).

Characterization: UPLC/MS Rt: 2.23 min (gradient 1), MS (ESI) m/z: 385.2 [M+H] ⁺ .[M+H] ⁺ Calculated for C ₂₂ H ₂₅ O ₆ : 385.2. ¹ H-NMR (400 MHz, DMSO-d ₆ ) δ 7.20 - 7.15 (m, 2H), 7.12 (s, 1H), 6.64 (d, J = 2.3 Hz, 1H), 6.48 (d, J = 2.3 Hz, 1H), 3.85 (s, 3H), 3.84 (s, 3H), 3.83 (s, 3H), 3.81 (s, 3H), 2.38 - 2.29 (m, 2H), 1.53 - 1.39 (m, 2H), 0.82 (t, J = 7.3 Hz, 3H).

Step 3. Synthesis of 2-(3,4-dihydroxyphenyl)-5,7-dihydroxy-3- propy1-4H-chromen-4-one (compound 5aa, Scheme 1).

The title compound was synthesized following the general procedure

C Method A described above using intermediate 4aa (54 mg, 0.14 mmol) and pyridinium chloride (162 mg, 1.4 mmol). Purification by silica (elution by gradient from 100 to 94:6 DCM/MeOH) gave the pure compound 5aa (35 mg, 76% yield).

Characterization: UPLC/MS Rt: 1.93 min (gradient 1), MS (ESI) m/z: 329.3 [M+H] ⁺ .[M+H] ⁺ Calculated for C ₁₈ H ₁₇ O ₆ : 329.1. ¹ H-NMR (400 MHz, DMSO-d ₆ ) δ 13.08 (s, 1H), 7.01 (d, J = 2.1 Hz, 1H) 6.93 (dd, J = 8.2, 2.1 Hz, 1H), 6.88 (d, J = 8.2 Hz, 1H), 6.28 (d, J = 2.1 Hz, 1H), 6.16 (d, J = 2.1 Hz, 1H), 2.44 - 2.35 (m, 2H), 1.56 - 1.42 (m, 2H), 0.84 (t, J = 7.3 Hz, 3H). ¹³ C-NMR (101 MHz, DMSO-d ₆ ) δ 182.20, 164.66, 162.78, 161.93, 157.77, 148.25, 145.67, 123.83,

120.86, 118.66, 116.21, 116.02, 103.53, 99.01, 93.77, 27.07,

22.25, 14.53.

Synthesis of 2-(3,4-dihydroxyphenyl)-7-fluoro-5-hydroxy-4H- chromen-4-one (compound 5ca, Scheme 1).

Step 1. Synthesis of (E)-3-(3,4-dimethoxyphenyl)-1-(4-fluoro-2- hydroxy-6-methoxyphenyl)prop-2-en-1-one (compound 3ca, Scheme 1).

The title compound was synthesized following the general procedure

A described above using: ketone lc (150 mg, 0.46 mmol), 3,4- dimethoxybenzaldehyde 2a (84 mg, 0.51 mmol), KOH (515 mg, 9.2 mmol) in anhydrous MeOH (3.9 mL). The crude product was purified by trituration with EtOH (1.5 mL) to give the title intermediate 3ca (120 mg, 92% yield).

Characterization: UPLC/MS Rt = 2.47 min (gradient 1), MS (ESI) m/z 333.5, [M+H] ⁺ . [M+H] ⁺ calculated for C ₁₈ H ₁₈ FO ₅ : 333.3. ¹ H NMR (400

MHz, CDC13) δ 13.90 (d, J = 1.4 Hz, 1H), 7.81 (d, J = 15.5 Hz,

1H), 7.72 (d, J = 15.5 Hz, 1H), 7.23 (dd, J = 8.3, 2.0 Hz, 1H),

7.13 (d, J = 2.0 Hz, 1H), 6.91 (d, J = 8.3 Hz, 1H), 6.32 (dd, J =

10.1, 2.4 Hz, 1H), 6.16 (dd, J = 11.0, 2.4 Hz, 1H), 3.95 (s, 3H),

3.94 (s, 3H), 3.94 (s, 3H).

Step 2, Synthesis of 2-(3,4-dimethoxyphenyl)-7-fluoro-5-methoxy- 4H-chromen-4-one (compound 4ca, Scheme 1).

Intermediate 4ca was prepared according to general procedure B described above using: Intermediate 3ca (150 mg, 0.45 mmol), I ₂ (6 mg, 0.02 mmol) in anhydrous DMSO (1.5 mL). The crude product was purified by trituration with EtOH (1 mL) to give the title intermediate 4ca (120 mg, 80% yield).

Characterization: UPLC/MS Rt = 1.93 min (gradient 1), MS (ESI) m/z 331.1, [M+H] ⁺ . [M+H] ⁺ calculated for C ₁₈ H ₁₆ FO ₅ : 331.3. 1H NMR (400

MHz, CDCI ₃ ) δ 7.50 (dd, J = 8.5, 2.2 Hz, 1H), 7.31 (d, J = 2.1 Hz, 1H), 6.97 (d, J = 8.5 Hz, 1H), 6.82 (dd, J = 9.0, 2.4 Hz, 1H), 6.64 (s, 1H), 6.57 (dd, J = 11.1, 2.4 Hz, 1H), 3.99 (s, 3H), 3.97 (s, 3H), 3.95 (s, 3H).

Step 3. Synthesis of 2-(3,4-dihydroxyphenyl)-7-fluoro-5-hydroxy-

4H-chromen-4-one (compound 5ca, Scheme 1).

The title compound was prepared according to general procedure C, Method B described above using: intermediate 4ca (100 mg, 0.30 mmol), 1 M BBr ₃ in DCM (1.4 mL, 1.4 mmol) in anhydrous DCM (6 mL). Crystallization from EtOH (1.5 mL) gave the pure compound 5ca (56 mg, 66% yield).

Characterization: UPLC/MS Rt = 1.99 min (gradient 1), MS (ESI) m/z 287.1, [M-H]-. [M-H]- calculated for C ₁₅ H ₈ FO ₅ : 287.2. NMR (400 MHz, DMSO-d ₆ ) δ 13.24 (s, 1H), 7.47 (dd, J = 8.4, 2.3 Hz, 1H), 7.44

(d, J = 2.3 Hz, 1H), 7.08 (dd, J = 10.0, 2.4 Hz, 1H), 6.91 (d, J = 8.3 Hz, 1H), 6.84 (s, 1H), 6.72 (dd, J = 10.9, 2.3 Hz, 1H).

Synthesis of 2-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)-5-hydroxy-7- methoxy-4H-chromen-4-one (compound 5.1bb, Scheme 1).

Step 1. Synthesis of (E)-3-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)- 1-(2-hydroxy-4,6-dimethoxyphenyl)prop-2-en-1-one) (intermediate 3bb, Scheme 1).

The intermediate 3bb was prepared according to general procedure A described above using: ketone lb (200 mg, 1.00 mmol), 2,2- Difluoro-1,3-benzodioxol-5-carboxaldehyde 2b (210 g, 1.1 mmol), KOH (1680 mg, 20 mmol) in anhydrous MeOH (8.5 mL). The crude product was purified by trituration with EtOH (3 mL) to give the title intermediate 3bb (200 mg, 55% yield).

Characterization: UPLC/MS Rt = 1.98 min (gradient 1), MS (ESI) m/z 365.0, [M+H] ⁺ . [M+H] ⁺ calculated for C ₁₈ H ₁₅ F ₂ O ₆ : 365.3. ¹ H NMR (400

MHz, CDCI ₃ ) δ 14.19 (s, 1H), 7.79 (d, J = 15.5 Hz, 1H), 7.70 (d, J = 15.6 Hz, 1H), 7.32 (d, J = 1.6 Hz, 1H), 7.30 (dd, J = 8.8, 1.7 Hz, 1H), 7.08 (d, J = 8.7 Hz, 1H), 6.11 (d, J = 2.3 Hz, 1H), 5.97 (d, J = 2.4 Hz, 1H), 3.92 (s, 3H), 3.84 (s, 3H).

Step 2. Synthesis of intermediate n (2-(2,2- difluorobenzo[d][1,3]dioxol-5-yl)-5,7-dimethoxy-4H-chromen-4 -one) (compound 4bb).

Intermediate 4bb was prepared according to general procedure B described above using: intermediate 3bb (180 mg, 0.51 mmol), I2 (6.4 mg, 0.02 mmol) in anhydrous DMSO (1.7 mL). The crude product was purified by trituration with EtOH (1 mL) to give the title compound 4bb (140 mg, 79% yield).

Characterization: UPLC/MS Rt = 1.06 min (gradient 2), MS (ESI) m/z 363.1, [M+H] ⁺ . [M+H] ⁺ calculated for C ₁₈ H ₁₃ F ₂ O ₆ : 363.3. ¹ H NMR (400

MHz, CDCl ₃ ) δ 7.64 (dd, J = 8.4, 1.8 Hz, 1H), 7.57 (d, J = 1.8 Hz, 1H), 7.17 (d, J = 8.5 Hz, 1H), 6.61 (d, J = 0.9 Hz, 1H), 6.55 (d, J = 2.2 Hz, 1H), 6.38 (d, J = 2.3 Hz, 1H), 3.95 (s, 3H), 3.91 (s, 3H).

Step 3. Synthesis of 2-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)-5- hydroxy-7-methoxy-4H-chromen-4-one (compound 5.1bb, Scheme 1).

Compound 5.lbb was prepared according to general procedure C, Method B described above using: intermediate 4bb (100 mg, 0.28 mmol), 1 MBBr ₃ in CH ₂ CI ₂ (0.42 mL, 0.42 mmol) in anhydrous CH ₂ CI ₂ (10 mL). Crystallization from EtOH (1.5 mL) gave the pure compound 5.lbb (56 mg, 57% yield).

Characterization: UPLC/MS Rt = 2.67 min (gradient 1), MS (ESI) m/z 247.0, [M-H]-. [M-H]- calculated for C ₁₇ H ₉ F ₂ O ₆ : 347.3. 1H NMR (400

MHz, DMSO-d ₆ ) δ 7.44 (dd, J = 8.3, 2.3 Hz, 1H), 7.41 (d, J = 2.3 Hz, 1H), 6.86 (d, J = 8.3 Hz, 1H), 6.72 (d, J = 2.3 Hz, 1H), 6.70 (s, 1H), 6.37 (d, J = 2.2 Hz, 1H), 3.87 (s, 3H).

Synthesis of 2-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)-5,7- dihydroxy-4H-chromen-4-one (compound 5.2bb, Scheme 1).

Compound 5.2bb was prepared according to general procedure C method A using: intermediate 4bb (100 mg, 0.28 mmol), pyridinium chloride (324, 2.8 mmol). Crystallization with EtOH (1.5 mL) gave the pure compound 5.2bb (28 mg, 30% yield).

Characterization: UPLC/MS Rt = 2.28 min (gradient 1), MS (ESI) m/z 333.1, calculated for C ₁₆ H ₇ F ₂ O ₆ : 333.2. 1H NMR (400

MHz, DMSO) δ 12.71 (s, 1H), 8.12 (d, J = 1.8 Hz, 1H), 7.94 (dd, J = 8.6, 1.8 Hz, 1H), 7.57 (d, J = 8.6 Hz, 1H), 6.96 (s, 1H), 6.51 (d, J = 2.1 Hz, 1H), 6.19 (d, J = 2.1 Hz, 1H).

Synthesis of 5-methoxy-2-(or—tolyl)-4H-chromen-4-one (compound 4dc, Scheme 1).

Step 1. Synthesis of (E)-1-(2-hydroxy-6-methoxy-phenyl)-3-(or- tolyl)prop-2-en-1-one (compound 3dc, Scheme 1).

The title compound was synthesized following general procedure A using 1-(2-hydroxy-6-methoxyphenyl)ethan-1-one Id (100 mg, 0.60 mmol), 2-methylbenzaldehyde 2c (0.08 mL, 0.66 mmol) and potassium hydroxide (675 mg, 12 mmol) in MeOH (5 mL). Purification by silica (elution by gradient from 100 to 95/5 cyclohexane/EtOAc) gave the pure intermediate 3dc (118 mg, 73% yield).

Characterization: UPLC/MS Rt: 2.60 min (gradient 1), MS (ESI) m/z: 269.1 [M+H] ⁺ .[M+H]+ Calculated for C ₁₇ H ₁₇ O ₃ : 269.1. 1H-NMR (400 MHz, CDC13) δ 13.15 (s, 1H), 8.11 (d, J = 15.5 Hz, 1H), 7.79 (d, J =

15.5 Hz, 1H), 7.69 - 7.62 (m, 1H), 7.37 (dd like t, J = 8.3 Hz,

1H), 7.33 - 7.21 (m, 3H), 6.62 (d, J = 8.3 Hz, 1H), 6.44 (d, J =

8.3 Hz, 1H), 3.95 (s, 3H), 2.51 (s, 3H).

Step 2. Synthesis of 5-methoxy-2-(or-tolyl)-4H-chromen-4-one (compound 4dc, Scheme 1).

The title compound was synthesized following general procedure B using compound 3dc (118 mg, 0.70 mmol) and I2 (8 mg, 0.03 mmol) in DMSO (1 mL). Purification by silica (elution by gradient from 100 to 50/50 cyclohexane/EtOAc) gave the pure intermediate 4dc (79 mg, 75% yield).

Characterization: UPLC MS Rt: 2.01 min (gradient 1), MS (ESI) m/z: 267.1 [M+H] ⁺ .[M+H] ⁺ Calculated for C ₁₇ H ₁₅ O ₃ : 267.1. ¹ H-NMR (400 MHz, DMSO-d ₆ ) δ 7.69 (dd like t, J = 8.4 Hz, 1H), 7.59 (dd, J = 7.7, 1.4 Hz, 1H), 7.46 (ddd like td, J = 7.5, 1.4 Hz, 1H), 7.41 - 7.31 (m, 2H), 7.18 (d, J = 8.3 Hz, 1H), 7.01 (d, J = 8.3 Hz, 1H), 6.33 (s,

1H), 3.87 (s, 3H), 2.43 (s, 3H). 1 ³ C-NMR (101 MHz, DMSO-d ₆ ) δ 176.24, 162.79, 159.21, 157.84,

136.42, 134.35, 131.77, 131.12, 130.70, 129.13, 126.26, 113.64, 112.69, 109.91, 107.29, 56.18, 20.12.

Synthesis of 2-(2-fluorophenyl)-5-methoxy-4H-chromen-4-one

(compound 4dd, Scheme 1)

Step 1. Synthesis of (E)-3-(2-fluorophenyl)-1-(2-hydroxy-6- methoxyphenyl)prop-2-en-1-one (compound 3dd, Scheme 1).

The title compound was synthesized following general procedure A using ketone Id (100 mg, 0.60 mmol), 2-fluorobenzaldehyde 2d (82.0 mg, 0.66 mmol) and KOH (675 mg, 12 mmol) in MeOH (5 mL). Trituration with EtOH gave the pure compound 3dd (96 mg, 59% yield).

Characterization: UPLC/MS Rt: 2.52 min (gradient 1), MS (ESI) m/z: 273.1 [M+H] ⁺ .[M+H] ⁺ Calculated for C ₁₆ H ₁₄ FO ₃ : 273.1. ¹ H-NMR (400 MHz, CDCI3) δ 13.10 (s, 1H), 7.98 (d, J = 15.8 Hz, 1H), 7.88 (d, J =

15.8 Hz, 1H), 7.61 (ddd like td, J = 7.6, 1.8 Hz, 1H), 7.42 - 7.32

(m, 2H), 7.19 (ddd like td, J = 7.5, 1.1 Hz, 1H), 7.15 (d, J = 1.1

Hz, 1H), 6.62 (dd, J = 8.4, 1.0 Hz, 1H), 6.44 (dd, J = 8.4, 1.0

Hz, 1H), 3.95 (s, 3H).

Step 2. Synthesis of 2-(2-fluorophenyl)-5-methoxy-4H-chromen-4-one

(compound 4dd, Scheme 1). The title compound was synthesized following general procedure B using intermediate 3dd (96 mg, 0.40 mmol) and I ₂ (5 mg, 0.02 mmol) in DMSO (1 mL). Purification by silica (elution by gradient from 100 to 50/50 Cyclohexane/EtOAc) gave the pure compound 4dd (79 mg, 63% yield).

Characterization: UPLC/MS Rt: 1.96 min (gradient 1), MS (ESI) m/z: 271.1 [M+H]+.[M+H] ⁺ Calculated for C ₁₆ H ₁₂ FO ₃ : 271.1. ¹ H-NMR (400 MHz, DMSO-d ₆ ) δ 8.01 (ddd like td, J = 7.9, 1.8 Hz, 1H), 7.72 (dd like t, J = 8.4 Hz, 1H), 7.68 - 7.62 (m, 1H), 7.49 - 7.39 (m, 2H), 7.24 (d, J = 8.4 Hz, 1H), 7.02 (d, J = 8.3 Hz, 1H), 6.61 (s, 1H), 3.87 (s, 3H).

Synthesis of 5-methoxy-2-(2-(trifluoromethyl)phenyl)-4H-chromen-4- one (compound 4de, Scheme 1)

Step 1. Synthesis of (E)-1-(2-hydroxy-6-methoxyphenyl)-3-(2-

(trifluoromethyl)phenyl)prop-2-en-1-one (intermediate 3de, Scheme

1)- The title compound was synthesized following general procedure A using ketone Id (100 mg, 0.60 mmol), 2- (trifluoromethyl)benzaldehyde 2e (0.09 mL, 0.66 mmol) and KOH (675 mg, 12 mmol) in MeOH (5 mL). Trituration with EtOH gave the pure compound 3de (126 mg, 65% yield).

Characterization: UPLC/MS Rt: 2.64 min (gradient 1), MS (ESI) m/z: 323.1 [M+H] ⁺ .[M+H] ⁺ Calculated for C ₁₇ H14F ₃ O ₃ : 323.1. ¹ H-NMR (400

MHz, CDCI ₃ ) δ 13.01 (s, 1H), 8.17 - 8.07 (m, 1H), 7.81 (d, J = 1.9 Hz, 1H), 7.78 (d, J = 6.0 Hz, 1H), 7.76 - 7.70 (m, 1H), 7.60 (t, J = 7.6 Hz, 1H), 7.49 (t, J = 7.7 Hz, 1H), 7.38 (t, J = 8.3 Hz, 1H), 6.63 (dd, J = 8.4, 1.0 Hz, 1H), 6.43 (dd, J = 8.3, 1.0 Hz, 1H), 3.94 (s, 3H).

Step 2. Synthesis of 5-methoxy-2-(2-(trifluoromethyl)phenyl)-4H- chromen-4-one (compound 4de, Scheme 5).

The title compound was synthesized following general procedure B using intermediate 3de (126 mg, 0.35 mmol) and I2 (5 mg, 0.02 mmol) in DMSO (1 mL). Purification by silica (elution by gradient from 100 to 94/6 DCM/MeOH) gave the pure compound 4de (108 mg, 84% yield).

Characterization: UPLC/MS Rt: 2.13 min (gradient 1), MS (ESI) m/z: 321.1 [M+H] ⁺ .[M+H] ⁺ Calculated for C ₁₇ H ₁₂ F ₃ O: 321.1. ¹ H-NMR (400 MHz, DMSO-d ₆ ) 6 8.00 - 7.93 (m, 1H), 7.89 - 7.77 (m, 3H), 7.71 (dd like t, J = 8.4 Hz, 1H), 7.07 (d, J = 8.4 Hz, 1H), 7.04 (d, J = 8.4 Hz, 1H), 6.42 (s, 1H), 3.88 (s, 3H).

Synthesis of 2-(4-hydroxy-2-methylphenyl)-5-methoxy-4H-chromen-4- one (compound 5df, Scheme 1).

Step 1. Synthesis of (E)-3-(4-benzyloxy)-2-methoxyphenyl)-1-(2- hydroxy-6-methoxyphenyl)prop-2-en-1-one (compound 3df, Scheme 1)

The title compound was synthesized following general procedure A using ketone Id (100 mg, 0.60 mmol), 4-benzyloxy-2-methyl- benzaldehyde 2f (149 mL, 0.66 mmol) and KOH (675 mg, 12 mmol) in MeOH (5 mL). The crude product was triturated with MeOH to give pure compound 3df (224 mg, quantitative yield).

Characterization: UPLC/MS Rt: 2.21 min (gradient 2), MS (ESI) m/z: 375.1 [M+H] ⁺ .[M+H]+ Calculated for C ₂₄ H ₂₃ O ₄ : 375.1. ¹ H-NMR (400 MHz, DMSO-d ₆ ) δ 7.71 (d, J = 8.6 Hz, 1H), 7.58 (d, J = 15.8 Hz, 1H), 7.48 - 7.30 (m, 5H), 7.26 (dd like t, J = 8.3 Hz, 1H), 7.01 (d, J = 15.8 Hz, 1H), 6.97 - 6.85 (m, 2H), 6.63 - 6.47 (m, 2H), 5.14 (s, 2H), 3.75 (s, 3H), 2.27 (s, 3H).

Step 2. Synthesis of 2-(4-benzyloxy)-2-methylphenyl)-5-methoxy-4H- chromen-4-one (compound 4df, Scheme 1).

The title compound was synthesized following general procedure B using intermediate 3df (224 mg, 0.59 mmol), I2 (7 mg, 0.03 mmol) in DMSO (1 mL). Purification by silica (elution by gradient from 100 to 20/80 Cyclohexane/EtOAc) gave the pure compound 4df (181 mg, 83% yield).

Characterization: UPLC/MS Rt: 1.39 min (gradient 1), MS (ESI) m/z: 373.1 [M+H] ⁺ .[M+H]+ Calculated for C ₂₄ H ₂₁ O ₄ : 373.1. ¹ H-NMR (400 MHz, DMSO-da) δ 7.67 (dd like t, J = 8.4 Hz, 1H), 7.55 (d, J = 8.6 Hz, 1H), 7.49 - 7.44 (m, 2H), 7.44 - 7.37 (m, 2H), 7.37 - 7.30 (m, 1H), 7.16 (dd, J = 8.5, 0.9 Hz, 1H), 7.05 (d, J = 2.6 Hz, 1H), 7.02 - 6.95 (m, 2H), 6.28 (s, 1H), 5.18 (s, 2H), 3.87 (s, 3H), 2.43 (s, 3H).

Step 3. Synthesis of 2-(4-hydroxy-2-methylphenyl)-5-methoxy-4H- chromen-4-one (compound 5df, Scheme 1).

The title compound 5df was prepared following general procedure D using: compound 4df (181 mg, 0.49 mmol), Pd/C (40 mg), Et ₃ SiH (0.5 mL, 2.35 mmol) in a 1:1 mixture of MeOH/DCM (10 mL). Purification by silica (elution by gradient from 100 to 0 cyclohexane/EtOAc) gave the pure intermediate 5df (69 mg, 50% yield).

Characterization: UPLC/MS Rt: 1.64 min (gradient 1), m/z: 283.1 [M+H] ⁺ .[M+H] ⁺ Calculated for C ₁₇ H ₁₅ O ₄ : 283.1. ¹ H-NMR (400 MHz, DMSO- d ₆ ) δ 7.66 (dd like t, J = 8.4 Hz, 1H), 7.44 (d, J = 8.3 Hz, 1H), 7.15 (dd, J = 8.4, 0.9 Hz, 1H), 6.98 (dd, J = 8.4, 0.9 Hz, 1H), 6.77 - 6.68 (m, 2H), 6.23 (s, 1H), 3.86 (s, 3H), 2.38 (s, 3H). Reference is now made to Scheme 2 in figure 3.

Synthesis of 2-(3,4-dihydroxyphenyl)-5,7-dihydroxy-3-(3- hydroxypropoxy)-4H-chromen-4-one (compound 9, Scheme 2).

Step 1. Synthesis of 7-(benzyloxy)-2-(3,4-bis(benzyloxy)phenyl)-5- hydroxy-3-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-

((((2R,3R,4R,5R,6S) -3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-

2-yl)oxy)methyl)tetrahydro-2H-pyran-2-yl)oxy)-4H-chromen- 4-one (compound 6, Scheme 2).

To a solution of rutin (2000 mg, 2.96 mmol) in DMF (20 mL) K ₂ CO ₃ (1716 mg, 12.44 mmol) and benzoyl bromide (2.8 mL, 23.68 mmol) were added. The reaction mixture was stirred overnight at room temperature. Then it was diluted with EtOAc (60 mL). The organic phase was divided, washed with water (2x50 mL), dried over magnesium sulfate, filtered and concentrated under vacuum giving the crude product 6, which was used without further purification for the next step (2000 mg).

Characterization: UPLC/MSRt: 2.44 min (gradient 1), MS (ESI) m/z: 881.3 [M+H] ⁺ .[M+H] ⁺ Calculated for C ₄₈ H ₄₉ O ₁₆ : 883.3.

Step 2. Synthesis of 7-(benzyloxy)-2-(3,4-bis(benzyloxy)phenyl)-

3,5-dihydroxy-4H-chromen-4-one (compound 7, Scheme 2).

Intermediate 6 (2000 mg, 2.27 mmol) was dissolved in EtOH (14 mL) to which 37% HC1 (2 mL) was added. The reaction mixture was refluxed for 120 minutes. After complete conversion of the starting material, the reaction mixture was cooled to room temperature and filtered. The precipitate was washed with water (5 mL) and cold MeOH (5 mL), giving the pure product 7 (1394 mg, yield: 82% in the two steps).

Characterization: UPLC/MS Rt: 2.55 min (gradient 2), MS (ESI) m/z: 573.2 [M+H] ⁺ .[M+H]+ Calculated for C ₃₆ H ₂₉ O ₇ : 573.2. 1H-NMR (400 MHz, DMSO-d ₆ ) δ 11.70 (s, 1H), 7.85 (d, J = 2.1 Hz, 1H), 777 (dd, J =

8.5, 2.0 Hz, 1H), 7.54 - 7.29 (m, 16H), 7.04 (d, J = 8.7 Hz, 1H),

6.52 (d, J = 2.2 Hz, 1H), 6.45 (d, J = 2.2 Hz, 1H), 5.25 (s, 4H),

5.15 (s, 2H). Step 3. Synthesis of 7-(benzyloxy)-2-(3,4-bis(benzyloxy)phenyl)-5- hydroxy-3-(3-hydroxypropoxy)-4H-chromen-4-one (compound 8, Scheme

2). K ₂ CO ₃ (248 mg, 0.18 mmol) and 3-bromo-1-propanol (0.14 mL, 0.9 mmol) were added to a solution of intermediate 8 (350 mg, 0.60 mmol) in DMF (10 mL) and the reaction mixture was stirred under argon atmosphere for three hours at room temperature. After reaction completion, the reaction mixture was poured into water and extracted with ethyl acetate (2x20 mL). The combined organic layer was divided, dried over magnesium sulfate, filtered and concentrated under vacuum, giving the crude product 8, which was used for the next step without purification (365 mg).

Characterization: UPLC/MS Rt: 2.40 min (gradient 2), MS (ESI) m/z: 631.2 [M+H] ⁺ .[M+H] ⁺ Calculated: 631.2 for C ₄₁ H ₃₇ O ₈ .

Step 4. Synthesis of 2-(3,4-dihydroxyphenyl)-5,7-dihydroxy-3-(3- hydroxypropoxy)-4H-chromen-4-one (compound 9, Scheme 2). Compound 9 was prepared following general procedure D using: compound 8 (365 mg, 0.58 mmol), Pd/C (72 mg), Et ₃ SiH (2.2 mL, 10.5 mmol) in a 1:1 mixture of MeOH/DCM (14 mL). Purification by silica (elution by gradient from 100 to 40/60 Cyclohexane/EtOAc) gave the pure compound 9 (33 mg, 15% yield in two steps).

Characterization: Rt: 1.40 min (gradient 1), MS (ESI) m/z: 361.1 [M+H] ⁺ .[M+H] ⁺ Calculated for C ₁₉ H ₁₉ O ₇ : 361.1.

¹ H-NMR (400 MHz, DMSO-d ₆ ) δ 12.72 (s, 1H), 7.53 (d, J = 2.2 Hz, 1H), 7.46 (dd, J = 8.4, 2.2 Hz, 1H), 6.89 (d, J = 8.4 Hz, 1H), 6.39 (d, J = 2.1 Hz, 1H), 6.18 (d, J = 2.1 Hz, 1H), 4.01 (t, J = 6.7 Hz, 2H), 3.49 (t, J = 6.4 Hz, 2H), 1.81 (p, J = 6.5 Hz, 2H). 13C-NMR (101 MHz, DMSO-d ₆ ) 5 178.04, 164.12, 161.29, 156.37,

155.95, 148.62, 145.18, 136.75, 120.96, 120.78, 115.64, 115.51,

104.17, 98.54, 93.57, 69.72, 57.71, 32.91.

Reference is now made to Scheme 3.

Synthesis of N-(3,4-dihydroxyphenyl)-5-hydroxy-4-oxo-4H-chromen-2- carboxamide (compound 13, Scheme 3).

Step 1. Synthesis of ethyl 5-methoxy-4-oxo-4H-chromen-2- carboxylate (compound 10).

NaOEt (210 mg, 3 mmol) was dissolved in absolute EtOH 4 mL). A mixture of diethyl oxalate (310 mg, 2.1 mmol) and 2-hydroxy-6- methoxyacetophenone lc (100 mg, 0.6 mmol) in absolute EtOH (2 mL) was slowly added to the NaOEt solution. The solution was reflux- stirred for 2 hours until complete consumption of starting material. The mixture was then allowed to cool to room temperature and neutralized with acq. HCl (2 M). The mixture was extracted with EtOAc (3 x 5 mL), the collected organic layer was dried over Na ₂ SO ₄ , filtered and concentrated under vacuum. Purification by silica (elution by gradient from 100:0 to 75:25 Cyclohexane/EtOAc) gave the pure title compound 10 (120 mg, 79%).

Characterization: UPLC/MS Rt: 1.50 min (gradient 1), MS (ESI) m/z 249.0, [M+H] ⁺ . [M+H] ⁺ calculated for C ₁₇ H ₁₅ O ₅ : 249.3. 1H NMR (400

MHz, CDCI ₃ ) δ 7.40 (dd, J = 8.4, 8.4 Hz, 1H), 6.60 - 6.58 (m, 2H), 6.42 (d, J = 8.4 Hz, 1H), 4. 38 (q, J = 7.2 Hz, 2H), 3.93 (s, 3H), 1.34 (t, J = 7.2 Hz, 3H).

Step 2. Synthesis of 5-methoxy-4-oxo-4H-chromen-2-carboxylic acid (compound 11). K ₂ CO ₃ (100 mg, 0.60 mmol) was added to a solution of intermediate 10 (100 mg, 0.40 mmol) in a 3:1 mixture of THF/EtOH (4 mL). The reaction mixture was stirred for 6 h at 50°C until complete consumption of the starting material, then acq. HC1 (2 M) was added up to reaching pH=5, and the mixture was extracted with EtOAc (3x4 mL). The collected organic layers were dried over Na ₂ SO ₄ , filtered and concentrated under vacuum to give the pure compound 11 (70 mg, 79% yield). Characterization: UPLC/MS Rt: 0.80 min (gradient 1), MS (ESI) m/z 220.9, [M+H] ⁺ . [M+H] ⁺ calculated for C ₁₁ H ₉ O ₅ : 221.2. NMR (400 MHz, DMSO-d ₆ ) δ 7.72 (dd, J = 8.4, 8.4 Hz, 1H), 7.18 (d, J = 8.4 Hz, 1H), 7.02 (d, J = 8.3 Hz, 1H), 3.86 (s, 3H).

Step 3. Synthesis of N-(3,4-dimethoxyphenyl)-5-methoxy-4-oxo-4H- chromen-2-carboxamide (compound 12).

HATU (170 mg, 0.43 mmol) and DIPEA (0.23 mL, 1.3 mmol) were sequentially added to a solution of intermediate 11 (60 mg, 0.29 mmol) in a 3:1 mixture of DMF/DCM (4 mL) under argon. The reaction mixture was stirred at room temperature for 15 minutes, after that 3,4-dimethoxyaniline (44 mg, 0.29 mmol) was added and the reaction mixture was stirred for another 4 hours until complete consumption of the starting material. Water (1 mL) and acq. HCl (2 M) were added to reach pH=7, the mixture was extracted with CH ₂ CI ₂ (3 x 3 mL), the collected organic layers were dried over Na ₂ SO ₄ , filtered and concentrated under vacuum. Purification by silica (elution by gradient from 100:0 to 55:45 DCM/EtOAc) gave the pure title compound 12 (92 mg, 90%).

Characterization: UPLC/MS Rt: 1.66 min (gradient 1), MS (ESI) m/z 356.0, [M+H] ⁺ . [M+H] ⁺ calculated for C ₁₉ H ₁₈ NO ₆ : 356.3. NMR (400 MHz, DMSO-d ₆ ) δ 7.78 (dd, J = 8.4, 8.4 Hz, 1H), 7.44 (d, J = 2.4 Hz, 1H), 7.36 (dd, J = 8.6, 2.4 Hz, 1H), 7.34 (d, J = 8.3 Hz, 1H), 7.05 (d, J = 8.3 Hz, 1H), 6.98 (d, J = 8.7 Hz, 1H), 6.76 (s, 1H),

3.88 (s, 3H), 3.77 (s, 3H), 3.76 (s, 3H).

Step 4, Synthesis of N-(3,4-dihydroxyphenyl)-5-hydroxy-4-oxo-4H- chromen-2-carboxamide (compound 13).

Compound 13 was prepared following general procedure C, method A using: intermediate 12 (45 mg, 0.13 mmol), BBrs (1 M in DCM) (0.59 mL, 0.59 mmol) in anhydrous CH2CI2 (2.6 mL). The crude product was purified by silica (elution by gradient from 100:0 to 98:2 DCM/MeOH) to give product 13 (0.016 g, 40% yield).

Characterization: UPLC/MS Rt: 1.65 min (gradient 1), m/z 312.0,

[M-H]-. [M-H]-. calculated for C ₁₆ H ₁₀ NO ₆ : 312.3. ¹ H NMR (400 MHz,

DMSO-d ₆ ) δ 12.30 (bs, 1H), 10.48 (bs, 1H), 9.01 (bs, 1H), 7.77 (dd, J = 8.4, 8.4 Hz, 1H), 7.30 (d, J = 2.5 Hz, 1H), 7.26 (d, J = 8.4 Hz, 1H), 7.03 (dd, J = 8.5, 2.5 Hz, 1H), 6.95 (s, 1H), 6.88 (d, J = 8.3 Hz, 1H), 6.74 (d, J = 8.5 Hz, 1H).

Activity measurement

Pol η activity was assessed by quantification of the final product of DNA synthesized with the enzyme in the presence of all four dNTPs/nucleotides. The double strand of DNA used in the reaction was created by annealing an IRD700-labeled primer to a template strand. A mixture of DNA template and Pol η was placed on ice in an assay buffer solution and then added in test tubes containing the inhibitors or DMSO (as vehicle; the final concentration in the assay is 1%). The reaction was started with the addition of 5 mM Mg ²⁺ and immediately transferred to 37°C. After 60 minutes of incubation, the reaction was stopped with the addition of 5X loading buffer for the sample and incubation for 5 minutes at 70°C. The Pol η extension products were then separated by denaturing electrophoresis (15% polyacrylamide/tbe/urea gel; BioRad) and scanned using the ChemiDoc imaging system (BioRad Laboratories).

Cell viability assay

Compound 9 was evaluated for the inherent antiproliferative activity thereof, individually and together with cisplatin on the cancer cell lines A549, A375 and OVCAR3. The antiproliferative activity of compound 9 was evaluated using the MTT cell viability assay. To assess antiproliferative activity, the cells were seeded at a density of 5000 cells/well (A549), 3000 cells/well (A375) and 10000 cells/well (OVCAR3). After 24 hours, the cells were first treated with Cisplatin (0.4-50 μM) alone and compound 9 (0.046-100 μM), then with Cisplatin (0.4-50 μM) with compound 9 at 50 μM, 75 μM and 100 μM for 48 hours. The MTT solution was added to a final concentration of 0.5 mg/ml and the cells were further incubated for 4 hours. The insoluble formazan crystals were solubilized by the addition of a 10% 0.01 N SDS/HC1 solution and the absorbance measured at 570 nm (reference 690 nm) in a plate reader (Tecan Spark). The inhibition curves were 8 serial dilutions in triplicate in each case, and the results were analyzed as sigmoidal dose-response curves using GraphPad Prism software. The values are reported as meaniSD of three experiments. a The concentration of compound 9 is 100 μM b The concentration of compound 9 is 50 μM c The concentration of compound 9 is 75 μM

From the above, the advantages linked to the present invention will be immediately apparent to those skilled in the art.

Firstly, the compounds described have been shown to be very active in inhibiting human polymerase eta (Pol η) representing a potential tool for the therapy of chemo-resistant tumors in combination with current therapies, for example comprising the use of Cis platinum.

The data obtained show the effectiveness in inhibiting the proliferation of cancer cells and the low cell toxicity.

Furthermore, data also show that the stability and solubility values are favorable for medical use.

The main structure and substructures define scaffolds which allow a good chemical diversity, allowing many compounds with similar structure to be investigated.

In addition to this, the compounds described are referred to as small molecules, thus being molecules of small size capable of being easily synthesized.

It is worth to emphasize that the present invention contemplates already known compounds, and therefore it applies to the so-called "repurposing" of drugs, thus making the evaluations of activity and toxicity of a possible drug for the new identified disease faster.