WO2025008012A1 - Pyrroloquinolines and their use as tubuline assembly inhibitors - Google Patents

Abstract

This invention concerns 1H-pyrrolo[3,2-c]quinoline compounds of formula (I) as inhibitors of tubuline polymerization. The invention also relates to the uses of such compounds and compositions, particularly their use in treating cancer. Formula (I)

Description

Pyrroloquinolines and their use as tubuline assembly inhibitors Field of the Invention The invention relates to 1H-pyrrolo[3,2-c]quinoline derivatives acting as tubuline assembly inhibitors and to their use in the treatment of cancer. Background Art Microtubules are cytoskeletal polymers of tubulin. They consist of α- and β-tubulin heterodimers which represent one of the most strategic molecular targets among the current targets for anticancer chemotherapy. Microtubules play a critical role in a number of cellular functions, such as chromosome segregation during cell division, intracellular transport, cell motility, and the maintenance of cell shape. According to a general mode of action, microtubule targeting agents (MTAs) are classified into microtubule destabilizing and microtubule stabilizing agents. Both of them inhibit the dynamic instability of tubulin polymerization. Based on their different binding sites on microtubule protein, MTAs are further classified into six types: colchicine, vinca alkaloids, taxanes, laulimalide, maytansine, and pironetin site-binding agents. Majority of MTAs reported to date are tubulin polymerization inhibitors which can decrease the mass of microtubules, whereas microtubule stabilizers promote the growth of microtubules. Additionally, MTAs cause vascular disruption, and target the tumor vasculature leading to death of cancer cells that are resistant to conventional chemotherapy and radiotherapy. Similarly to other drug classes, the clinical outcome of microtubule-targeted chemotherapy is often limited by drug resistance. The resistance can stem from a variety of mechanisms including altered expression of drug efflux pumps, expression of tubulin isoforms such as βIII-tubulin isoform, alterations in regulatory post-translational modifications or microtubule-regulatory proteins. P-glycoprotein (MDR1), multidrug resistance-associate protein 1 (MRP1) and breast cancer resistance protein (BCRP) are the three drug efflux pumps that can decrease intracellular concentration of drugs. Expression of MDR1 was shown to limit the response to paclitaxel and vinca alcaloids. Aberrant overexpression of βIII-tubulin isoform is a common feature of aggressive and treatment refractory cancers. Microtubule- associated proteins such as Microtubule Associated Protein 4 MAP4 or stathmin can also modulate microtubule dynamics and response to MTAs. Disclosure of the invention The invention relates to a compound of general formula I:

or an N-oxide, a pharmaceutically acceptable salt, a hydrate or a solvate thereof, wherein: R₁ is selected from F, Cl, Br, I, OH, H and C1-C3 alkyl; X is selected from C1-C2 alkylenyl, -C(=O)- and -CH₂-C(=O)- linker; R₂ is selected from phenyl and 5-8 membered heteroaryl containing at least one heteroatom selected from N, O, S; wherein said phenyl or heteroaryl is unsubstituted or substituted by at least one substituent selected from hydroxy, F, Cl, Br, I, C1-C4 alkoxy, hydroxy-C1-C4-alkoxy, halogen- C1-C4-alkoxy, amino-C1-C5-alkoxy, C1-C4 alkylamino, hydroxy-C1-C4-alkylamino, halogen- C1-C4-alkylamino, amino-C1-C4-alkylamino. The term “alkyl” as used herein denotes a univalent saturated branched or linear hydrocarbon chain, having the number of carbon atoms within the indicated range. Representative examples of alkyl groups are methyl, ethyl, n-propyl, isopropyl, butyl, isobutyl, tert-butyl. The term “alkylenyl” denotes a bivalent saturated linear hydrocarbon chain, having the number of carbon atoms within the indicated range. Alkylenyl is, for example, methylenyl (-CH₂-) or ethylenyl (- CH₂-CH₂-). The term “alkoxy” refers to the group alkyl-O-, wherein alkyl is as defined herein above. The term “alkylamino” refers to the group alkyl-NH-, wherein alkyl is as defined herein above. The term “halogen” includes F, Cl, Br, I. The term “heteroaryl” refers to monocyclic or bicyclic aromatic group, containing at least one heteroatom selected from N, O, S. Examples of heteroaryl group include furyl, thienyl, pyrroryl, oxazolyl, thiazolyl, imidazolyl, triazolyl, isoxazolyl, isothiazolyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, 1,3,5-triazinyl. The term “substituted” means that the specified group or moiety bears one or more substituents, preferably one substituent. Where any group may carry multiple substituents, and a variety of possible substituents is provided, the substituents are independently selected, and do not need to be the same. The term “unsubstituted” means that the specified group bears no substituents others than H atoms. The term “N-oxide” refers to N-oxide form of a compound of general formula I. The N-oxide group may form on the quinoline ring nitrogen atom. The term “pharmaceutically acceptable salt” refers to salts of a compound of general formula I with pharmaceutically acceptable cations or anions. Pharmaceutically acceptable salts include also addition salts with acids. Suitable acids are those forming pharmaceutically acceptable anions. Examples of pharmaceutically acceptable cations and anions include: Cations: aluminum, arginine, benzathine, calcium, chloroprocaine, choline, diethanolamine, ethanolamine, ethylenediamine, lysine, magnesium, histidine, lithium, meglumine, potassium, procaine, sodium, triethylamine, zinc. Anions: acetate, aspartate, benzenesulfonate, benzoate, besylate, bicarbonate, bitartrate, bromide, camsylate, carbonate, chloride, citrate, decanoate, edetate, esylate, fumarate, gluceptate, gluconate, glutamate, glycolate, hexanoate, hydroxynaphthoate, iodide, isethionate, lactate, lactobionate, malate, maleate, mandelate, mesylate, methylsulfate, mucate, napsylate, nitrate, octanoate, oleate, pamoate, pantothenate, phosphate, polygalacturonate, propionate, salicylate, stearate, succinate, sulfate, tartrate, teoclate, tosylate. The term “hydrate” refers to solvate of a compound of general formula I with water. In a preferred embodiment, the compound of general formula I is a compound of general formula II:

or an N-oxide, a pharmaceutically acceptable salt, a hydrate or a solvate thereof, wherein: R₁ is selected from H, F, Cl, Br, I, OH and C1-C3 alkyl; X is selected from C1-C2 alkylenyl, -C(=O)- and -CH₂-C(=O)- linker; R₃ represents one or more substituents which are independently selected from hydroxy, F, Cl, Br, I, C1-C4 alkoxy, hydroxy-C1-C4-alkoxy, halogen-C1-C4-alkoxy, amino-C1-C5-alkoxy, C1-C4 alkylamino, hydroxy-C1-C4-alkylamino, halogen-C1-C4-alkylamino, amino-C1-C4-alkylamino. More preferably, R₃ is selected from 3-methoxy, 4-methoxy and 4-ethoxy, or R₃ represents two substituents wherein one substituent is selected from 3-methoxy, 4-methoxy and 4-ethoxy and one substituent is selected from halogen (preferably fluoro). More preferably, X is selected from methylenyl (-CH₂-) or ethylenyl (-CH₂CH₂-). More preferably, R₁ is Cl or Br. The compounds of general formula I act as inhibitors of tubuline assembly and are useful in treating cancer. In particular, the compounds of general formula I are useful in treatment of cancers selected from leukaemia, lung adenocarcinoma, colorectal cancer and osteosarcoma. The term ”treatment” as used herein refers to any treatment a mammalian, for example human, disease, and includes: (i) inhibiting the disease, i.e., arresting its development, (ii) relieving the disease, i.e.; causing the condition to regress, or (iii) stopping the symptoms of the disease. In another aspect of the invention, a pharmaceutical composition is provided which contains at least one compound of general formula I and at least one pharmaceutically acceptable excipient. Pharmaceutically acceptable excipients include, in particular, fillers, binders, lubricants, solvents, dispersing agents and preservatives. Suitable excipients for any desired form of a pharmaceutical composition are known to a person skilled in the art of pharmaceutical formulations. The term “composition” as used herein encompasses a product comprising specified ingredients. In relation to pharmaceutical compositions, this term encompasses a product comprising one or more active ingredients, and at least one pharmaceutically acceptable excipient. In general, pharmaceutical compositions are prepared by uniformly and intimately bringing the active ingredients into association with said pharmaceutically acceptable excipient(s) and then, if necessary, shaping the product into desired formulation. The pharmaceutical composition includes enough of the active object compound to produce the desired therapeutic effect. Accordingly, the pharmaceutical compositions of the present invention encompass any composition made by admixing a compound of the present invention and at least one pharmaceutically acceptable excipient. By “pharmaceutically acceptable” it is meant the excipient must be compatible with the other ingredients of the formulation and not deleterious to the recipient thereof. The typical daily dose of the active ingredient varies within a wide range and will depend on various factors such as the relevant indication, the route of administration, the age, weight and sex of the patient and may be determined by a physician. In general, total daily dose administration to a patient a single or individual doses, may be in amounts, for example from 0.001 to 10 mg/kg body weight daily, and more usually 0.01 to 1000 mg per day, of total active ingredients. Generally, compounds described in the scope of this patent application can be synthesized by the route described in Scheme 1 and further illustrated in the Examples. Scheme 1 Compounds of formula (I) can be prepared by the following reactions: (i) Condensation of 2-nitrobenzaldehyde I and p-toluenesulfonamide II using pyrrolidine, anhydrous CH₂Cl₂ and molecular sieves 4 A°; (ii) Reaction of imine III with ethyl buta-2,3-dienoate IV with triphenylphosphine in anhydrous CH₂Cl₂; (iii) Reaction of N-tosyl 2,5-dihydro-1H-pyrrole-3-carboxylate V with tBuOK in N,N- dimethylformamide; (iv) Catalytic hydrogenation of intermediate VI using palladium on carbon (Pd/C) and ethanol as a solvent and simultaneous cyclization in a presence of catalytic amount of acetic acid; (v) Chlorination of lactam VII using POCl₃ to yield the intermediate VIII; (vi) Alkylation or acylation of the intermediate VIII with alkyl halides or acyl chlorides to receive intermediate IX, optionally followed by demethylation of methoxy groups to hydroxyl analogues; (vii) Further modifications using nucleophiles such as halides or MeMgI to obtain compounds X. Brief description of Drawings Figure 1: Results of the tubulin binding assay for compound IXd. Effect of compounds on the inhibition of the bisthioalkylation of Cys-239 and Cys-354 of β-tubulin by N,N'-ethylene-bis(iodoacetamide). IXd occupies colchicine binding site at β-tubulin. CCRF-CEM cells were pre-treated with 50 μmol/L of IXd, nocodazole (NOC), colchicine (COL) and vincristine (VCR) or corresponding volume of DMSO and subjected to 100 μmol/L EBI. MW of β-tubulin is 55 kDa/EBI:β-tubulin mobility shift to 50 kDa. Examples of carrying out the Invention Methods and materials: Solvents and chemicals were purchased from Sigma-Aldrich (Milwaukee, WI, www.sigmaaldrich.com), Acros Organic (Geel, Belgium, www.acros.com) and Fluorochem (Hadfield, United Kingdom, www.fluorochem.co.uk). Anhydrous solvents were dried over 4 Å molecular sieves or stored as received from commercial suppliers. Reactions were performed in round-bottom flasks fitted with rubber septa under positive pressure of nitrogen or in pressure ampoules, unless stated otherwise. Reactions were monitored by LC-MS analysis or by thin-layer chromatography (TLC) using aluminum plates pre-coated with silica gel (silica gel 60 F₂₅₄, Merck, USA) impregnated with a fluorescent indicator. TLC plates were visualized by exposure to ultraviolet light (λ = 254 nm) and/or by submersion in aqueous ceric ammonium molybdate (CAM) solution and/or ninhydrin solution followed by brief heating. The LC-MS analyses were carried out on UHPLC-MS system consisting of UHPLC chromatograph Acuity with photodiode array detector and single quadrupole mass spectrometer (Waters), using X-Select C18 column with the mobile phase consisting of 10 mM ammonium acetate (AmAc) in H₂O and CH₃CN. The ESI source operated at discharge current of 5 μA, vaporizer temperature of 350 °C and capillary temperature of 200 ^ºC. For the LC-MS analysis, samples were extracted into CH₃CN/H₂O (20% or 50%; 1 mL). Purification was carried out on silica gel chromatography, on semipreparative reverse-phase (RP)-HPLC chromatography on C18 reverse phase column (Phenomenex Gemini, 50 × 2.00 mm, 3 µm particles, C18) with gradient and/or isocratic of 10 mM aqueous ammonium acetate (AmAc) buffer and CH₃CN, after direct infusion of samples into the mass spectrometer using autosampler, flow rate 15 mL/min and/or on flash column chromatography (FCC) using silica gel (60 Å, 230–400 mesh, Sigma-Aldrich, Milwaukee, WI, www.sigmaaldrich.com) after direct infusion of samples into the column, flow rate 40 mL/min. Residual solvents (H₂O, AmAc, DMSO etc.) were lyophilized by the ScanVac Coolsafe 110- 4 working at -110 °C. All 1D NMR experiments were performed with using Varian BB 200 (Varian Inc., California, USA) and/or ECX500 spectrometer (JEOL RESONANCE, Tokyo, Japan) at magnetic field strengths of 7.05 T and 11.75 T corresponding to ¹H, ¹³C and ¹⁹F resonance frequencies of 300.01 MHz (for ¹H) and 500.16 MHz (for ¹H), 125.77 MHz (for ¹³C) and 470.62 MHz (for ¹⁹F) at 26-27 ºC. Chemical shifts (δ) are reported in parts per million (ppm) and coupling constants (J) are reported in Hertz (Hz). The signals of CDCl₃, MeOH-d₄, DMSO-d₆ and THF-d₈ were set at 7.26 ppm, 3.31 ppm, 2.50 ppm and 1.73 ppm in ¹H NMR spectra and at 77.0 ppm, 49.0 ppm, 39.5 ppm and 25.3 ppm in ¹³C NMR spectra, respectively. The residual signal of AmAc (from HPLC purification) exhibited one resonance at 1.90 ppm in ¹H NMR spectra and at two resonances at 21.3 ppm and 172.0 ppm in ¹³C NMR spectra. Abbreviations in NMR spectra: br. s – broad singlets – singlet, d – doublet, dd – doublet of doublets, ddd – doublet of doublets of doublets, dddd – doublets of doublets of doublets of doublets, t – triplet, td – triplet of doublets, m – multiplet. HRMS analysis was performed using LC-MS (Dionex Ultimate 3000, Thermo Fischer Scientific, MA, USA) with Exactive Plus Orbitrap high-resolution mass spectrometer (Thermo Exactive plus, Thermo Fischer Scientific, MA, USA) operating at positive or negative full scan mode (120000 FWMH) in the range of 100-1000 m/z with electrospray ionization working at 150 °C and the source voltage of 3.6 kV. Chromatographic separation was performed on C18 column (Phenomenex Gemini, 50 x 2 mm, 3 µm particle) with isocratic elution and mobile phase (MP) containing CH₃CN/10 mM AmAc (80/20; v/v). The samples were dissolved in CH₃CN or CH₃CN/H₂O (1/1; v/v). Melting points (not corrected) were measured by Thermovar apparatus (Reichert, Vienna, Austria) and reported in Celsius degree (°C). Cytotoxic activity was screened against representative cancer cell lines: CCRF-CEM (T-lymphoblastic leukaemia), CEM-DNR (T-lymphoblastic leukaemia, daunorubicin resistant), K562 (acute myeloid leukaemia), K562-TAX (acute myeloid leukaemia, paclitaxel resistant), A549 (human lung adenocarcinoma), HCT116 (human colorectal cancer), HCT116p53-/-, (human colorectal cancer, p53 deficient), U2OS (human osteosarcoma). The therapeutic index (TI) was evaluated using one representative of non-malignant cell lines BJ (human fibroblasts). General synthetic procedures and analytical data of prepared derivatives: 1. Synthesis of 4-methyl-N-(2-nitrobenzylidene)benzenesulfonamide III. To a suspension of p- toluenesulfonamide 2 (4.0 g, 23.360 mmol, 1 equiv) in extra anhydrous CH₂Cl₂ (60 mL) containing pyrrolidine (215 µL, 2.570 mmol, 0.11 equiv) and 4 Å molecular sieves (23.36 g), 2-nitrobenzaldehyde I (2.82 g, 18.688 mmol, 0.8 equiv) was added. The reaction mixture was stirred at reflux for 4 h under Dean-Stark conditions. The reaction was monitored by TLC. Then the reaction was filtered through Celite, and the residue was washed with extra anhydrous CH₂Cl₂. The organic fraction was evaporated to dryness in vacuo and the crude product was precipitated twice from EtOAc/hexane (3/7; v/v) to give the imine III as pale yellow solid (4.47 g, 14.7 mmol, 66%). 2. [3+2] Annulation-catalyzed triphenylphosphine (TPP) to ethyl 2-(2-nitrophenyl)-1-tosyl-2,5-dihydro- 1H-pyrrole-3-carboxylate V. To the intensively stirred solution of imine III (4.47 g, 14.704 mmol, 1 equiv) and TPP (387 mg, 1.470 mmol, 0.1 equiv) in anhydrous CH₂Cl₂ (93 mL) cooled at -10 °C, and then ethyl 2,3-butadienoate IV (12.048 mL, 17.645 mmol, 1.2 equiv) was added. The reaction mixture was stirred at -10 °C to room temperature overnight and monitored by TLC. After completion, the solvent was evaporated in vacuo and the crude product was purified by silica gel chromatography in EtOAc/hexane (4/6; v/v) to give the product V as yellow solid (5.6 g, 13.46 mmol, 92%), HPLC purity 99%. 3. Synthesis of ethyl 2-(2-nitrophenyl)-1H-pyrrole-3-carboxylate VI. The compound was prepared by literature procedure.¹ To a solution of 2,5-dihydro-1-tosyl-pyrrole 5 (5.6 g, 13.462 mmol, 1 equiv) in DMF (89.5 mL), potassium tert-butoxide (4.53 g, 40.38 mmol, 3 equiv) was added. The reaction was stirred for 2 h at room temperature and monitored by TLC or LC-MS. The resulting mixture was diluted with CH₂Cl₂ (27 mL), washed three times with a saturated solution of NH₄Cl (27 mL). Then the organic layer was washed three times with a saturated solution of NaHCO₃, twice with water and once with brine. The organic fractions were collected and dried using Na₂SO₄, filtered and evaporated to dryness in vacuo. Crude product was purified by silica gel chromatography in EtOAc/hexane (4/6; v/v) to give the pyrrole VI as orange oil (2.55 g, 9.79 mmol, 73%), HPLC purity 99%. 4. Catalytical hydrogenation and spontaneous cyclization to 1,5-dihydro-4H-pyrrolo[3,2-c]quinolin-4- one 7. To the nitro-derivative VI (2.55 g, 9.792 mmol, 1 equiv) dissolved in anhydrous EtOH (83 mL), 10% Pd/C (254.6 mg, 10 wt%) and AcOH (252 µL, 4.406 mmol, 0.45 equiv) were added under nitrogen atmosphere. The reaction mixture was stirred under a hydrogen atmosphere overnight at atmospheric pressure and monitored by TLC with ninhydrin visualization. After completion, the reaction mixture was filtered through Celite, washed with EtOH and redistilled five times with Et₂O to remove AcOH residue to give the lactam VI as brown solid (1.77 g, 9.6 mmol, 98%), HPLC purity 99%. 5. Synthesis of 4-chloro-1H-pyrrolo[3,2-c]quinoline VIII. The mixture of the lactam 7 (1.80 g, 9.792 mmol, 1 equiv) and POCl₃ (25 mL) was heated at 105 °C for 5 h in pressure ampoule. The reaction was monitored by TLC. After completion, the reaction mixture was evaporated under stream of nitrogen and ice (39.4 g) was added. After melting, the product was either filtered and dried in vacuo, or extracted three times with CH₂Cl₂, the collected fractions were dried over MgSO₄, filtered and evaporated to dryness in vacuo. Crude product was crystallized from EtOAc to give the product VIII as creme solid (1.83 g, 9.06 mmol, 92%), HPLC purity 99%. 6. General procedures for N-alkylation (products IX). Method A: To a solution of 4-chloro-1H- pyrrolo[3,2-c]quinoline VII, 4-chloro-1H-pyrrolo[3,2-c]pyridine Xa–b (1 equiv) dissolved in extra anhydrous DMSO/CH₃CN or DMSO (2 mL/0.248 mmol of the starting material; 1/1; v/v), BTPP (6 equiv) was added and stirred for 20-30 min at room temperature. Then alkylating or acylating reagent (2 or 6 equiv) was slowly added dropwise and heated up at 90 °C for 3 h-5 days or at 50 °C for 1 h. The reaction was monitored by LC-MS or TLC analysis. After completion, the reaction mixture was cooled down to room temperature, diluted with aqueous NH₄Cl (100 mL/0.248 mmol of the starting material) and extracted four times with CH₂Cl₂ (70 mL/0.248 mmol of the starting material). The combined organic layers were dried using MgSO₄, filtered and evaporated to dryness in vacuo. The crude products were purified by silica gel chromatography or FCC chromatography or semipreparative RP-HPLC chromatography or crystallized from appropriate solvent. Method B: To a solution of 4-chloro-1H-pyrrolo[3,2-c]quinoline VIII (1 equiv) dissolved in extra anhydrous CH₂Cl₂ (2 mL/0.248 mmol of the starting material), BTPP (2,5 equiv) was added and stirred for 30 min at room temperature. Then phenethyl bromide (3,5 equiv) was slowly added dropwise and refluxed at 40 °C for 22 h in a pressure ampoule. The reaction was monitored by LC-MS or TLC analysis. After completion, the reaction mixture was cooled down to room temperature, diluted with aqueous NH₄Cl (100 mL/0.248 mmol of the starting material) and extracted four times with CH₂Cl₂ (70 mL/0.248 mmol of the starting material). The combined organic layers were dried using MgSO₄, filtered and evaporated to dryness in vacuo. The crude products were purified by silica gel chromatography or FCC chromatography or semipreparative RP-HPLC chromatography or crystallized from appropriate solvent. urther modification of compounds IX to products X a) To a solution of IXd (25 mg, 0.074 mmol, 1 equiv) in anhydrous and properly nitrogen flushed isopropanol (2 mL), triphenyl phosphite (40 µL, 0.148 mmol, 2 equiv), tBuOK (36 mg, 0.333 mmol, 4.5 equiv) and Pd₂dba₃ (20 mg, 0.022 mmol, 30 mol%) were added at room temperature and the resulting mixture was heated up to 80 °C for 24 h. The reaction was monitored by LC- MS. After completion, the reaction mixture was cooled down to room temperature, diluted with aqueous NH₄Cl (40 mL) and extracted four times with EtOAc (40 mL). The combined organic extracts were dried using MgSO₄, filtered and evaporated to dryness in vacuo. The crude product was purified by semipreparative RP-HPLC chromatography in isocratic gradient containing CH₃CN/10 mM AmAc in water (4/1; v/v) to give the product as colorless oil (6.0 mg, 0.020 mmol, 27%), HPLC purity 99%. b) To a solution of IXd (25 mg, 0.074 mmol, 1 equiv) in anhydrous DMSO (1.5 mL), CsF (112 mg, 0.740 mmol, 10 equiv) was added at ambient temperature. The resulting mixture was heated up to 130 °C for 36 h. The reaction was monitored by LC-MS. Then, the reaction mixture was cooled down to ambient temperature, diluted with CH₂Cl₂ (70 mL) and extracted four times with aqueous NH₄Cl (100 mL). The combined organic extracts were dried using MgSO₄, filtered and evaporated to dryness in vacuo. The crude product was purified by semipreparative RP- HPLC chromatography in isocratic gradient containing CH₃CN/10 mM AmAc in water (4/1; v/v) to give the product as white solid (10.0 mg, 0.031 mmol, 42%), HPLC purity 99%. c) To a solution of IXd (38 mg, 0.113 mmol, 1 equiv) in anhydrous CH₃CN (2 mL), bromotrimethylsilane (TMBS; 195 µL, 1.47 mmol, 13 equiv) was added at room temperature. The resulting mixture was heated up to 90 °C for 20 h. The reaction was monitored by LC-MS. After completion, the reaction mixture was cooled down to room temperature, residual solvents were evaporated to dryness and the crude product was purified by FCC in CH₂Cl₂/EtOAc (1:1; v/v) and semipreparative RP-HPLC chromatography in isocratic gradient containing CH₃CN/10 mM AmAc in water (4/1; v/v) to give the product as white solid (10.0 mg, 0.026 mmol, 24%), HPLC purity 99%. d) To a solution of IXd (46 mg, 0.137 mmol, 1 equiv) in anhydrous CH₃CN (1 mL), NaI (177 mg, 1.374 mmol, 10 equiv) and acetyl chloride (132 µL, 1.856 mmol, 13.5 equiv) were added. The reaction mixture was refluxed for 24 h under nitrogen atmosphere and monitored by TLC. Then the reaction was quenched with CH₂Cl₂ (20 mL) and washed three times with a solution of 10% K₂CO₃/10% Na₂S₂O₃ (20 mL, 1/1; v/v). The combined organic fractions were dried using Na₂SO₄, filtered and evaporated to dryness in vacuo. The crude product was purified by semipreparative RP-HPLC chromatography in CH₃CN/10 mM AmAc in water (7/3; v/v) to give the product as white solid (5.7 mg, 0.013 mmol, 10%), HPLC purity 99%. e) A flame-dried flask was charged under nitrogen with IXd (50 mg, 0.149 mmol, 1 equiv), Fe(acac)₃ (28 mg, 0.08 mmol, 0.53 equiv), and anhydrous THF/NMP (3.5 mL, 10:1; v/v). A solution of methyl magnesium iodide (3.0M in Et₂O, 160 µL, 0.480 mmol, 3.2 equiv) was added via syringe to the solution. The resulting mixture was stirred for 30 min at room temperature under nitrogen atmosphere. Then, the reaction mixture was diluted with Et₂O (20 mL) and carefully quenched upon addition of saturated solution of NH₄Cl (10 mL). The organic layer was washed with water (420 mL) and with brine (210 mL), dried using Na₂SO₄, filtered and evaporated to dryness in vacuo. The crude product was crystallized from EtOAc/hexane to give the product as yellow-brown solid (25.3 mg, 0.080 mmol, 53%), HPLC purity 98%. Example 1: 4-chloro-1-phenethyl-1H-pyrrolo[3,2-c]quinoline IXa.

Cream solid (22.7 mg, 0.074 mmol, 12%), HPLC purity 99%. ¹H NMR (500 MHz, CDCl₃): δ = 8.18- 8.31 (m, 2H), 7.62-7.66 (m, 2H), 7.27-7.30 (m, 3H), 7.06-7.09 (m, 2H), 6.86 (d, J = 3.1 Hz, 1H), 6.70 (d, J = 3.1 Hz, 1H), 4.78 (t, J = 7.3 Hz, 2H), 3.26 (t, J = 7.3 Hz, 2H).¹³C NMR (126 MHz, CDCl₃): δ = 146.0, 144.1, 137.4, 134.4, 130.1, 129.2, 129.1, 127.5, 127.3, 126.7, 121.3, 120.7, 118.4, 103.0, 52.1, 37.1. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₁₉H₁₆ClN₂307.0997 found 307.0998. mp 120-124 °C. Example 2: 4-chloro-1-(3-chlorophenethyl)-1H-pyrrolo[3,2-c]quinoline IXb

Orange solid (13.2 mg, 0.039 mmol, 26%), HPLC purity 97%. ¹H NMR (500 MHz, CDCl₃): δ = 8.24- 8.26 (m, 1H), 8.18-8.20 (m, 1H), 7.61-7.67 (m, 2H), 7.23-7.25 (m, 1H), 7.19 (t, J = 7.7 Hz, 1H), 7.11- 7.12 (m, 1H), 6.85-6.86 (m, 2H), 6.72 (d, J = 3.2 Hz, 1H), 4.79 (t, J = 7.2 Hz, 2H), 3.24 (t, J = 7.2 Hz, 2H). ¹³C NMR (126 MHz, CDCl₃): δ = 145.9, 144.1, 139.1, 134.7, 134.0, 130.1, 129.5, 128.8, 127.4, 127.0, 126.3, 121.1, 120.1, 118.1, 102.8, 51.4, 36.5. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₁₉H₁₅Cl₂N₂341.0607 found 341.0611. mp 112-115 °C. Example 3: 4-chloro-1-(2-methoxyphenethyl)-1H-pyrrolo[3,2-c]quinoline IXc.

Pale yellow solid (92.0 mg, 0.274 mmol, 54%), HPLC purity 99%. ¹H NMR (500 MHz, DMSO-d₆): δ = 8.61 (dd, J = 8.3, 1.4 Hz, 1H), 8.04 (dd, J = 8.2, 1.5 Hz, 1H), 7.72 (dddd, J = 25.5, 8.3, 7.0, 1.4 Hz, 2H), 7.43 (d, J = 3.1 Hz, 1H), 7.22 (td, J = 7.7, 1.7 Hz, 1H), 7.10 (dd, J = 7.4, 1.7 Hz, 1H), 6.98 (dd, J = 8.3, 1.1 Hz, 1H), 6.85 (td, J = 7.4, 1.1 Hz, 1H), 6.68 (d, J = 3.1 Hz, 1H), 4.79 (t, J = 7.3 Hz, 2H), 3.80 (s, 3H), 3.15 (t, J = 7.3 Hz, 2H).¹³C NMR (126 MHz, DMSO-d₆): δ = 157.3, 144.2, 143.1, 133.7, 131.1, 130.5, 129.0, 128.2, 126.9, 126.4, 125.1, 121.1, 120.3, 119.8, 117.7, 110.6, 101.4, 55.3, 49.2, 31.4. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₂₀H₁₈ClN₂O 337.1102 found 337.1103. mp 105-107 °C. Example 4: 4-chloro-1-(3-methoxyphenethyl)-1H-pyrrolo[3,2-c]quinoline IXd.

White solid (100.0 mg, 0.298 mmol, 59%), HPLC purity 99%. ¹H NMR (500 MHz, CDCl₃): δ = 8.29 (dd, J = 8.0, 1.5 Hz, 1H), 8.18 (dd, J = 8.1, 1.6 Hz, 1H), 7.63-7.65 (m, 2H), 7.20-7.23 (m, 1H), 6.88 (d, J = 3.2 Hz, 1H), 6.79-6.81 (m, 1H), 6.70 (d, J = 3.2 Hz, 1H), 6.68 (d, J = 7.5 Hz, 1H), 6.55-6.56 (m, 1H), 4.77 (t, J = 7.3 Hz, 2H), 3.73 (s, 3H), 3.22 (t, J = 7.3 Hz, 2H). ¹³C NMR (126 MHz, CDCl₃): δ = 159.9, 145.8, 144.1, 138.7, 134.1, 130.0, 129.9, 129.6, 126.9, 126.2, 121.0, 121.0, 120.3, 118.2, 114.5, 112.4, 102.6, 55.2, 51.6, 36.8. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₂₀H₁₈ClN₂O 337.1102 found 337.1102. mp 155-158 °C. Example 5: 4-chloro-1-(4-methoxyphenethyl)-1H-pyrrolo[3,2-c]quinoline IXe.

Pale yellow solid (105.0 mg, 0.313 mmol, 62%), HPLC purity 99%.¹H NMR (500 MHz, DMSO-d₆): δ = 8.42-8.48 (m, 1H), 8.04 (dd, J = 7.7, 1.9 Hz, 1H), 7.67-7.75 (m, 2H), 7.38 (d, J = 3.1 Hz, 1H), 7.06- 7.11 (m, 2H), 6.81-6.86 (m, 2H), 6.67 (d, J = 3.1 Hz, 1H), 4.88 (t, J = 7.4 Hz, 2H), 3.71 (s, 3H), 3.11 (t, J = 7.3 Hz, 2H). ¹³C NMR (126 MHz, DMSO-d₆): δ = 158.0, 144.3, 143.1, 133.5, 131.1, 129.7, 129.3, 129.0, 126.9, 126.5, 121.1, 119.9, 117.6, 113.8, 101.3, 54.9, 50.7, 35.0. HRMS (ESI, pos.): [M+H]⁺ m/z calcd for C₂₀H₁₈ClN₂O 337.1102 found 337.1101. mp 122-124 °C. Example 6: 2-(2-(4-chloro-1H-pyrrolo[3,2-c]quinolin-1-yl)ethyl)phenol IXf.

Pale yellow solid (15.0 mg, 0.047 mmol, 33%), HPLC purity 95%. ¹H NMR (500 MHz, DMSO-d₆): δ = 9.77 (br. s, 1H), 8.68-8.73 (m, 1H), 8.01-8.06 (m, 1H), 7.66-7.72 (m, 2H), 7.46 (d, J = 3.1 Hz, 1H), 7.06 (td, J = 7.7, 1.7 Hz, 1H), 7.01 (dd, J = 7.5, 1.7 Hz, 1H), 6.85 (dd, J = 8.1, 1.1 Hz, 1H), 6.67-6.72 (m, 2H), 4.80-4.85 (m, 2H), 3.11-3.14(m, 2H). ¹³C NMR (126 MHz, DMSO-d₆): δ = 155.5, 144.3, 143.1, 133.7, 131.0, 130.5, 128.9, 127.8, 126.9, 126.3, 123.5, 121.4, 119.8, 119.0, 117.7, 114.9, 101.4, 49.1, 31.5. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₁₉H₁₆ClN₂O 323.0946 found 323.0942. mp 217- 221 °C. Example 7: 3-(2-(4-chloro-1H-pyrrolo[3,2-c]quinolin-1-yl)ethyl)phenol IXg. White solid (39.9 mg, 0.124 mmol, 83%). HPLC purity 99%. ¹H NMR (500 MHz, CH₃CN-d₃): δ = 8.43-8.45 (m, 1H), 8.05-8.07 (m, 1H), 7.67-7.70 (m, 2H), 7.10 (d, J = 3.2 Hz, 1H), 7.08 (d, J = 7.7 Hz, 1H), 6.91 (br. s, 1H), 6.64-6.66 (m, 2H), 6.62 (d, J = 7.4 Hz, 1H), 6.51 (t, J = 2.0 Hz, 1H), 4.84 (t, J = 7.2 Hz, 2H), 3.17 (t, J = 7.2 Hz, 2H).¹³C NMR (126 MHz, CH₃CN-d₃): δ = 158.1, 146.0, 144.8, 140.4, 135.1, 131.61, 130.61, 130.4, 127.8, 127.4, 122.2, 121.6, 121.3, 119.1, 116.7, 114.5, 102.5, 52.1, 37.1. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₁₉H₁₅ClN₂O+H 323.0946 found 323.0945. mp 172-175 °C. Example 8: 4-(2-(4-chloro-1H-pyrrolo[3,2-c]quinolin-1-yl)ethyl)phenol IXh.

Pale brown solid (35.5 mg, 0.110 mmol, 74%), HPLC purity 99%. ¹H NMR (500 MHz, DMSO-d₆): δ = 9.22 (br. s, 1H), 8.44 (dd, J = 8.1, 1.6 Hz, 1H), 8.04 (dd, J = 8.1, 1.6 Hz, 1H), 7.67-7.75 (m, 2H), 7.38 (d, J = 3.1 Hz, 1H), 6.94-6.97 (m, 2H), 6.64-6.67 (m, 3H), 4.85 (t, J = 7.4 Hz, 2H), 3.06 (t, J = 7.4 Hz, 2H).¹³C NMR (126 MHz, DMSO-d₆): δ = 155.9, 144.3, 143.1, 133.5, 131.4, 131.2, 129.6, 129.0, 128.7, 126.9, 126.5, 121.1, 117.6, 115.2, 101.2, 50.8, 35.1. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₁₉H₁₆ClN₂O 323.0946 found 323.0946. mp 187-190 °C. Example 9: 4-chloro-1-(3-ethoxyphenethyl)-1H-pyrrolo[3,2-c]quinoline IXi. Cream solid (24.7 mg, 0.071 mmol, 57%), HPLC purity 99%. ¹H NMR (500 MHz, CDCl₃): δ = 8.28- 8.30 (m, 1H), 8.17-8.19 (m, 1H), 7.61-7.66 (m, 2H), 7.19-7.22 (m, 1H), 6.89 (d, J = 3.1 Hz, 1H), 6.79 (ddd, J = 8.3, 2.5, 0.8 Hz, 1H), 6.71 (d, J = 3.1 Hz, 1H), 6.66-6.67 (m, 1H), 6.56 (t, J = 2.0 Hz, 1H), 4.78 (t, J = 7.3 Hz, 2H), 3.94 (q, J = 7.0 Hz, 2H), 3.22 (t, J = 7.3 Hz, 2H), 1.39 (t, J = 7.0 Hz, 3H).¹³C NMR (126 MHz, CDCl₃): δ = 159.4, 146.1, 144.1, 138.5, 133.9, 130.0, 129.8, 129.6, 126.8, 126.2, 121.0, 120.8, 120.3, 118.2, 115.1, 112.9, 102.6, 63.4, 51.6, 36.8, 14.8. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₂₁H₂₀ClN₂O 351.1259 found 351.1258. mp 169-171 °C. Example 10: 4-chloro-1-(3-isopropoxyphenethyl)-1H-pyrrolo[3,2-c]quinoline IXj.

Brown solid (36.1 mg, 0.099 mmol, 91%), HPLC purity 98%. ¹H NMR (500 MHz, CDCl₃): δ = 8.27- 8.29 (m, 1H), 8.17-8.18 (m, 1H), 7.63 (d, J = 2.0 Hz, 2H), 7.19 (d, J = 7.9 Hz, 1H), 6.88 (d, J = 3.2 Hz, 1H), 6.77 (dd, J = 8.2, 2.3 Hz, 1H), 6.70 (d, J = 3.2 Hz, 1H), 6.66 (d, J = 7.6 Hz, 1H), 6.53 (t, J = 1.9 Hz, 1H), 4.77 (t, J = 7.3 Hz, 2H), 4.39-4.44 (m, 1H), 3.21 (t, J = 7.3 Hz, 2H), 1.28 (d, J = 6.0 Hz, 6H). ¹³C NMR (126 MHz, CDCl₃): δ = 158.3, 145.8, 144.1, 138.7, 134.1, 130.0, 129.8, 129.6, 126.8, 126.2, 121.0, 120.8, 120.3, 118.2, 116.5, 114.3, 102.6, 77.3, 77.0, 76.7, 69.9, 51.6, 36.8, 22.0. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₂₂H₂₂ClN₂O 365.1415 found 365.1415. mp 97-101 °C. Example 11: 2-(3-(2-(4-chloro-1H-pyrrolo[3,2-c]quinolin-1-yl)ethyl)phenoxy)ethan-1-ol IXk. Pale yellow solid (48.4 mg, 0.132 mmol, 93%), HPLC purity 98%. ¹H NMR (500 MHz, MeOH-d₄): δ = 8.44-8.46 (m, 1H), 8.03-8.05 (m, 1H), 7.67-7.69 (m, 2H), 7.11-7.14 (m, 2H), 6.76 (ddd, J = 8.2, 2.5, 0.6 Hz, 1H), 6.69 (d, J = 3.2 Hz, 1H), 6.65 (d, J = 7.4 Hz, 1H), 6.57 (t, J = 1.9 Hz, 1H), 4.92 (t, J = 7.0 Hz, 2H), 3.88-3.90 (m, 2H), 3.80-3.82 (m, 2H), 3.22 (t, J = 7.0 Hz, 2H).¹³C NMR (126 MHz, MeOH- d₄): δ = 160.6, 146.7, 144.6, 140.4, 135.7, 132.2, 130.7, 129.6, 128.2, 127.7, 122.4, 122.3, 121.9, 119.4, 116.1, 114.2, 103.1, 70.5, 61.7, 52.4, 37.7. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₂₁H₂₀ClN₂O₂ 367.1208 found 367.1208. mp 72-75 °C. Example 12: 4-chloro-1-(4-ethoxyphenethyl)-1H-pyrrolo[3,2-c]quinoline IXl.

Pale yellow solid (26.2 mg, 0.075 mmol, 80%), HPLC purity 98%. ¹H NMR (500 MHz, CDCl₃): δ = 8.28-8.30 (m, 1H), 8.17-8.19 (m, 1H), 7.61-7.65 (m, 2H), 6.94-6.96 (m, 2H), 6.85 (d, J = 3.0 Hz, 1H), 6.80-6.82 (m, 2H), 6.69 (d, J = 3.2 Hz, 1H), 4.74 (t, J = 7.2 Hz, 2H), 4.01 (q, J = 7.0 Hz, 2H), 3.19 (t, J = 7.2 Hz, 2H), 1.41 (t, J = 7.0 Hz, 3H). ¹³C NMR (126 MHz, CDCl₃): δ = 158.1, 145.8, 144.1, 134.0, 130.0, 129.7, 129.6, 129.0, 126.8, 126.2, 121.0, 120.3, 118.2, 114.8, 102.5, 63.5, 52.0, 35.9, 14.8. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₂₁H₂₀ClN₂O 351.1259 found 351.1258. mp 144-147 °C. Example 13: 2-(4-(2-(4-chloro-1H-pyrrolo[3,2-c]quinolin-1-yl)ethyl)phenoxy)ethan-1-ol IXm. Green oil (28.9 mg, 0.079 mmol, 78%), HPLC purity 97%. ¹H NMR (500 MHz, MeOH-d₄): δ = 8.34- 8.36 (m, 1H), 7.98-7.99 (m, 1H), 7.61-7.64 (m, 2H), 7.03 (d, J = 3.2 Hz, 1H), 6.88-6.90 (m, 2H), 6.76- 6.78 (m, 2H), 6.61 (d, J = 3.2 Hz, 1H), 4.78 (t, J = 7.0 Hz, 2H), 3.93-3.95 (m, 2H), 3.80-3.82 (m, 2H), 3.12 (t, J = 7.0 Hz, 2H).¹³C NMR (126 MHz, MeOH-d₄): δ = 159.4, 146.6, 144.7, 135.0, 132.1, 130.9, 130.9, 129.6, 128.1, 127.6, 122.2, 121.9, 119.3, 115.8, 103.0, 70.5, 61.7, 52.7, 36.8. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₂₁H₂₀ClN₂O₂367.1208 found 367.1208. mp 127-130 °C. Example 14: 4-chloro-1-(3-methoxybenzyl)-1H-pyrrolo[3,2-c]quinolone IXn.

White powder (30.0 mg, 0.093 mmol, 37%) and HPLC purity 99%. ¹H NMR (500 MHz, CDCl₃): ^ = 8.11 (dd, J = 8.3, 0.9 Hz, 1H), 7.98 (dd, J = 8.4, 0.9 Hz, 1H), 7.54 (ddd, J = 8.3, 7.0, 1.3 Hz, 1H), 7.39 (ddd, J = 8.4, 7.0, 1.3 Hz, 1H), 7.22-7.26 (m, 1H), 7.18 (d, J = 3.2 Hz, 1H), 6.87 (d, J = 3.2 Hz, 1H), 6.81 (dd, J = 8.2, 2.1 Hz, 1H), 6.63-6.64 (m, 1H), 6.57 (t, J = 2.1 Hz, 1H), 5.73 (s, 2H), 3.70 (s, 3H).¹³C NMR (126 MHz, CDCl₃): ^ = 160.3, 145.6, 144.1, 137.8, 135.0, 130.3, 129.8, 129.6, 126.9, 126.0, 120.9, 120.7, 118.2, 117.9, 113.0, 112.0, 103.3, 55.2, 53.3. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₁₉H₁₆ClN₂O 323.0946 found 323.0943. mp 182-185 °C. Example 15: 4-chloro-1-(4-methoxybenzyl)-1H-pyrrolo[3,2-c]quinoline IXo. Pale yellow solid (72.1 mg, 0.224 mmol, 90%), HPLC purity 99%. ¹H NMR (500 MHz, CDCl₃): δ = 8.09-8.11 (m, 1H), 8.00 (d, J = 8.4 Hz, 1H), 7.51-7.55 (m, 1H), 7.37-7.40 (m, 1H), 7.12 (d, J = 3.2 Hz, 1H), 6.97 (d, J = 8.6 Hz, 2H), 6.82-6.83 (m, 3H), 5.66 (s, 2H), 3.73 (s, 3H).¹³C NMR (126 MHz, CDCl₃): δ = 159.3, 145.6, 144.0, 134.8, 129.6, 129.5, 127.9, 127.4, 126.8, 125.9, 120.8, 120.7, 117.9, 114.5, 103.0, 55.2, 52.9. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₁₉H₁₆ClN₂O 323.0946 found 323.0943. mp 107-109 °C. Example 16: 1-(3-methoxyphenethyl)-4-methyl-1H-pyrrolo[3,2-c]quinoline IXp.

Pink oil (11.0 mg, 0.032 mmol, 12%). HPLC purity 98%.¹H NMR (500 MHz, CDCl₃): ^ = 8.12 (dd, J = 8.3, 1.0 Hz, 1H), 7.97 (dd, J = 8.4, 1.0 Hz, 1H), 7.57-7.55 (m, 1H), 7.44-7.42 (m, J = 2.4 Hz, 1H), 7.18 (d, J = 3.2 Hz, 1H), 6.89-6.82 (m, 3H), 6.71 (ddd, J = 8.4, 2.2, 1.1 Hz, 1H), 5.71 (s, 2H), 3.84 (s, 3H). ¹³C NMR (126 MHz, CDCl₃): ^ = 152.7 (d, ¹J_C-F = 248.8 Hz), 147.5, 147.4, 145.7, 144.2, 134.8, 129.7 (d, ²J_C-F = 17.4 Hz), 128.9 (d, ⁴J_C-F = 5.8 Hz), 127.0, 126.1, 121.8 (d, ⁵J_C-F = 3.5 Hz), 121.0, 120.5, 117.9, 114.1 (d, ³J_C-F = 16.7 Hz), 114.0, 103.4, 56.3, 52.6. HRMS (ESI, pos.): m/z calcd for C₁₉H₁₄ClFN₂O+H [M+H]⁺ 341.0851 found 341.0852. Example 17: (4-chloro-1H-pyrrolo[3,2-c]quinonine-1-yl)(4-methoxyphenyl)methanone IXq. Yellow solid (265.5 mg, 0.790 mmol, 80%), HPLC purity 99%.¹H NMR (500 MHz, THF-d₈): δ = 8.24 (dd, J = 8.6, 0.8 Hz, 1H), 8.04 (dd, J = 8.4, 0.9 Hz, 1H), 7.97-7.94 (m, 2H), 7.63-7.59 (m, 2H), 7.47- 7.44 (m, 1H), 7.13-7.10 (m, 2H), 6.90 (d, J = 3.5 Hz, 1H), 3.91 (s, 3H).¹³C NMR (126 MHz, THF-d₈): δ = 169.1, 165.9, 146.2, 145.3, 137.7, 134.4, 130.5, 130.3, 128.5, 126.7, 125.4, 124.6, 123.5, 119.0, 115.2, 105.7, 56.1. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₁₉H₁₄ClN₂O₂337.0738 found 337.0737. mp 177-178 °C. Example 18: 1-(3-methoxyphenethyl)-1H-pyrrolo[3,2-c]quinoline Xa.

Colorless oil (6.0 mg, 0.020 mmol, 27%), HPLC purity 99%.¹H NMR (500 MHz, DMSO-d₆): δ = 9.08 (s, 1H), 8.42-8.46 (m, 1H), 8.11 (dd, J = 7.9, 1.8 Hz, 1H), 7.62-7.70 (m, 2H), 7.32 (d, J = 3.1 Hz, 1H), 7.16.-7.21 (m, 1H), 6.71-6.79 (m, 3H), 6.71 (d, J = 3.1 Hz, 1H), 4.90 (t, J = 7.4 Hz, 2H), 3.68 (s, 3H), 3.15 (t, J = 7.3 Hz, 2H).¹³C NMR (126 MHz, DMSO-d₆): δ = 159.2, 146.1, 143.9, 139.3, 131.9, 130.3, 130.0, 129.4, 125.8, 122.0, 120.9, 120.8, 118.1, 114.2, 112.1, 101.5, 54.8, 50.1, 36.0. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₂₀H₁₉N₂O 303.1492 found 303.1491. Example 19: 4-fluoro-1-(3-methoxyphenethyl)-1H-pyrrolo[3,2-c]quinoline Xb. White solid (10.0 mg, 0.031 mmol, 42%), HPLC purity 99%.¹H NMR (500 MHz, DMSO-d₆): δ = 8.42- 8.46 (m, 1H), 7.92-7.96 (m, 1H), 7.65-7.69 (m, 2H), 7.39 (d, J = 3.2 Hz, 1H), 7.15-7.20 (m, 1H), 6.76- 6.78 (m, 1H), 6.71-6.75 (m, 2H), 6.70 (d, J = 3.1 Hz, 1H), 4.93 (t, J = 7.3 Hz, 2H), 3.68 (s, 3H), 3.15 (t, J = 7.3 Hz, 2H).¹³C NMR (126 MHz, DMSO-d₆): δ = 159.2, 156.3 (d, ¹J_C-F = 236.9 Hz), 141.2 (d, ³J_C-F = 19.9 Hz), 139.0, 136.6 (d, ³J_C-F = 13.5 Hz), 131.1, 129.4, 128.5, 126.9, 125.6, 121.0, 120.9, 117.8, 114.3, 112.1, 109.4 (d, ²J_C-F = 43.4 Hz), 99.1 (d, ⁴J_C-F = 4.1 Hz), 54.8, 50.2, 35.9. ¹⁹F NMR (471 MHz, DMSO-d₆): δ= -68.0 (s). HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₂₀H₁₈FN₂O 321.1398 found 321.1397. mp 76-79 °C. Example 20: 4-bromo-1-(3-methoxyphenethyl)-1H-pyrrolo[3,2-c]quinoline Xc. White solid (10.0 mg, 0.026 mmol, 24%),

8.47 (m, 1H), 8.05 (dd, J = 8.1, 1.5 Hz, 1H), 7.68-7.72 (m, 2H), 7.40 (d, J = 3.2 Hz, 1H), 7.15-7.19(m, 1H), 6.76-6.78 (m, 1H), 6.71-6.74 (m, 2H), 6.59 (d, J = 3.1 Hz, 1H), 4.92 (t, J = 7.4 Hz, 2H), 3.68 (s, 3H), 3.15 (t, J = 7.3 Hz, 2H). ¹³C NMR (126 MHz, DMSO-d₆): δ = 159.2, 143.5, 139.0, 136.6, 132.6, 131.0, 129.4, 129.1, 126.9, 126.6, 122.4, 121.2, 120.9, 117.7, 114.2, 112.2, 102.8, 54.8, 50.4, 35.9. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₂₀H₁₈BrN₂O 381.0597 found 381.0598. mp 123-125 °C. Example 21: 4-iodo-1-(3-methoxyphenethyl)-1H-pyrrolo[3,2-c]quinoline Xd. White solid (5.7 mg, 0.013 mmol, 10%), HPLC purity 99%.¹H NMR (500 MHz, CDCl₃): δ = 8.28-8.30 (m, 1H), 8.20-8.22 (m, 1H), 7.61-7.65 (m, 2H), 7.22 (t, J = 7.9 Hz, 1H), 6.88 (d, J = 3.2 Hz, 1H), 6.77- 6.84 (m, 1H), 6.67-6.71 (m, 1H), 6.55 (t, J = 2.0 Hz, 1H), 6.52 (d, J = 3.2 Hz, 1H), 4.76 (t, J = 7.3 Hz, 2H), 3.73 (s, 3H), 3.22 (t, J = 7.3 Hz, 2H).¹³C NMR (126 MHz, CDCl₃): δ = 159.9, 145.4, 138.7, 131.1, 130.4, 129.9, 128.9, 127.8, 126.6, 126.4, 121.0, 120.4, 118.4, 117.0, 114.5, 112.5, 106.9, 55.2, 51.9, 36.8. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₂₀H₁₈IN₂O 429.0458 found 429.0457. mp 146-149 °C. Example 22: 1-(3-methoxyphenethyl)-4-methyl-1H-pyrrolo[3,2-c]quinoline Xe.

Yellow-brown solid (25.3 mg, 0.080 mmol, 53%), HPLC purity 98%.¹H NMR (500 MHz, CDCl₃): δ = 8.30 (dd, J = 8.3, 1.1 Hz, 1H), 8.21 (d, J = 7.7 Hz, 1H), 7.60-7.63 (m, 1H), 7.54-7.62 (m, 1H), 7.22 (t, J = 7.9 Hz, 1H), 6.88 (d, J = 2.9 Hz, 1H), 6.80 (dd, J = 8.3, 1.7 Hz, 1H), 6.71 (d, J = 7.4 Hz, 1H), 6.64 (d, J = 3.2 Hz, 1H), 6.58 (t, J = 2.0 Hz, 1H), 4.78 (t, J = 7.4 Hz, 2H), 3.73 (s, 3H), 3.24 (t, J = 7.3 Hz, 2H), 2.89 (s, 3H). ¹³C NMR (126 MHz, CDCl₃): δ = 159.9, 154.5, 144.4, 144.3, 139.1, 132.6, 129.8, 128.9, 126.2, 125.1, 122.1, 121.0, 120.1, 117.9, 114.5, 112.3, 101.9, 55.2, 51.5, 36.9, 22.5. HRMS (ESI, pos.) m/z: [M+H]⁺ calcd for C₂₁H₂₁N₂O 317.1648 found 317.1646. mp 91-94 °C. In vitro cytotoxicity (MTT test) The cancer cell lines used in this test were derived from T-lymphoblastic leukemia CCRF-CEM, leukemia K562 and their multiresistant counterparts (CEM-DNR, K562-TAX), solid tumors including lung (A549) and colon (HCT116, HCT116p53-/-) carcinomas, osteosarcoma cell line (U2OS), and for comparison, on two human non-cancer fibroblast lines (BJ). Cell suspensions were prepared and diluted according to the particular cell type and the expected target cell density (2500–30000 cells/well based on cell growth characteristics). Cells were added by pipette (80 ^L) into 96-well microtiter plates. Inoculates were allowed a pre-incubation period of 24 h at 37 °C and 5 % CO₂ for stabilisation. Four- fold dilutions, in 20- ^L aliquots, of the intended test concentration were added to the microtiter plate wells at time zero. All test compound concentrations were examined in duplicate. Incubation of the cells with the test compounds lasted for 72 h at 37 °C, in a 5 % CO₂ atmosphere at 100 % humidity. At the end of the incubation period, the cells were assayed using MTT. Aliquots (10 μL) of the MTT stock solution were pipetted into each well and incubated for a further 1–4 h. After this incubation period the formazan produced was dissolved by the addition of 100 ^L/well of 10 % aq SDS (pH = 5.5), followed by a further incubation at 37 °C overnight. The optical density (OD) was measured at 540 nm with a Labsystem iEMS Reader MF. Tumour cell survival (TCS) was calculated using the following equation: TCS = (OD_{drug-exposed well} / mean OD_{control wells}) x 100 %. The IC₅₀ value, the drug concentration in μM lethal to 50 % of the tumor cells, was calculated from dose-response curves. (NT = not tested) 3 ¹ CEM- K562 HCT116 Cmpd. R X R CEM K562 A549 HCT116 U2OS BJ DNR Tax p53-/- IXa H (CH2)2 Cl 0.28 0.31 0.17 0.33 0.18 0.27 0.34 0.22 1.23 IXb 3-Cl (CH₂)₂ Cl 0.29 0.36 0.15 0.36 0.70 0.45 0.66 0.34 ≥50 IXc 2-OMe (CH2)2 Cl 0.28 0.36 0.15 0.40 1.84 0.54 0.62 0.31 ≥50 IXd 3-OMe (CH2)2 Cl 0.058 0.097 0.029 0.087 0.033 0.029 0.029 0.038 ≥50 IXe 4-OMe (CH₂)₂ Cl 0.018 0.025 0.025 0.028 0.093 0.033 0.037 0.043 ≥50 IXf 2-OH (CH₂)₂ Cl 1.22 1.58 0.76 2.59 5.03 2.06 2.79 1.86 ≥50 IXg 3-OH (CH2)2 Cl 0.16 0.35 0.082 0.27 2.33 0.18 0.18 0.43 48 IXh 4-OH (CH₂)₂ Cl 0.93 1.04 0.51 1.52 9.08 1.75 2.21 1.57 ≥50 IXi 3-OEt (CH₂)₂ Cl 0.65 0.86 0.44 0.72 1.49 0.61 0.71 0.78 47 IXj 3-OiPr (CH2)2 Cl 8.4 13.9 5.5 13.1 21.8 15.1 12.9 16.05 ≥50 3- IXk (CH₂)₂ Cl 0.22 0.34 0.15 0.27 0.76 0.32 0.62 0.33 47 OEtOH IXl 4-OEt (CH2)2 Cl 0.086 0.086 0.042 0.082 0.15 0.080 0.10 0.087 ≥50 4- IXm (CH2)2 Cl 0.13 0.13 0.17 0.13 0.26 0.13 0.19 0.23 ≥50 OEtOH IXn 3-OMe CH₂ Cl 0.071 0.085 0.027 0.11 0.17 0.13 0.17 NT ≥50 IXo 4-OMe CH2 Cl 0.018 0.029 0.013 0.03 0.034 0.017 0.021 NT ≥50 3-OMe- IXp CH₂ Cl 0.013 NT 0.011 NA 0.025 0.013 NT 0.016 ≥50 4-F IXq 4-OMe C=O Cl 0.45 0.57 9.57 0.40 3.33 0.45 0.64 NT ≥50 Xa 3-OMe (CH₂)₂ H 1.63 1.90 0.69 1.36 7.77 1.96 2.41 1.57 ≥50 Xb 3-OMe (CH2)2 F 0.23 0.24 0.12 0.22 1.11 0.25 0.52 0.19 ≥50 Xc 3-OMe (CH₂)₂ Br 0.064 0.083 0.027 0.10 0.50 0.093 0.16 0.087 ≥50 Xd 3-OMe (CH₂)₂ I 0.19 0.26 0.11 0.14 0.44 0.15 0.19 0.20 ≥50 Xe 3-OMe (CH2)2 Me 0.19 0.27 0.11 0.14 0.44 0.15 0.19 0.28 ≥50 Tubulin binding assay The tubulin binding site for pyrroloquinolines derivatives was tested with compound IXd. The method was based on the ability of ethylene-bisiodoacetamide (EBI) to crosslink in living cells the cysteine residues Cys-239 and Cys-354 inside the colchicine-binding site. CCRF-CEM cells were pre-treated with 50 μmol/L of IXd, nocodazole, colchicine and vincristine or corresponding volume of DMSO and subjected to 100 μmol/L EBI. MW of β-tubulin is 55 kDa/EBI:β-tubulin mobility shift to 50 kDa. EBI:β- tubulin adduct has a higher mobility in PAGE than the native β-tubulin whose crosslinking was prevented by a ligand present in the cavity. Comparison of IXd with nocodazole and colchicine (Figure 1) demonstrates that they share the same binding site.

Claims

CLAIMS 1. Compound of general formula I:

or an N-oxide, a pharmaceutically acceptable salt, a hydrate or a solvate thereof, wherein: R₁ is selected from H, F, Cl, Br, I, NH₂, OH and C1-C3 alkyl; X is selected from C1-C2 alkylenyl, -C(=O)- and -CH₂-C(=O)- linker; R₂ is selected from phenyl and 5-8 membered heteroaryl containing at least one heteroatom selected from N, O, S; wherein said phenyl or heteroaryl is unsubstituted or substituted by at least one substituent selected from hydroxy, F, Cl, Br, I, C1-C4 alkoxy, hydroxy-C1-C4-alkoxy, halogen- C1-C4-alkoxy, amino-C1-C5-alkoxy, C1-C4 alkylamino, hydroxy-C1-C4-alkylamino, halogen- C1-C4-alkylamino, amino-C1-C4-alkylamino. 2. Compound according to claim 1, which is a compound of general formula II:

or an N-oxide, a pharmaceutically acceptable salt, a hydrate or a solvate thereof, wherein: R₁ is selected from H, F, Cl, Br, I, NH₂, OH and C1-C3 alkyl; X is selected from C1-C2 alkylenyl, -C(=O)- and -CH₂-C(=O)- linker; R₃ represents one or more substituents which are independently selected from hydroxy, F, Cl, Br, I, C1-C4 alkoxy, hydroxy-C1-C4-alkoxy, halogen-C1-C4-alkoxy, amino-C1-C5-alkoxy, C1-C4 alkylamino, hydroxy-C1-C4-alkylamino, halogen-C1-C4-alkylamino, amino-C1-C4-alkylamino. 3. Compound according to claim 2, wherein R₃ is selected from 3-methoxy, 4-methoxy and 4-ethoxy; or R₃ represents two substituents wherein one substituent is selected from 3-methoxy, 4-methoxy and 4-ethoxy and one substituent is selected from halogen; and/or X is selected from -CH₂- and -CH₂CH₂-; and/or R₁ is Cl or Br. 4. The compound according to any one of claims 1 to 3 for use as a medicament. 5. The compound according to any one of claims 1 to 3 for use in treating and preventing cancer. 6. The compound according to any one of claims 1 to 3 for use in treatment of cancer selected from leukaemia, lung adenocarcinoma, colorectal cancer and osteosarcoma. 7. A pharmaceutical composition containing at least one compound according to any one of claims 1 to 3, and at least one pharmaceutically acceptable excipient.