EP1546129A1 - Substituted pyridine derivatives as antitumor agent - Google Patents

Substituted pyridine derivatives as antitumor agent

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Publication number
EP1546129A1
EP1546129A1 EP03794969A EP03794969A EP1546129A1 EP 1546129 A1 EP1546129 A1 EP 1546129A1 EP 03794969 A EP03794969 A EP 03794969A EP 03794969 A EP03794969 A EP 03794969A EP 1546129 A1 EP1546129 A1 EP 1546129A1
Authority
EP
European Patent Office
Prior art keywords
pyrrolidin
diamine
pyridine
dichloro
pyridin
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP03794969A
Other languages
German (de)
French (fr)
Inventor
Mauro Angiolini
Maria Menichincheri
Domenico Fusar Bassini
Markus Gude
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Pfizer Italia SRL
Original Assignee
Pharmacia Italia SpA
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Filing date
Publication date
Application filed by Pharmacia Italia SpA filed Critical Pharmacia Italia SpA
Publication of EP1546129A1 publication Critical patent/EP1546129A1/en
Withdrawn legal-status Critical Current

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Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D213/00—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members
    • C07D213/02—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members
    • C07D213/04—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D213/60—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
    • C07D213/72—Nitrogen atoms
    • C07D213/73—Unsubstituted amino or imino radicals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D213/00—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members
    • C07D213/02—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members
    • C07D213/04—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D213/60—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
    • C07D213/72—Nitrogen atoms
    • C07D213/74—Amino or imino radicals substituted by hydrocarbon or substituted hydrocarbon radicals

Definitions

  • the present invention relates to novel substituted pyridine derivatives active as Telomerase inhibitors, to the use of them as therapeutic agents, in particular as antitumoral agents, to a process for their preparation and to • pharmaceutical compositions comprising them.
  • Cancer is one of the major causes of disease and the second leading cause of death in the western world. Most cancer patients still die due to metastatic disease. Despite the great increase in the knowledge and understanding of the regulatory mechanisms involved in the onset of malignancy, currently available treatments (including surgery, radiation and a variety of cytoreductive and hormone-based drugs, used alone or in combination) , are still highly non specific and toxic to the patient, causing severe side effects including nausea and vomiting, hair loss, diarrhea, fatigue and ulcerations. This evidence indicates the need for new and more effective anti-cancer therapies. Recently an understanding of the mechanisms by which normal cells reach the state of replicative senescence, i.e.
  • telomerase is a ribonucleoprotein enzyme responsible in most eukaryotes for the complete replication and maintenance of chromosome ends, or telomeres, which are composed of repeated DNA sequences (in particular human telomeres are formed by 5'-TTAGGG repeats). Telomerase binds to telomeric DNA using as a template a sequence contained within the RNA component of the enzyme necessary for the addition of the short sequence repeats to the chromosome 3' end (see Blackburn 1992, Annu . Rev. Biochem. , 61 , 113 -129) .
  • telomere activity cannot be detected and telomeres shorten with successive cell division: in fact actively dividing normal cells have the potential to lose 50- 200 base pairs after each round of cell division, finally resulting . in shortening of telomeres .
  • telomeres shortening will eventually lead to cellular senescence by various mechanisms.
  • telomere shortening in tumors and subsequent senescent phenotype (see Feng et al . Science, 1995, 269 , 1236-1241) . Moreover it has been recently shown ⁇ Hahn et al . Nature Med .
  • telomere shortening can cause telomere shortening and arrest of cell growth and apoptosis.
  • peptide-nucleic .acids, and 2 ' -O-MeRNA oligomers complementary 'to-, the template region of the RNA component of the- enzyme .. have been reported to cause inhibition of telomerase ' activity, telomere shortening and cell death in certain tumor cell lines (see Herbert et al .
  • telomere activity can be used to treat cancer, as cancer cells express telomerase activity, while normal human somatic cells usually do not express telomerase activity at biologically relevant levels (i.e., at levels sufficient to maintain telomere length over many cell divisions) . Also telomere length in tumors is reduced compared with non-transformed cells giving the possibility of a therapeutic window (see Nakamura et al . Cancer Letters 158, 2000, 179-184) . Therefore a need exists to find molecules that inhibit the activity of telomerase and interfere with the growth of many types of cancer.
  • the present invention fulfills such a need by providing a high general method of treating many - if not most malignancies, as demonstrated by the highly number of human tumor cell lines and tumors having telomerase activity.
  • the compounds of the present invention can be effective in providing treatments that discriminate between malignant and normal cells to a high degree, avoiding many of the deleterious side-effects present with most current chemotherapeutic regimes which rely on agents that kill
  • Embodiments of the present invention are substituted pyridine derivatives having the following formula (I)
  • R4 represents hydrogen or C1-C4 alkyl and R5 represents an unsubstituted or substituted C1-C18 n-alkyl or C3-C6 cycloalkyl ; or b) R4 and R5 taken together with N to which they are linked represent an unsubstituted or substituted saturated 5 to 7 membered heterocyclic ring;
  • halogen atom we intend a chlorine, bromine, fluorine or iodine atom.
  • C1-C18 n-alkyl we intend a straight alkyl chain having a carbon chain length of from 1 to 18 carbon atoms, unsubstituted or substituted by a group selected from C1-C4 alkoxycarbonyl, C1-C4 alkoxy, adamantyl, 2 ' -tetrahydrofuryl, methylene dimethoxy. or phenyl unsubstituted or substituted by C1-C4 alkoxy or C3-C6 cycloalkyl.
  • C1-C18 n-alkyl is an unsubstituted C18 straight alkyl chain or a C1-C4 straight alkyl chain unsubstituted or substituted by a group selected from C1-C4 alkoxycarbonyl, C1-C4 alkoxy, adamantyl, 2 ' -tetrahydrofuryl, methylene dimethoxy or phenyl unsubstituted or substituted by from 1 to 3 C1-C4 alkoxy or C3-C6 cycloalkyl.
  • C1-C4 alkyl we intend a group such as, for instance, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl or tert-butyl group.
  • C3-C6 cycloalkyl we intend a cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl group.
  • C1-C4 acyl we intend, for instance, formyl, acetyl , and propanoyl .
  • C1-C4 alkoxy is typically methoxy, ethoxy, propoxy or butoxy.
  • R4 and R5 form a 5 to 7 membered saturated heterocycle ring with the nitrogen atom to which they are linked, one carbon atom may be further replaced by heteroatoms such as nitrogen, oxygen and sulphur.
  • 5 to 7 membered optionally substituted saturated heterocyclic rings according to the present invention are pyrrolidine, pyrroline, pyrazolidine, pyrazoline, piperidine, piperazine, N-C1-C4 alkyl piperidine, N-C1-C4 alkyl-piperazine, N-phenyl piperazine, N-benzyl piperazine, N-pyrimidinyl piperazine, N- C1-C4 alkoxycarbonyl piperazine, morpholine, and the like.
  • salts of the compounds of formula (I) are, e.g., the acid addition salts with inorganic or organic, e.g. nitric, hydrochloric, hydrobromic, sulphuric, perchloric, phosphoric, acetic, trifluoroacetic, propionic, glycolic, lactic, oxalic, malonic, malic, maleic, tartaric, citric, benzoic, cinnamic, mandelic, methanesulphonic, isethionic and salicylic acid.
  • a preferred class of compounds of the present invention is represented by compounds of formula (I) above wherein: n is 1;
  • R represents hydrogen; Rl and R2 represent either both hydrogen or both Cl;
  • R3 represents NR4R5 wherein a') R4 represents hydrogen or C1-C4 alkyl, and R5 represents C1-C18 n-alkyl unsubstituted or substituted by adamantyl, 2 ' -tetrahydrofuryl, methylene dimethoxy CH(OMe)2, phenyl unsubstituted or substituted by C1-C4 alkoxy or C3-C6 cycloalkyl; or b ' ) R4 and R5 taken together with N to which they are linked represent an unsubstituted or substituted 5-7 membered saturated heterocyclic ring represented by the following formulae:
  • R6 represents hydrogen, methyl, phenyl, benzyl, ethoxycarbonyl (COOEt)or 2-pyriminidinyl; and the pharmaceutically acceptable salts thereof.
  • Examples of preferred compounds of the present invention, whenever appropriate in the form pharmaceutically acceptable salts, e.g. with hydrochloric or hydrobromic acid are listed, but not limited to, the following ones: 1)2- (4-Phenyl-piperazin-l-yl) -6-pyrrolidin-l-yl-pyridin-4- ylamine;
  • a further embodiment of the present invention is to provide a process for the preparation of compounds of formula (I) as defined above according to the method described in the following Scheme 1.
  • Hal is a halogen atom, i.e. Cl, Br, I or F; and n, R, R4 and R5 have the same meanings as reported in formula ( I ) above .
  • Compounds of formula (II) may be easily prepared starting from commercially available reagents following procedures well known in the art or they are commercially available from, for example, Aldrich Catalogue.
  • Novel intermediates of formula (III) may be prepared by reacting a compound of formula (II) with a proper nucleophile, i.e. an amine of formula (IV), wherein R4 and R5 are as defined above.
  • a proper nucleophile i.e. an amine of formula (IV)
  • R4 and R5 are as defined above.
  • proper reaction conditions for this transformation involve temperatures ranging from room temperature to reflux conditions in an organic solvent such as, for example, methanol, ethanol, isopropanol, n-buthanol, t-buthanol, tetrahydrofuran, dioxane, ethyl acetate, toluene or N-methyl-pyrrolidone for a time ranging from 30' to 24h, using 1 or more equivalents of amine of formula (IV) in high yield improving methods reported in the literature (see for example J " . Org. Chem . 1990, 55, 4992) .
  • Novel intermediates of formula (III) are then reacted with an amine of formula (V), i.e. pyrrolidine or piperidine, to get a compound of general formula (lb), i.e. a compound of formula
  • R is hydrogen and Rl and R2 are both halogen, in an organic solvent like methanol, ethanol, isopropanol, n- buthanol , t-buthanol, tetrahydrofuran, dioxane, ethyl acetate, toluene or N-methyl-pyrrolidone, at a temperature ranging from room temperature to reflux conditions for a time which may vary from 30' to 24h, using 1 or more equivalents of a compound of formula (V) , in high yield improving methods reported in the literature (see for example J " . Org. Chem. 1990, 55, 4992) .
  • a compound of formula (lb) may be converted in to a compound of formula (Ic) , i.e. a compound of formula (I) wherein R, Rl and R2 are hydrogen, under hydrogenolytic conditions, for example in ethanol or methanol as the solvent, by using ammonium formate as the source of hydrogen and Pd/C as catalyst at room temperature .
  • Other hydrogenolitic conditions can be applied (see, for example, classical hydrogenolysis in J " . March “Advanced Organic Chemistry", III Edition, Wiley - Interscience Publ . ) but the previous method is particularly suitable for parallel synthesis.
  • the stepwise solution phase method outlined in Scheme 1 has been conveniently adapted for the parallel synthesis of multiple analogs of general formula (Ic) in one shot.
  • Compounds of formula (lb) and (Ic) may be converted by acylation into compounds of formula (Ie) , i.e. compounds of formula (I) wherein R is C1-C4 acyl and Rl and R2 are both halogen, and (Ic) , i.e. compounds of formula (I) wherein R is C1-C4 acyl and Rl and R2 are both hydrogen, respectively, in dry pyridine under argon pressure, at temperatures ranging from 0° to 5° C, in presence of a catalytic amount of dimethylamino pyridine (DMAP) and corresponding chlorides as reagents .
  • DMAP dimethylamino pyridine
  • a compound of formula (I) may be converted into a pharmaceutically acceptable salt thereof using the suitable pharmaceutically acceptable acid, following the procedures well know in the art .
  • telomerase activity of compounds of the invention have been evaluated using a FlashPlate ® -based assay (PerkinElmer, Boston, Mass.) .
  • the method proved to be sensitive, accurate and able to reproducibly identify compounds that inhibit telomerase activity in a dose-dependent manner.
  • the assay mixture is constituted of:
  • telomerase enzyme diluted in a buffer, the composition of which has been selected to maintain the enzyme activity stable along the duration of the assay.
  • test compounds/positive control - increasing concentrations of test compounds/positive control .
  • telomeric repeats added are evaluated by hybridization in solution with a 3' -labeled short oligonucleotide probe.
  • FlashPlate ® where the binding between biotin and streptavidin occurs .
  • telomerase activity is proportional to the radioactivity measured and the inhibitory activity of the compounds is evaluated as IC 50 using the Sigma Plot fit program.
  • a human or animal body can thus be treated by a method, which comprises the administration of a pharmaceutically effective amount of a compound of formula (I) or a salt thereof.
  • the condition of the human or animal can thereby be improved.
  • Compounds of the invention can be used to treat, prevent, and/or ameliorate tumors and cancer.
  • the present invention provides a pharmaceutical composition, which comprises a therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof as an active ingredient and a pharmaceutically acceptable excipient or carrier.
  • the present invention also provides methods for the preparation of a pharmaceutical composition of a compound of formula (I) or a pharmaceutically acceptable salt thereof, which comprises admixing a therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof with a pharmaceutically acceptable excipient or carrier.
  • Pharmaceutical compositions of the invention containing pharmaceutically acceptable excipients or carriers and a compound of the invention can be prepared by any of the well known techniques of pharmacy that comprise admixing the excipients or carriers with a drug or therapeutic agent.
  • compositions of the present invention can be formulated and used as, e.g., tablets, capsules or elixirs for oral administration, suppositories for rectal administration, sterile solutions, suspensions for injectable administration, and the other compositions known in the art.
  • a pharmaceutical composition refers to a composition in a form suitable for administration, e.g., systemic administration, into a cell or subject, including for example a human. Suitable forms, in part, depend upon the use or the route of entry, for example oral, transdermal, intravenous, subcutaneous, intraperitoneal, inhalation, oral, intrapul onary and intramuscular.
  • Acceptable carriers or diluents for therapeutic use are well known in the pharmaceutical art, and are described, for example, in Remington 's Pharmaceutical Sciences, Mack Publishing Co. (A.R. Gennaro edit. 1985) hereby incorporated by reference herein.
  • preservatives, stabilizers, dyes and flavoring agents can be provided.
  • antioxidants and suspending agents can be used.
  • the pharmaceutical compositions containing compounds of the invention can be in a form suitable for oral use, for example, as tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsion, hard or soft capsules, or syrups or elixirs.
  • compositions intended for oral use can be prepared according to any method known to the art for the manufacture of pharmaceutical compositions and such compositions can contain one or more such sweetening agents, flavoring agents, coloring agents or preservative agents in order to provide pharmaceutically elegant and palatable preparations.
  • Tablets contain the active ingredient in admixture with non-toxic pharmaceutically acceptable excipients that are suitable for the manufacture of tablets.
  • excipients can be, for example, inert diluents; such as calcium carbonate, sodium carbonate, lactose, calcium phosphate or sodium phosphate; granulating and disintegrating agents, for example, corn starch, or alginic acid; binding agents, for example starch, gelatin or acacia; and lubricating agents, for example magnesium stearate, stearic acid or talc.
  • the tablets can be uncoated or they can be coated by known techniques . In some cases such coatings can be prepared by known techniques to delay disintegration and absorption in the gastrointestinal tract and thereby provide a sustained action over a longer period.
  • a time delay material such as glyceryl monosterate or glyceryl distearate can be employed.
  • Formulations for oral use can also be presented as hard gelatin capsules wherein the active ingredient is mixed with an inert solid diluent, for example, calcium carbonate, calcium phosphate or kaolin, or as soft gelatin capsules wherein the active ingredient is mixed with water or an oil medium, for example peanut oil, liquid paraffin or olive oil.
  • Dispersible powders and granules suitable for preparation of an aqueous suspension by the addition of water provide the active ingredient in admixture with a dispersing or wetting agent, suspending agent and one or more preservatives. Suitable dispersing or wetting agents or suspending agents are exemplified by those already mentioned above. Additional excipients, for example sweetening, flavoring and coloring agents, can also be present.
  • compositions of the invention can also be in the form of oil-in-water emulsions.
  • the oily phase can be a vegetable oil or a mineral oil or mixtures of these.
  • Suitable emulsifying agents can be naturally-occurring gums, for example gum acacia or gum tragacanth, naturally-occurring phosphatides, for example soy bean, lecithin, and esters or partial esters derived from fatty acids and hexitol, anhydrides, for example sorbitan monooleate, and condensation products of the said partial esters with ethylene oxide, for example polyoxyethylene sorbitan monooleate.
  • the emulsions can also contain sweetening and flavoring agents.
  • Syrups and elixirs can be formulated with sweetening agents, for example glycerol, propylene glycol, sorbitol, glucose or sucrose. Such formulations can also contain a demulcent, a preservative and flavoring and coloring agents.
  • the pharmaceutical compositions can be in the form of a sterile injectable aqueous or oleaginous suspension. This suspension can be formulated according to the known art using those suitable dispersing or wetting agents and suspending agents that have been mentioned above.
  • the sterile injectable preparation can also be a sterile injectable solution or suspension in a non-toxic parentally acceptable diluent or solvent, for example as a solution in 1, 3-butanediol .
  • Suitable vehicles and solvents that can be employed are water, Ringer's solution and isotonic sodium chloride solution.
  • sterile, fixed oils are conventionally employed as a solvent or suspending medium.
  • any bland fixed oil can be employed including synthetic mono-or diglycerides .
  • fatty acids such as oleic acid find use in the preparation of injectables .
  • the specific dose of a compound administered according to this invention to obtain a therapeutic effect will, of course, be determined by the particular circumstances surrounding the administration, including, for example, the age, weight, condition of the patient, gender, diet, time of administration, rate of excretion, drug combination, severity of disease, and administration route; specific dosage regimens can be fit to any particular subject on the basis of the individual need and the professional judgment of the person administering or supervising the administration of the aforesaid compounds.
  • the dosage range adopted will depend on the route of administration and on the age, weight and condition of the patient being treated.
  • daily doses of the compounds of the invention typically administered by parenteral route, for example, intravenously by bolus or infusion is from 1 to 1000 mg/m 2 body surface area, for instance from 10 to 500 mg/m 2 .
  • the dosages can be administered at once or can be divided into a number of smaller doses to be administered at varying intervals.
  • a pharmaceutically effective dose is that dose required to prevent, inhibit the occurrence, or treat (alleviate a symptom to some extent, preferably all of the symptoms) of a disease state.
  • the pharmaceutically effective dose depends on, for example, the type of disease, the composition used, the route of administration, the type of mammal being treated, the physical characteristics of the specific mammal under consideration, concurrent medication, and other factors that those skilled in the medical arts will recognize.
  • Compounds and pharmaceutical compositions according to the invention are useful for the prevention, amelioration, and/or treatment of benign and malignant tumors or neoplasias including cancer, such as, for example, brain cancer, bone cancer, epithelial cell-derived neoplasia (epithelial carcinoma) such as basal cell carcinoma, adenocarcinoma, esophageal cancer, small bowel cancer and stomach cancer, colon cancer, liver cancer, gastrointestinal cancer, colorectal cancer, bladder cancer, pancreas cancer, ovary cancer, cervical cancer, pancreatic cancer, lung cancers, breast cancer and skin cancer, prostate cancer, and renal cell carcinoma.
  • a method for treating a patient having a cancer or tumor which comprises administering a therapeutically effective amount
  • the invention further comprises methods of treatment, prevention, or amelioration of benign and malignant tumors or neoplasias comprising administering a pharmaceutically effective amount of a compound of the invention to a subject in need thereof such that the benign or malignant tumor or neoplasia is treated, prevented or ameliorated.
  • N*2*-cyclohexyl-6-pyrrolidin-l-yl-pyridine-2, 4-diamine (2) : X H-NMR (400Mhz, CDC1 3 ) , ppm: 1.06-2.12 (m, 14H) , 3.37 (m, 5H) , 3.67 (sb, 2H) , 4.16 (sb, IH) , 5.08 (s, IH) , 5.10 (s, IH) .
  • N*2*-methyl-6-pyrrolidin-l-yl-pyridine-2, 4-diamine (9) X H- NMR (400Mhz, CDC1 3 ) , ppm: 1.88 (m, 4H) , 2.78 (s, 3H) , 3.40 (m, 4H) , 3.76 (sb, 2H)., 4.25 (sb, IH) , 5.05 (s, IH) , 5.10 (s, IH) .

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Abstract

The present invention relates to novel substituted pyridine derivatives active as Telomerase inhibitors, to the use of them as therapeutic agents, in particular as antitumoral agents, to a process for their preparation and to pharmaceutical compositions comprising them.

Description

TITLE
Substituted pyridine derivatives as antitumor agent
SUMMARY OF THE INVENTION
The present invention relates to novel substituted pyridine derivatives active as Telomerase inhibitors, to the use of them as therapeutic agents, in particular as antitumoral agents, to a process for their preparation and to • pharmaceutical compositions comprising them.
BACKGROUND OF THE INVENTION
Cancer is one of the major causes of disease and the second leading cause of death in the western world. Most cancer patients still die due to metastatic disease. Despite the great increase in the knowledge and understanding of the regulatory mechanisms involved in the onset of malignancy, currently available treatments (including surgery, radiation and a variety of cytoreductive and hormone-based drugs, used alone or in combination) , are still highly non specific and toxic to the patient, causing severe side effects including nausea and vomiting, hair loss, diarrhea, fatigue and ulcerations. This evidence indicates the need for new and more effective anti-cancer therapies. Recently an understanding of the mechanisms by which normal cells reach the state of replicative senescence, i.e. the loss of proliferative capacity that cells normally undergo in the cellular aging process, has begun to emerge and in this respect telomerase appears to have a central role . Telomerase is a ribonucleoprotein enzyme responsible in most eukaryotes for the complete replication and maintenance of chromosome ends, or telomeres, which are composed of repeated DNA sequences (in particular human telomeres are formed by 5'-TTAGGG repeats). Telomerase binds to telomeric DNA using as a template a sequence contained within the RNA component of the enzyme necessary for the addition of the short sequence repeats to the chromosome 3' end (see Blackburn 1992, Annu . Rev. Biochem. , 61 , 113 -129) . In most human somatic cells telomerase activity cannot be detected and telomeres shorten with successive cell division: in fact actively dividing normal cells have the potential to lose 50- 200 base pairs after each round of cell division, finally resulting . in shortening of telomeres . Recently it has been hypothesized that the cumulative loss of telomeric DNA over repeated cell divisions can act as a trigger for cellular senescence and aging, and that regulation of telomerase can have important biological implications (see Barley 1991, Mutation Research, 256, 271 -282) . In fact in the absence of telomerase, telomeres shortening will eventually lead to cellular senescence by various mechanisms. This phenomenon, thought to be responsible for cellular aging, is termed the "mitotic clock" (see Holt et al . Nat . Biotechnol . , 1996, 15, 1734 -1741 ) . Telomerase activity is restored in immortalized cell lines and in more than 85% of human tumors, thus maintaining telomeres length stable (see Shay, J. W. and Bacchetti , S. Eur. J. Cancer, 1997, 33 , 787- 791) . Thus in cancer cells having telomerase activity and where the malignant phenotype is due to the loss of cell cycle or growth controls or other genetic damage, telomeric DNA is not lost during cell division and telomeres are maintained, thereby allowing the cancer cells to become immortal, leading to a terminal prognosis for the patient. Telomerase inhibition can lead to telomere shortening in tumors and subsequent senescent phenotype (see Feng et al . Science, 1995, 269 , 1236-1241) . Moreover it has been recently shown {Hahn et al . Nature Med . , 1999, 5, 1164-1170) that inhibition of telomerase activity by expressing in tumor cells a catalytically-inactive form of human TERT (TElomerase Reverse Transcriptase, the catalytic subunit of the enzyme) can cause telomere shortening and arrest of cell growth and apoptosis. In addition peptide-nucleic .acids, and 2 ' -O-MeRNA oligomers complementary 'to-, the template region of the RNA component of the- enzyme .. have been reported to cause inhibition of telomerase ' activity, telomere shortening and cell death in certain tumor cell lines (see Herbert et al . PNAS, 1999, 96, 14276-14281; Shammas et al . Oncogene, 1999, 18, 6191 -6200) . These data strongly support inhibition of telomerase activity as an innovative, selective and useful method for the development of new anticancer agents. Thus compounds that inhibit telomerase activity can be used to treat cancer, as cancer cells express telomerase activity, while normal human somatic cells usually do not express telomerase activity at biologically relevant levels (i.e., at levels sufficient to maintain telomere length over many cell divisions) . Also telomere length in tumors is reduced compared with non-transformed cells giving the possibility of a therapeutic window (see Nakamura et al . Cancer Letters 158, 2000, 179-184) . Therefore a need exists to find molecules that inhibit the activity of telomerase and interfere with the growth of many types of cancer.
The present invention fulfills such a need by providing a high general method of treating many - if not most malignancies, as demonstrated by the highly number of human tumor cell lines and tumors having telomerase activity. The compounds of the present invention can be effective in providing treatments that discriminate between malignant and normal cells to a high degree, avoiding many of the deleterious side-effects present with most current chemotherapeutic regimes which rely on agents that kill
• dividing cells indiscriminately. Therefore • they are expected to exhibit greater safety and1 lack of toxic effects in comparison with traditional' ' chemotherapeutic anticancer agents .
DETAILED DESCRIPTION OF THE INVENTION
Embodiments of the present invention are substituted pyridine derivatives having the following formula (I)
(I) wherein n is 1 or 2; R represents hydrogen or C1-C4 acyl; Rl and R2 represent either both hydrogen or both halogen; R3 represents NR4R5 wherein:
a) R4 represents hydrogen or C1-C4 alkyl and R5 represents an unsubstituted or substituted C1-C18 n-alkyl or C3-C6 cycloalkyl ; or b) R4 and R5 taken together with N to which they are linked represent an unsubstituted or substituted saturated 5 to 7 membered heterocyclic ring;
and the pharmaceutically acceptable salts thereof.
In the present description, unless otherwise specified, with the term halogen atom we intend a chlorine, bromine, fluorine or iodine atom.
With the term C1-C18 n-alkyl we intend a straight alkyl chain having a carbon chain length of from 1 to 18 carbon atoms, unsubstituted or substituted by a group selected from C1-C4 alkoxycarbonyl, C1-C4 alkoxy, adamantyl, 2 ' -tetrahydrofuryl, methylene dimethoxy. or phenyl unsubstituted or substituted by C1-C4 alkoxy or C3-C6 cycloalkyl. Preferably, C1-C18 n-alkyl is an unsubstituted C18 straight alkyl chain or a C1-C4 straight alkyl chain unsubstituted or substituted by a group selected from C1-C4 alkoxycarbonyl, C1-C4 alkoxy, adamantyl, 2 ' -tetrahydrofuryl, methylene dimethoxy or phenyl unsubstituted or substituted by from 1 to 3 C1-C4 alkoxy or C3-C6 cycloalkyl. With the term C1-C4 alkyl we intend a group such as, for instance, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl or tert-butyl group.
With the term C3-C6 cycloalkyl we intend a cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl group.
With the term C1-C4 acyl we intend, for instance, formyl, acetyl , and propanoyl .
C1-C4 alkoxy is typically methoxy, ethoxy, propoxy or butoxy.
When R4 and R5 form a 5 to 7 membered saturated heterocycle ring with the nitrogen atom to which they are linked, one carbon atom may be further replaced by heteroatoms such as nitrogen, oxygen and sulphur. Examples of 5 to 7 membered optionally substituted saturated heterocyclic rings according to the present invention are pyrrolidine, pyrroline, pyrazolidine, pyrazoline, piperidine, piperazine, N-C1-C4 alkyl piperidine, N-C1-C4 alkyl-piperazine, N-phenyl piperazine, N-benzyl piperazine, N-pyrimidinyl piperazine, N- C1-C4 alkoxycarbonyl piperazine, morpholine, and the like.
Pharmaceutically acceptable salts of the compounds of formula (I) are, e.g., the acid addition salts with inorganic or organic, e.g. nitric, hydrochloric, hydrobromic, sulphuric, perchloric, phosphoric, acetic, trifluoroacetic, propionic, glycolic, lactic, oxalic, malonic, malic, maleic, tartaric, citric, benzoic, cinnamic, mandelic, methanesulphonic, isethionic and salicylic acid. A preferred class of compounds of the present invention is represented by compounds of formula (I) above wherein: n is 1;
R represents hydrogen; Rl and R2 represent either both hydrogen or both Cl;
R3 represents NR4R5 wherein a') R4 represents hydrogen or C1-C4 alkyl, and R5 represents C1-C18 n-alkyl unsubstituted or substituted by adamantyl, 2 ' -tetrahydrofuryl, methylene dimethoxy CH(OMe)2, phenyl unsubstituted or substituted by C1-C4 alkoxy or C3-C6 cycloalkyl; or b ' ) R4 and R5 taken together with N to which they are linked represent an unsubstituted or substituted 5-7 membered saturated heterocyclic ring represented by the following formulae:
wherein R6 represents hydrogen, methyl, phenyl, benzyl, ethoxycarbonyl (COOEt)or 2-pyriminidinyl; and the pharmaceutically acceptable salts thereof.
Examples of preferred compounds of the present invention, whenever appropriate in the form pharmaceutically acceptable salts, e.g. with hydrochloric or hydrobromic acid are listed, but not limited to, the following ones: 1)2- (4-Phenyl-piperazin-l-yl) -6-pyrrolidin-l-yl-pyridin-4- ylamine;
2)N-2-Cyclohexyl-6-pyrrolidin-l-yl-pyridine-2 , 4-diamine;
3 ) N-2-Cyclohexylmethyl-6-pyrrolidin-1-yl-pyridine-2,4- diamine;
4) 6-Pyrrolidin-l-yl-N-2- (3,4, 5-trimethoxy-benzyl) -pyridine-
2, 4 -diamine;
5) N-2-Benzyl-6-pyrrolidin-1-yl-pyridine-2,4-diamine ;
6)N-2- (2 , 2 -Dimethoxy-ethyl) -6-pyrrolidin-1-yl-pyridine-2 , 4- diamine;
7) 2-piperazin-1-yl-6-pyrrolidin-1-yl-pyridin- -ylamine;
8) 4- (4-Amino-6-pyrrolidin-l-yl-pyridin-2-yl) -piperazine-1- carboxylic acid ethyl ester;
9) -2 -methyl-6-pyrrolidin-1-yl-pyridine-2 , 4-diamine; 10) 2-Morpholin-4-yl-6-pyrrolidin-l-yl-pyridin-4-ylamine;
11) 6-Pyrrolidin-l-yl-N-2- (tetrahydro-furan-2-ylmethyl) - pyridine-2 , 4-diamine;
12 ) N-2- (4-Phenyl-butyl) -6-pyrrolidin-1-yl-pyridine-2 , 4- diamine; 13 ) N-2- henethyl-6-pyrrolidin-1-yl-pyridine-2,4-diamine ;
14) 2- (4-Pyrimidin-2-yl-piperazin-l-yl) -6-pyrrolidin-1-yl- py idin-4-ylamine;
15) N-2-Methyl-N-2-octadecyl-6-pyrrolidin-l-yl-pyridine-2 , 4- diamine ; 16 ) 4-Methyl-6 ' -pyrrolidin- 1-yl -3 , 4 , 5 , 6-tetrahydro-2H-
[1 , 2 ' ] bipyridinyl-4 ' -ylamine ;
17) 2 , 6-Di-pyrrolidin- l-yl -pyridin-4 -yl-amine ;
18 ) N- (2 , 6-Di-pyrrolidin-l-yl -pyridin-4 -yl ) -acetamide ;
19) 6 ' -Pyrrolidin-1-yl-3, , 5, 6-tetrahydro-2H- [1,2'] bipyridinyl-4 ' -ylamine; 20) (4-Amino-6-pyrrolidin-l-yl-pyridin-2-ylamino) -acetic acid ethyl ester;
21) 2-pyrrolidin-l-yl-6- (1,3,4, 9-tetrahydro-beta-carbolin-2- yl) -pyridin-4-ylamine; 22) [ (4-Amino-6-pyrrolidin-l-yl-pyridin-2-yl) -methyl-amino] - acetic acid ethyl ester;
23) 2- (4-Methyl-piperazin-l-yl) -6-pyrrolidin-l-yl-pyridin-4- ylamine;
24) N-2-adamantan-l-yl-6-pyrrolidin-l-yl-pyridine-2 , 4-diamine; 25) N-2-adamantan-2-ylmethyl-6-pyrrolidin-l-yl-pyridine-2 , 4- diamine;
26)N-2- (3-Methoxy-phenyl) -6-pyrrolidin-l-yl-pyridine-2 , 4- diamine;
27)N-2- (2-Methoxy-ethyl) -βpyrrolidin -1-yl-pyridine-2 , 4- diamine;
28)N-2- (4-Methoxy-phenyl) -6-pyrrolidin-l-yl-pyridine-2, 4- diamine;
29) -2-octadecyl-6-pyrrolidin-1-yl-pyridine-2,4-diamine;
30)3, 5-Dichloro-2- (4-phenyl-piperazin-l-yl) -6-pyrrolidin-l- yl-pyridin- -ylamine;
31)3, 5-Dichloro-N-2-cyclohexyl-6-pyrrolidin-l-yl-pyridine-
2 , 4-diamine;
32) 3 , 5-Dichloro-N-2-cyclohexylmethyl-6-pyrrolidin-l-yl- pyridine-2 , 4-diamine; 33) 3, 5-Dichloro-6-pyrrolidin-l-yl-N-2- (3 , 4, 5-trimethoxy- benzyl) -pyridine-2 , 4-diamine;
34 )N-2-Benzyl-3 , 5-dichloro-6-pyrrolidin-1-yl-pyridine-2 , 4- diamine;
35) 3 , 5-Dichloro-N-2- (2, 2-dimethoxy-ethyl) -6-pyrrolidin-l-yl- pyridine-2 , 4-diamine; 36) 2- (4-Benzyl-piperazin-l-yl) -3, 5-dichloro-6-pyrrolidin-l- yl-pyridin-4-ylamine;
37)4- (4-Amino-3, 5-dichloro-6-pyrrolidin-l-yl-pyridin-2-yl) - piperazine-1-carboxylic acid ethyl ester; 38)N-2-Benzyl-3, 5-dichloro-N-2 -methyl-6-pyrrolidin-1-yl- pyridine-2 , 4-diamine;
39)3, 5-Dichloro-2-morpholin-4-yl-6-pyrrolidin-l-yl-pyridin-4- ylamine ;
40)3, 5-Dichloro-6-pyrrolidin-l-yl-N-2- (tetrahydro-furan-2- ylmethyl) -pyridine-2 , 4-diamine;
41) 3, 5-Dichloro-N-2- (4-phenyl-butyl) -6-pyrrolidin-l-yl- pyridine-2 , 4-diamine;
42) 3 , 5-Dichloro-N-2-phenethyl-6-pyrrolidin-l-yl-pyridine-2 , 4- diamine; 43) 3 , 5-Dichloro-2- (4-pyrimidin-2-yl-piperazin-l-yl) -6- pyrrolidin-1-yl-pyridin-4-ylamine ;
44) 3 , 5-Dichloro-N-2-methyl-N-2-octadecyl-6-pyrrolidin-l-yl- pyridine-2 , 4-diamine;
45) 3 ' ,5' -Dichloro-4-methyl-6 ' -pyrrolidin-1-yl-3 , 4, 5,6- tetrahydro-2H- [1,2 ' ] bipyridinyl-4 ' -ylamine;
46)3, 5-Dichloro-2 , 6-di-pyrrolidin-l-yl-pyridin-4-ylamine;
47)3' , 5 ' -Dichloro-6 ' -pyrrolidin-1-yl-3 , 4, 5, 6-tetrahydro-2H- [1,2'] bipyridinyl-4 ' -ylamine;
48)3, 5-Dichloro-2-fluoro-6-pyrrolidin-l-yl-pyridin-4-ylamine; 49) (4-Amino-3 , 5-dichloro-6-pyrrolidin-l-yl-pyridin-2- ylamino) -acetic acid ethyl ester;
50) 3, 5-dichloro-2-pyrrolidin-l-yl-6- (1,3,4, 9-tetrahydro-beta- carbolin-2 -yl) -pyridin- -ylamine;
51) [ (4-Amino-3 , 5-dichloro-6-pyrrolidin-l-yl-pyridin-2-yl) - methyl-amino] -acetic acid ethyl ester; 52) 3 , 5-dichloro-2- (4-Methyl-piperazin-1-yl) -6-pyrrolidin-1- yl-pyridin-4-ylamine;
53 ) N-2-adamantan-1-yl-3 , 5-dichloro-6-pyrrolidin-l-yl- pyridine-2 , 4-diamine; 54) N-2-adamantan-2-ylmethyl-3 , 5-dichloro-6-pyrrolidin-l-yl- pyridine-2 , 4-diamine;
55) 3 , 5-dichloro-N-2- (3-Methoxy-phenyl) -6-pyrrolidin-1-yl- pyridine-2 , 4-diamine;
56)3, 5-dichloro-N-2- (2-Methoxy-ethyl) -6-pyrrolidin-l-yl- pyridine-2 , 4-diamine ;
57)3, 5-dichloro-N-2- (4-Methoxy-phenyl) -6-pyrrolidin-l-yl- pyridine-2 , 4-diamine and
58)3, 5-dichloro-N-2-octadecyl-6-pyrrolidin-l-yl-pyridine-2 , 4- diamine.
A further embodiment of the present invention is to provide a process for the preparation of compounds of formula (I) as defined above according to the method described in the following Scheme 1.
Scheme 1
NHL
(ID Hal Hal
XX.
In the above Scheme 1 Hal is a halogen atom, i.e. Cl, Br, I or F; and n, R, R4 and R5 have the same meanings as reported in formula ( I ) above . Compounds of formula (II) may be easily prepared starting from commercially available reagents following procedures well known in the art or they are commercially available from, for example, Aldrich Catalogue.
Novel intermediates of formula (III) may be prepared by reacting a compound of formula (II) with a proper nucleophile, i.e. an amine of formula (IV), wherein R4 and R5 are as defined above. Typically proper reaction conditions for this transformation involve temperatures ranging from room temperature to reflux conditions in an organic solvent such as, for example, methanol, ethanol, isopropanol, n-buthanol, t-buthanol, tetrahydrofuran, dioxane, ethyl acetate, toluene or N-methyl-pyrrolidone for a time ranging from 30' to 24h, using 1 or more equivalents of amine of formula (IV) in high yield improving methods reported in the literature (see for example J". Org. Chem . 1990, 55, 4992) .
Novel intermediates of formula (III) are then reacted with an amine of formula (V), i.e. pyrrolidine or piperidine, to get a compound of general formula (lb), i.e. a compound of formula
(I) wherein R is hydrogen and Rl and R2 are both halogen, in an organic solvent like methanol, ethanol, isopropanol, n- buthanol , t-buthanol, tetrahydrofuran, dioxane, ethyl acetate, toluene or N-methyl-pyrrolidone, at a temperature ranging from room temperature to reflux conditions for a time which may vary from 30' to 24h, using 1 or more equivalents of a compound of formula (V) , in high yield improving methods reported in the literature (see for example J". Org. Chem. 1990, 55, 4992) . A compound of formula (lb) may be converted in to a compound of formula (Ic) , i.e. a compound of formula (I) wherein R, Rl and R2 are hydrogen, under hydrogenolytic conditions, for example in ethanol or methanol as the solvent, by using ammonium formate as the source of hydrogen and Pd/C as catalyst at room temperature . Other hydrogenolitic conditions can be applied (see, for example, classical hydrogenolysis in J". March "Advanced Organic Chemistry", III Edition, Wiley - Interscience Publ . ) but the previous method is particularly suitable for parallel synthesis. In fact the stepwise solution phase method outlined in Scheme 1 has been conveniently adapted for the parallel synthesis of multiple analogs of general formula (Ic) in one shot.
Compounds of formula (lb) and (Ic) may be converted by acylation into compounds of formula (Ie) , i.e. compounds of formula (I) wherein R is C1-C4 acyl and Rl and R2 are both halogen, and (Ic) , i.e. compounds of formula (I) wherein R is C1-C4 acyl and Rl and R2 are both hydrogen, respectively, in dry pyridine under argon pressure, at temperatures ranging from 0° to 5° C, in presence of a catalytic amount of dimethylamino pyridine (DMAP) and corresponding chlorides as reagents .
If desired, a compound of formula (I) may be converted into a pharmaceutically acceptable salt thereof using the suitable pharmaceutically acceptable acid, following the procedures well know in the art . Biological activity
The telomerase activity of compounds of the invention have been evaluated using a FlashPlate®-based assay (PerkinElmer, Boston, Mass.) . The method proved to be sensitive, accurate and able to reproducibly identify compounds that inhibit telomerase activity in a dose-dependent manner. Briefly the assay mixture is constituted of:
- telomerase enzyme diluted in a buffer, the composition of which has been selected to maintain the enzyme activity stable along the duration of the assay.
- dNTPs, deoxynucleotides 5' -triphosphate .
- biotinylated oligo as primer.
- increasing concentrations of test compounds/positive control .
After two hours of incubation at 37° degrees the telomeric repeats added are evaluated by hybridization in solution with a 3' -labeled short oligonucleotide probe.
The extent of hybridization is then quantitated by transferring the reaction mixture in a streptavidin-coated
FlashPlate®, where the binding between biotin and streptavidin occurs .
The telomerase activity is proportional to the radioactivity measured and the inhibitory activity of the compounds is evaluated as IC50 using the Sigma Plot fit program.
With the above-described method IC50 values of the compounds of the present invention were determined.
The results relative to a representative selection of compounds of the invention are shown in Table 1. Table 1
The data reported in Table 1 clearly show the activity of the compounds according to the invention as telomerase inhibitors .
A human or animal body can thus be treated by a method, which comprises the administration of a pharmaceutically effective amount of a compound of formula (I) or a salt thereof. The condition of the human or animal can thereby be improved.
Therapeutic Use
Compounds of the invention can be used to treat, prevent, and/or ameliorate tumors and cancer.
The present invention provides a pharmaceutical composition, which comprises a therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof as an active ingredient and a pharmaceutically acceptable excipient or carrier.
The present invention also provides methods for the preparation of a pharmaceutical composition of a compound of formula (I) or a pharmaceutically acceptable salt thereof, which comprises admixing a therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof with a pharmaceutically acceptable excipient or carrier. Pharmaceutical compositions of the invention containing pharmaceutically acceptable excipients or carriers and a compound of the invention can be prepared by any of the well known techniques of pharmacy that comprise admixing the excipients or carriers with a drug or therapeutic agent.
The compositions of the present invention can be formulated and used as, e.g., tablets, capsules or elixirs for oral administration, suppositories for rectal administration, sterile solutions, suspensions for injectable administration, and the other compositions known in the art.
A pharmaceutical composition refers to a composition in a form suitable for administration, e.g., systemic administration, into a cell or subject, including for example a human. Suitable forms, in part, depend upon the use or the route of entry, for example oral, transdermal, intravenous, subcutaneous, intraperitoneal, inhalation, oral, intrapul onary and intramuscular.
Acceptable carriers or diluents for therapeutic use are well known in the pharmaceutical art, and are described, for example, in Remington 's Pharmaceutical Sciences, Mack Publishing Co. (A.R. Gennaro edit. 1985) hereby incorporated by reference herein. For example, preservatives, stabilizers, dyes and flavoring agents can be provided. In addition, antioxidants and suspending agents can be used. The pharmaceutical compositions containing compounds of the invention can be in a form suitable for oral use, for example, as tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsion, hard or soft capsules, or syrups or elixirs. Compositions intended for oral use can be prepared according to any method known to the art for the manufacture of pharmaceutical compositions and such compositions can contain one or more such sweetening agents, flavoring agents, coloring agents or preservative agents in order to provide pharmaceutically elegant and palatable preparations. Tablets contain the active ingredient in admixture with non-toxic pharmaceutically acceptable excipients that are suitable for the manufacture of tablets. These excipients can be, for example, inert diluents; such as calcium carbonate, sodium carbonate, lactose, calcium phosphate or sodium phosphate; granulating and disintegrating agents, for example, corn starch, or alginic acid; binding agents, for example starch, gelatin or acacia; and lubricating agents, for example magnesium stearate, stearic acid or talc. The tablets can be uncoated or they can be coated by known techniques . In some cases such coatings can be prepared by known techniques to delay disintegration and absorption in the gastrointestinal tract and thereby provide a sustained action over a longer period. For example, a time delay material such as glyceryl monosterate or glyceryl distearate can be employed. Formulations for oral use can also be presented as hard gelatin capsules wherein the active ingredient is mixed with an inert solid diluent, for example, calcium carbonate, calcium phosphate or kaolin, or as soft gelatin capsules wherein the active ingredient is mixed with water or an oil medium, for example peanut oil, liquid paraffin or olive oil. Dispersible powders and granules suitable for preparation of an aqueous suspension by the addition of water provide the active ingredient in admixture with a dispersing or wetting agent, suspending agent and one or more preservatives. Suitable dispersing or wetting agents or suspending agents are exemplified by those already mentioned above. Additional excipients, for example sweetening, flavoring and coloring agents, can also be present.
Pharmaceutical compositions of the invention can also be in the form of oil-in-water emulsions. The oily phase can be a vegetable oil or a mineral oil or mixtures of these. Suitable emulsifying agents can be naturally-occurring gums, for example gum acacia or gum tragacanth, naturally-occurring phosphatides, for example soy bean, lecithin, and esters or partial esters derived from fatty acids and hexitol, anhydrides, for example sorbitan monooleate, and condensation products of the said partial esters with ethylene oxide, for example polyoxyethylene sorbitan monooleate. The emulsions can also contain sweetening and flavoring agents. Syrups and elixirs can be formulated with sweetening agents, for example glycerol, propylene glycol, sorbitol, glucose or sucrose. Such formulations can also contain a demulcent, a preservative and flavoring and coloring agents. The pharmaceutical compositions can be in the form of a sterile injectable aqueous or oleaginous suspension. This suspension can be formulated according to the known art using those suitable dispersing or wetting agents and suspending agents that have been mentioned above. The sterile injectable preparation can also be a sterile injectable solution or suspension in a non-toxic parentally acceptable diluent or solvent, for example as a solution in 1, 3-butanediol . Among the acceptable vehicles and solvents that can be employed are water, Ringer's solution and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose, any bland fixed oil can be employed including synthetic mono-or diglycerides . In addition, fatty acids such as oleic acid find use in the preparation of injectables .
It is understood that the specific dose of a compound administered according to this invention to obtain a therapeutic effect will, of course, be determined by the particular circumstances surrounding the administration, including, for example, the age, weight, condition of the patient, gender, diet, time of administration, rate of excretion, drug combination, severity of disease, and administration route; specific dosage regimens can be fit to any particular subject on the basis of the individual need and the professional judgment of the person administering or supervising the administration of the aforesaid compounds. The dosage range adopted will depend on the route of administration and on the age, weight and condition of the patient being treated. As an example, daily doses of the compounds of the invention, typically administered by parenteral route, for example, intravenously by bolus or infusion is from 1 to 1000 mg/m2 body surface area, for instance from 10 to 500 mg/m2. The dosages can be administered at once or can be divided into a number of smaller doses to be administered at varying intervals. A pharmaceutically effective dose is that dose required to prevent, inhibit the occurrence, or treat (alleviate a symptom to some extent, preferably all of the symptoms) of a disease state. The pharmaceutically effective dose depends on, for example, the type of disease, the composition used, the route of administration, the type of mammal being treated, the physical characteristics of the specific mammal under consideration, concurrent medication, and other factors that those skilled in the medical arts will recognize. Compounds and pharmaceutical compositions according to the invention are useful for the prevention, amelioration, and/or treatment of benign and malignant tumors or neoplasias including cancer, such as, for example, brain cancer, bone cancer, epithelial cell-derived neoplasia (epithelial carcinoma) such as basal cell carcinoma, adenocarcinoma, esophageal cancer, small bowel cancer and stomach cancer, colon cancer, liver cancer, gastrointestinal cancer, colorectal cancer, bladder cancer, pancreas cancer, ovary cancer, cervical cancer, pancreatic cancer, lung cancers, breast cancer and skin cancer, prostate cancer, and renal cell carcinoma. Further disclosed is a method for treating a patient having a cancer or tumor, which comprises administering a therapeutically effective amount of a compound according to the invention.
The invention further comprises methods of treatment, prevention, or amelioration of benign and malignant tumors or neoplasias comprising administering a pharmaceutically effective amount of a compound of the invention to a subject in need thereof such that the benign or malignant tumor or neoplasia is treated, prevented or ameliorated.
The following examples illustrate but do not limit the invention. All references cited herein are incorporated by reference in their entirety.
Examples of single compound preparation Example 1: Synthesis of 3 , 5-Dichloro-2-fluoro-6-pyrrolidin-l- yl-pyridin-4-ylamine (48)
cl
A solution of 2 , 6-difluoro-3 , 5-dichloro-4-amino-pyridine (1000 mg, 5 mmol) and pyrrolidine (426 mg, 6 mmol) in 50 ml of toluene was stirred for 18 h under magnetic stirring at room temperature. To the reaction mixture was then added 0.1 ml of triethylamine and the solvent evaporated to dryness. The crude was purified by flash column chromatography (eluant: hexane/ethyl acetate 2:1) to yield 3 , 5-Dichloro-2- fluoro-6-pyrrolidin-1-yl-pyridin-4-ylamine (800 mg, 65%) as a pure white solid. Elemental analysis calculated for CsHioFxClaNs : C 43.22, H 4.03, N 16.80; found C 43.29, H 4.04, N 16.81. MS m/z: 250 (MH+) . -R-NMR (400Mhz, DMS0-d6) , ppm 1.80 (m,4H), 3.47 (m,4H), 6.46 (sb, 2H) .
Example 2: Synthesis of 3 , 5-Dichloro-2 , 6-di-pyrrolidin-l-yl- pyridin-4-ylamine (46)
To a solution of 2, 6-difluoro-3 , 5-dichloro-4-amino-pyridine
(5000 mg; 25 mmol) in 30 mL of pyrrolidine was heated to reflux for lh and the mixture was poured into water. The precipitated solid was filtered and washed with water. The collected solid was then dried and purified by flash column chromatography (eluant: hexane/ethyl acetate 2:1) to yield 3 , 5-Dichloro-2, 6-di-pyrrolidin-l-yl-pyridin-4-ylamine (6850 mg, 91%) as a pure slight brown solid. Elemental analysis calculated for Cι3H18Cl2N4 : C 51.84, H 6.02, N 18.60, Cl 23.54; found C 51.85, H 6.04, N 18.37, Cl 23.22. MS m/z: 301 (MH+) . αH-NMR (400Mhz, DMSO-d6) , ppm 1.92 (m, 8H) , 3.45 (m, 8H) , 5.82 (sb, 2H) .
Example 3: Synthesis of 3 ' , 5 ' -Dichloro-6 ' -pyrrolidin-1-yl- 3,4,5,6-tetrahydro-2H- [1, 2 '] bipyridinyl-4 ' -ylamine (47)
A mixture of substrate (86 mg; 0.34 mmol) and piperidine (1.2 ml) in 30 ml of ethyl acetate was heated to reflux for 8h. The solvent was evaporated and the crude purified by flash column chromatography (eluant: hexane/ethyl acetate 2:1) to yield 3 ' , 5 ' -Dichloro-6 ' -pyrrolidin-1-yl-3 ,4,5, 6-tetrahydro- 2H- [1, 2 '] bipyridinyl-4 ' -ylamine (60 mg, 55%) as a pure white solid. Elemental analysis calculated for CιH2oCl2N : C 53.34, H 6.39, N 17.77, Cl 22.49; found C 56.21, H 7.01, N 16.02, Cl 20.81. MS m/z: 315 (MH+) .
XH-NMR (400Mhz, DMSO-d6) , ppm 1.42-1.63 (m, 6H) , 1.80 (m, 4H) , 3.15 (m, 4H) , 3.44 (m, 4H) , 5.91 (sb, 2H) .
Example 4: Synthesis of N- (2 , 6-Di-pyrrolidin-1-yl-pyridin-4- yl) -acetamide (18)
To a solution of substrate (116 mg; 0.5 mmol) in 7 mL of dry pyridine under argon pressure at 5° C were added 12 mg of DMAP and 0.036 ml (40 mg, 0.52 mmol) of acetyl chloride. After 3h stirring the mixture was washed with 15 ml of water and extracted three times with ethyl acetate . The combined organic phase was dried and evaporated to dryness. Purification by flash column chromatography (eluant: DCM/methanol 190:20) afforded N- (2 , 6-Di-pyrrolidin-l-yl- pyridin-4-yl) -acetamide (65 mg, 47%) as a pure white solid. Elemental analysis calculated for C15H22N4O : C 65.67, H 8.08, N 20.42; found C 65.51, H 8.11, N 19.97. MS m/z: 275 (MH+) . ^H-NMR (400Mhz, DMSO-d6) , ppm: 1.87 (m, 8H) , 2.10 (s, 3H) , 3.33 (m, 8H) , 5.96 (sb, 2H) , 9.62 (sb, 1H) .
Example 5: Synthesis of 2 , 6-Di-pyrrolidin-l-yl-pyridin-4-yl- amine (17)
To a solution of substrate (900 mg, 3 mmol) dissolved in 60 ml of methanol was added palladium catalyst on carbon
(10%) and the mixture was hydrogenated at 60 psi for 2h. The catalyst was filtered and the solvent evaporated to dryness affording 600 mg of product (86%) . MS m/z: 233 (MH+) . XH-NMR ( 400Mhz , DMSO- d6 ) , ppm : 1 . 81 (m, 4H) , 3 . 22 (m, 4H) , 4 . 92 ( s , 2H) , 5 . 11 ( sb , 2H) .
Example 6 : General procedure for parallel synthesis of compounds having general formula (lb)
( II ) (III) (lb)
To a solution of 2, 6-difluoro-3 , 5-dichloro-4-amino-pyridine in N-methyl-pyrrolidone (0.2M) were added 3 equivalents of the proper amine, 3 equivalents of N,N-diisopropylethylamine and the mixture was heated to 90° C for 18h. Then 6 equivalents of pyrrolidine were added and for 18h the mixture was heated. The solvent was evaporated to dryness and the residue was purified by flash column chromatography (eluant: n-hexane/ethyl acetate from 5/1 to l/l) . Following compounds have been prepared using this procedure:
3 , 5-Dichloro-2- (4-phenyl-piperazin-l-yl) -6-pyrrolidin-1-yl- pyridin-4-ylamine (30): yield 61%. MS m/z: 392 (MH+) . XH-NMR (400Mhz, DMS0-d6) , ppm: 1.80 (m, 4H) , 3.20 (m, 4H) , 3.34 (m, 4H) , 3.54 (m, 4H) , 5.87 (sb, 2H) , 6.77 (t, 1H) , 6.96 (d, 2H) , 7.21 (d, 2H) .
3 , 5-Dichloro-N*2*-cyclohexyl-6-pyrrolidin-l-yl-pyridine-2 , 4- diamine (31): yield 69%. MS m/z: 329 (MH+) . ^-NMR (400Mhz, DMSO-d6) , ppm: 1.06-1.92 (m, 14H) , 3.41 (m, 4H) , 3.73 (m, IH) , 5.13 (d, IH) , 5.57(sb, 2H) .
3 , 5-Dichloro-N*2*-cyclohexylmethyl-6-pyrrolidin-1-yl- pyridine-2, 4-diamine (32) : yield 76%. MS m/z: 343 (MH+) .
^Η-NMR (400Mhz, DMSO-d6) , ppm: 0.80-1.20 (m, 5H) , 1.45-1.66 (m, 6H) , 1.79 (m, 4H) , 3.12 (t, 2H) , 3.42 (m, 4H) , 5.54 (sb, 2H) , 5.74 (t, IH) .
3, 5-Dichloro-6-pyrrolidin-l-yl-N*2*- (3,4, 5-trimethoxy- benzyl) -pyridine-2 , 4-diamine (33): yield 72%. MS m/z: 427 (MH+) .
^Η-NMR (400Mhz, DMSO-d6) , ppm: 1.77 (m, 4H) , 3.42 (m, 4H) , 3.60 (s, 3H) , 3.70 (s, 6H) , 4.38 (d, 2H) , 5.60 (sb, 2H) , 6.38 (t, IH) , 6.84 (s, 2H) .
N*2*-Benzyl-3 , 5-dichloro-6-pyrrolidin-l-yl-pyridine-2 , 4- diamine (34) : yield 28%. MS m/z: 337 (MH+) .
XH-NMR (400Mhz, DMSO-d6) , ppm: 1.76 (m, 4H) , 3.39 (m, 4H) , 4.47 (d, 2H) , 5.59 (sb, 2H) , 6.41 (t, IH) , 7.12-7.30 (m, 5H) .
3, 5-Dichloro-N*2*- (2 , 2-dimethoxy-ethyl) -6-pyrrolidin-1-yl- pyridine-2, 4-diamine (35) : yield 48%. MS m/z: 335 (MH+) . ^-NMR (400Mhz, DMSO-dβ) , ppm: 1.78 (m, 4H) , 3.22 (s, 3H) , 3.30 (s, 3H) , 3.38 (t, 2H) , 3.42 (m, 4H) , 4.56 (t, IH) , 5.62 (m, 3H) .
2- (4-Benzyl-piperazin-l-yl) -3 , 5-dichloro-6-pyrrolidin-l-yl- pyridin-4-ylamine (36): yield 45%. MS m/z: 406 (MH+) . XH-NMR (400Mhz, DMSO-d6) , ppm: 1.77 (m, 4H) , 2.43 (m, 4H) , 3.17 (m, 4H) , 3.42 (m, 6H) , 5.82 (sb, 2H) , 7.21-7.32 (m, 5H) .
4- (4-Amino-3, 5-dichloro-6-pyrrolidin-l-yl-pyridin-2-yl) - piperazine- 1-carboxylic acid ethyl ester (37) : yield 64%. MS m/z: 388 (MH+) .
XH-NMR (400Mhz, DMSO-dg) , ppm: 1.18 (t, 3H) , 1.79(m,4H) , 3.13 (m, 4H) , 3.41-3.52 (m, 8H) , 4.05 (q, 2H) , 5.90 (sb, 2H) .
N*2*-Benzyl-3 , 5-dichloro-N*2*-methyl-6-pyrrolidin-l-yl- pyridine-2,4-diamine (38): yield 83%. MS m/z: 351 (MH+) . XH-NMR (400Mhz, DMSO-dg), ppm: 1.77 (m, 4H) , 2.76 (s, 3H) , 3.44 (m, 4H) , 4.38 (s,, 2H) , 5.81 (sb, 2H) , 7.31(m, 5H) .
3 , 5-Dichloro-2-morpholin-4-yl-6-pyrrolidin-l-yl-pyridin-4- ylamine (39): yield 37%. MS m/z: 317 (MH+) .
^-NMR (400Mhz, DMSO-dg), ppm: 1.79 (m, 4H) , 3.16 (m, 4H) , 3.50 (m, 4H) , 3.63 (m, 4H) , 5.84 (sb, 2H) .
3, 5-Dichloro-6-pyrrolidin-l-yl-N*2*- (tetrahydro-furan-2- ylmethyl) -pyridine-2, 4-diamine (40): yield 70%. MS m/z: 330 (MH+) .
^-NMR (400Mhz, DMSO-dg), ppm: 1.56 (m, 4H) , 1.78 (m, 4H) , 3.36 (m, 2H) , 3.43 (m, 4H) , 3.58 (m, IH) , 3.77 (m, IH) , 3.96 (qp, IH) , 5.60 (sb, 3H) .
3, 5-Dichloro-N*2*- (4-phenyl-butyl) - 6-pyrrolidin- 1-yl- pyridine-2, 4-diamine (41): yield 64%. MS m/z: 379 (MH+) . ^H-NMR (400Mhz, DMSO-dg) , ppm: 1.57 (m, 4H) , 1.78 (m, 4H) , 2.58 (t, 2H) , 3.23 (q, 2H) , 3.40 (m, 4H) , 5.47 (sb, 2H) , 5.78 (t, IH) , 7.10-7.24 (m, 5H) .
3, 5-Dichloro-N*2*-phenethyl-6-pyrrolidin-l-yl-pyridine-2,4- diamine (42) : yield 63%. MS m/z: 351 (MH+) .
XH-NMR (400Mhz, DMSO-dg) , ppm: 1.80 (m, 4H) , 2.80 (dd, 2H) ,
3.42-3.57 (m, 6H) , 5.57 (sb, 2H) , 5.83 (t, IH) , 7.16-7.32 (m,
5H) .
3 , 5-Dichloro-2- (4-pyrimidin-2-yl-piperazin-l-yl) -6- pyrrolidin-l-yl-pyridin-4-ylamine (43) : yield 68%. MS m/z:
394 (MH+) .
XH-NMR (400Mhz, DMSO-dg) , ppm: 1.80 (m, 4H) , 3.20 (m, 4H) , 3.44 (m, 4H) , 3.82 (m, 4H) , 5.88 (sb, 2H) , 6.62 (t, IH) , 8.38
(d, 2H) .
3,5-Dichloro-N*2*-methyl-N*2*-octadecyl-6-pyrrolidin-l-yl- pyridine-2 , 4-diamine (44): yield 70%. XH-NMR (400Mhz, DMSO-dg), ppm: 0.83 (s, 3H) , 1.22 (s, 32H) , 1.92 (m, 4H) , 2.96 (s, 3H) , 3.40 (m, 4H) , 3.42 (m, 2H) , 5.84 (sb, 2H) .
3 ' , 5 ' -Dichloro-4-methyl-6 ' -pyrrolidin-l-yl-3 , 4, 5 , 6- tetrahydro-2H- [1,2 '] bipyridinyl-4 ' -ylamine (45) :yield 55%. MS m/z: 329 (MH+) . 'XH-NMR (400Mhz, DMSO-d6) , ppm: 0.94 (d, 3H) , 1.65-2.03 (m, 9H) , 3.15 (m, 2H) , 3.71 (m, 4H) , 4.03 (m, 2H) , 5.84 (sb, 2H) . (4-Amino-3 , 5-dichloro-6-pyrrolidin-l-yl-pyridin-2-ylamino) ■ acetic acid ethyl ester (49)
3, 5-dichloro-2-pyrrolidin-l-yl-6- (1,3,4, 9-tetrahydro-beta- carbolin-2-yl) -pyridin-4-ylamine (50)
[ (4-Amino-3 , 5-dichloro-6-pyrrolidin-l-yl-pyridin-2-yl) - methyl-amino] -acetic acid ethyl ester (51)
3, 5-dichloro-2- (4-Methyl-piperazin-l-yl) -6-pyrrolidin-1-yl■ pyridin-4-ylamine (52)
N-2 -adamantan-1-yl-3 , 5-dichloro-6-pyrrolidin-1-yl-pyridine- 2, 4-diamine (53)
N-2-adamantan-2-ylmethyl-3, 5-dichloro-6-pyrrolidin-1-yl- pyridine-2 , 4-diamine (54)
3, 5-dichloro-N-2- (3-Methoxy-phenyl) -6-pyrrolidin-1-yl- pyridine-2 , 4-diamine (55)
3 , 5-dichloro-N-2- (2-Methoxy-ethyl) -6-pyrrolidin-l-yl- pyridine-2 , 4-diamine (56)
3 , 5-dichloro-N-2- (4-Methoxy-phenyl) -6-pyrrolidin-1-yl- pyridine-2 , 4-diamine (57)
3 , 5-dichloro-N-2-octadecyl-6-pyrrolidin-l-yl-pyridine-2 , 4- diamine (58) Example 7: General procedure for parallel synthesis of compounds having general formula (Ic) .
n
(lb) (Ic)
Derivative of general formula (lb) was dissolved in 2 ml of methanol and 10 mg of Pd-C 10% catalyst and 150 mg of ammonium formate was added. The mixture was then stirred for 3h and solution was filtered over Celite and the solvent evaporated. The residue was dissolved in DCM, washed with aqueous saturated NaHC03, dried with Na2S04 and evaporated to dryness. No further purification has been made. For all the compounds reported in Example 6 the yields of hydrogenation were almost quantitative. Following compounds have been prepared using this procedure:
2- (4-phenyl-piperazin-l-yl) -6-pyrrolidin-l-yl-pyridin-4- ylamine (1): XH-NMR (400Mhz, CDC13) , ppm: 1.84 (m, 4H) , 3.20 (m, 4H) , 3.34 (m, 4H) , 3.54 (m, 4H) , 4.80 (sb, 2H) , 5.17 (s, IH) , 5.38 (s, IH) , 6.84 (t, IH) , 6.96(d, 2H) , 7.24 (d, 2H) .
N*2*-cyclohexyl-6-pyrrolidin-l-yl-pyridine-2, 4-diamine (2) : XH-NMR (400Mhz, CDC13) , ppm: 1.06-2.12 (m, 14H) , 3.37 (m, 5H) , 3.67 (sb, 2H) , 4.16 (sb, IH) , 5.08 (s, IH) , 5.10 (s, IH) . N*2* -cyclohexylmethyl-6 -pyrrolidin- 1-yl-pyridine-2,4-diamine (3): ^H-NMR (400Mhz, CDC13) , ppm: 0.80-1.28 (m, 5H) , 1.45- 1.83 (m, 10H) , 2.97 (t, 2H) , 3.42 (m, 4H) , 3.78 (sb, 2H) , 4.26 (sb, IH) , 5.03 (s, 2H) .
6-pyrrolidin-l-yl-N*2*- (3 , 4, 5-trimethoxy-benzyl) -pyridine- 2, 4-diamine (4): ^-NMR (400Mhz, CDCl3) , ppm: 1.92 (m, 4H) , 3.39 (m, 4H) , 3.78 (sb, 2H) , 3.83 (s, 9H) , 4.38 (d, 2H) , 5.08 (s, IH) , 5.13 (s, IH) , 6.61 (m, 2H) .
N*2*-Benzyl-6-pyrrolidin-l-yl-pyridine-2 , 4-diamine (5): 1H- NMR (400Mhz, CDC13) , ppm: 1.90 (m, 4H) , 3.39 (m, 4H) , 3.77 (sb, 2H) , 4.42 (d, 2H) , 4.53 (sb, IH) , 5.03 (s, IH) , 5.06 (s, IH) , 7.12-7.41 (m, 5H) .
N*2*- (2, 2 -dimethoxy-ethyl) -6-pyrrolidin-1-yl-pyridine-2 ,4- diamine (6): ^-NMR (400Mhz, CDC13) , ppm: 1.92 (m, 4H) , 3.34 (s, 3H) , 3.37 (s, 3H) , 3.40 (m, 6H) , 3.76 (sb, 2H) , 4.13 (sb, IH) , 4.62 (t, IH) , 5.04 (s, IH) , 5.11 (s, IH) .
2-piperazin-l-yl-6-pyrrolidin-l-yl-pyridin-4-ylamine (7) : 1H- NMR (400Mhz, CDC13) , ppm: 1.87 (m, 4H) , 2.88-3.40 (m, 8H) , 3.62 (m, 4H) , 4.77 (sb, 3H) , 5.29 (s, 2H) .
4- (4-Amino-6-pyrrolidin-l-yl-pyridin-2-yl) -piperazine-1- carboxylic acid ethyl ester (8) : ^H-NMR (400Mhz, CDC13) , ppm: 1.26 (t, 3H) , 1.86(m,4H), 3.16-3.60 (m, 12H) , 3.78 (sb, 2H) , 4.22 (q, 2H) , 4.76 (s, IH) , 5.13 (s, IH) . N*2*-methyl-6-pyrrolidin-l-yl-pyridine-2, 4-diamine (9) : XH- NMR (400Mhz, CDC13) , ppm: 1.88 (m, 4H) , 2.78 (s, 3H) , 3.40 (m, 4H) , 3.76 (sb, 2H)., 4.25 (sb, IH) , 5.05 (s, IH) , 5.10 (s, IH) .
2-morpholin-4-yl-6-pyrrolidin-l-yl-pyridin-4-ylamine (10) : αH-NMR (400Mhz, CDCl3) , ppm: 1.86 (m, 4H) , 3.25-3.77 (m, 12H) , 4.76 (sb, 2H) , 5.15 (s, IH) , 5.30 (s, IH) .
6-pyrrolidin-l-yl-N*2*- (tetrahydro-furan-2-ylmethyl) - pyridine-2, 4-diamine (11) : ^- MR (400Mhz, CDC13) , ppm: 1.61- 2.0 (m, 8H) , 3.22 (m, 2H) , 3.41 (m, 4H) , 3.74 (m, 3H) , 3.84 (m, IH) , 4.08 (m, IH) , 4.18 (sb, IH) , 5.03 (s, IH) , 5.07 (s, IH)
N*2*- (4 -phenyl -butyl) -6-pyrrolidin-l-yl-pyridine-2 , 4-diamine (12) : ^-NMR (400Mhz, CDCl3) , ppm: 1.63 (m, 4H) , 1.84 (m, 4H) , 2.64 (t, 2H) , 3.17 (m, 2H) , 3.40 (m, 4H) , 3.76 (sb, 2H) , 4.20 (sb, IH) , 5.02 (s, IH) , 5.09 (s, IH) , 7.07-7.25 (m, 5H) .
N*2*-phenethyl-6-pyrrolidin-l-yl-pyridine-2, 4-diamine (13) :
^- MR (400Mhz, CDC13) , ppm: 1.91 (m, 4H) , 2.92 (t, 2H) , 3.37-3.48 (m, 6H) , 3.73 (sb, 2H) , 4.19 (sb, IH) , 5.08 (s, 2H) , 7.16-7.37 (m, 5H) .
2- (4-pyrimidin-2-yl-piperazin-l-yl) -6-pyrrolidin-l-yl- pyridin- 4 -ylamine (14) : ^-N R (400Mhz, CDC13) , ppm: 1.91 (m, 4H) , 3.27-3.93 (m, 12H) , 4.76 (sb, 2H) , 5.15 (s, IH) , 5.32 (S, IH) , 6.50 (t, IH) , 8.33 (d, 2H) . N*2*-methyl-N*2*-octadecyl-6-pyrrolidin-1-yl-pyridine-2 , 4- diamine (15): ^-NMR (400Mhz, CDC13) , ppm: 0.83 (s, 3H) , 1.22
(s, 32H) , 1.92 (m, 4H) , 2.96 (s, 3H) , 3.38-3.44 (m, 6H) , 3.66
(sb, 2H) , 5.04 (s, IH) , 5.17 (s, IH) .
4-methyl-6 ' -pyrrolidin-l-yl-3 ,4,5, 6-tetrahydro-2H- [1, 2 '] bipyridinyl-41 -ylamine (16): ^-NMR (400Mhz, CDC13) , ppm: 0.98 (d, 3H) , 1.68-2.09 (m, 9H) , 3.17 (m, 2H) , 3.65 (m, 4H) , 3.81 (sb, 2H) , 4.05 (m, 2H) , 4.91 (s, IH) , 4.96 (s, IH) .
6 ' -Pyrrolidin-l-yl-3, 4, 5, 6-tetrahydro-2H- [1,2 ' ] bipyridinyl- 4 ' -ylamine (19)
(4-Amino-6-pyrrolidin-l-yl-pyridin-2-ylamino) -acetic acid ethyl ester (20)
2-pyrrolidin-l-yl-6- (1,3,4, 9-tetrahydro-beta-carbolin-2-yl) - pyridin-4-ylamine (21)
[ (4-Amino-6-pyrrolidin-l-yl-pyridin-2-yl) -methyl-amino] - ' acetic acid ethyl ester (22)
2- (4-Methyl-piperazin-l-yl) -6-pyrrolidin-l-yl-pyridin-4- ylamine (23)
N-2-adamantan-l-yl-6-pyrrolidin-l-yl-pyridine-2 , 4-diamine (24)
N-2-adamantan-2-ylmethyl-6-pyrrolidin- 1-yl-py idine-2,4- diamine (25) N-2- (3-Methoxy-phenyl) -6-pyrrolidin-l-yl-pyridine-2 , 4-diamine (26)
N-2- (2-Methoxy-ethyl) -6-pyrrolidin-l-yl-pyridine-2 , 4-diamine (27)
N-2- (4-Methoxy-phenyl) -6-pyrrolidin-l-yl-pyridine-2 , 4-diamine (28)
N-2-octadecyl-6-pyrrolidin-l-yl-pyridine-2, 4-diamine (29)

Claims

1. A compound of formula (I)
∞ wherein n is 1 or 2 ;
R represents hydrogen or C1-C4 acyl;
Rl and R2 represent either both hydrogen or both halogen; R3 represents NR4R5 wherein:
a) R4 represents hydrogen or C1-C4 alkyl and R5 represents an unsubstituted or substituted C1-C18 n-alkyl or C3-C6 cycloalkyl ; or
b) R4 and R5 taken together with N to which they are linked represent an unsubstituted or substituted saturated 5 to 7 membered heterocyclic ring;
and the pharmaceutically acceptable salts thereof.
2. A compound of formula (I) as claimed in claim 1 wherein: n is 1;
R represents hydrogen; Rl and R2 represent either both hydrogen or both Cl; R3 represents NR4R5 wherein a') R4 represents hydrogen or C1-C4 alkyl, and R5 represents C1-C18 n-alkyl unsubstituted or substituted by adamantyl, 2 ' -tetrahydrofuryl, methylene dimethoxy CH(0Me)2, phenyl unsubstituted or substituted by C1-C4 alkoxy or C3-C6 cycloalkyl; or
b1) R4 and R5 taken together with N to which they are linked represent an unsubstituted or substituted 5-7 membered saturated heterocyclic ring represented by the following formulae:
wherein R6 represents hydrogen, methyl, phenyl, benzyl, ethoxycarbonyl (COOEt) or 2 -pyriminidinyl ; and the pharmaceutically acceptable salts thereof.
3. A compound of formula (I) as claimed in claim 1, whenever appropriate in the form a pharmaceutically acceptable salt, which is
1)2- (4-Phenyl-piperazin-1-yl) -6-pyrrolidin-l-yl-pyridin-4- y1amine; 2) N-2-Cyclohexyl-6-pyrrolidin-1-yl-pyridine-2 , 4-diamine; 3) N-2 -Cyclohexylmethyl-6-pyrrolidin-1-yl-pyridine-2 , 4- diamine; 4) 6-Pyrrolidin-1-yl-N-2- (3,4, 5-trimethoxy-benzyl) -pyridine-
2, 4-diamine;
5) N-2-Benzyl-6-pyrrolidin-1-yl-pyridine-2 , 4-diamine;
6) N-2- (2, 2 -Dimethoxy-ethyl) -6-pyrrolidin-1-yl-pyridine-2 , 4- diamine;
7) 2-piperazin-1-yl-6-pyrrolidin-1-yl-pyridin-4-ylamine;
8) 4- (4-Amino-6-pyrrolidin-l-yl-pyridin-2-yl) -piperazine-1- carboxylic acid ethyl ester;
9) N-2-methyl-6-pyrrolidin-1-yl-pyridine-2,4-diamine; 10) 2-Morpholin-4-yl-6-pyrrolidin-l-yl-pyridin-4-ylamine;
11) 6-Pyrrolidin-1-yl-N-2- (tetrahydro-furan-2-ylmethyl) - pyridine-2 , 4-diamine;
12) N-2- (4-Phenyl-butyl) -6-pyrrolidin-1-yl-pyridine-2 , 4- diamine; 13 ) N-2-Phenethyl-6-pyrrolidin-1-yl-pyridine-2 , 4-diamine;
14)2- (4-Pyrimidin-2-yl-piperazin-l-yl) -6-pyrrolidin-1-yl- pyridin-4-ylamine,•
15) N-2-Methyl-N-2 -octadecyl-6-pyrrolidin-1-yl-pyridine-2, 4- diamine; 16) 4-Methyl-6 ' -pyrrolidin-l-yl-3 , 4, 5, 6-tetrahydro-2H-
[1,2' ] bipyridinyl-4 ' -ylamine;
17)2, 6-Di-pyrrolidin-1-yl-pyridin-4-yl-amine;
18) N- (2, 6-Di-pyrrolidin-l-yl-pyridin-4-yl) -acetamide;
19) 6' -Pyrrolidin-1-yl-3, 4, 5, 6-tetrahydro-2H- [1,2'] bipyridinyl-4 ' -ylamine;
20) (4-Amino-6-pyrrolidin-l-yl-pyridin-2-ylamino) -acetic acid ethyl ester;
21) 2-pyrrolidin-l-yl-6- (1,3,4, 9-tetrahydro-beta-carbolin-2- yl) -pyridin-4-ylamine; 22) [ (4-Amino-6-pyrrolidin-l-yl-pyridin-2-yl) -methyl-amino] - acetic acid ethyl ester;
23) 2- (4-Methyl-piperazin-l-yl) -6-pyrrolidin-l-yl-pyridin-4- ylamine; 24) -2-adamantan-1-yl-6-pyrrolidin-1-yl-pyridine-2,4-diamine;
25) N-2-adamantan-2-ylmethyl-6-pyrrolidin-l-yl-pyridine-2 , 4- diamine;
26) N-2- (3-Methoxy-phenyl) -6-pyrrolidin-l-yl-pyridine-2, 4- diamine; 27)N-2- (2-Methoxy-ethyl) -δpyrrolidin -l-yl-pyridine-2 , 4- diamine;
28) N-2- (4-Methoxy-phenyl) -6-pyrrolidin-l-yl-pyridine-2 , 4- diamine;
29) -2-octadecyl-6-pyrrolidin-l-yl-pyridine-2 , 4-diamine; 30)3, 5-Dichloro-2- (4-phenyl-piperazin-l-yl) -6-pyrrolidin-1- yl-pyridin-4-ylamine;
31)3, 5-Dichloro-N-2-cyclohexyl-6-pyrrolidin-l-yl-pyridine-
2, 4-diamine;
32) 3 , 5-Dichloro-N-2-cyclohexylmethyl-6-pyrrolidin-l-yl- pyridine-2 , 4-diamine;
33) 3, 5-Dichloro-6-pyrrolidin-1-yl-N-2- (3,4, 5-trimethoxy- benzyl) -pyridine-2 , 4-diamine;
34) N-2-Benzyl-3 , 5-dichloro-6-pyrrolidin-1-yl-pyridine-2 , 4- diamine; 35)3, 5-Dichloro-N-2- (2 , 2-dimethoxy-ethyl) -6-pyrrolidin-l-yl- pyridine-2 , 4-diamine;
36)2- (4-Benzyl-piperazin-l-yl) -3 , 5-dichloro-6-pyrrolidin-l- yl-pyridin-4-ylamine;
37) 4- (4-Amino-3 , 5-dichloro-6-pyrrolidin-l-yl-pyridin-2-yl) - piperazine-1-carboxylic acid ethyl ester; 38) N-2-Benzyl-3,5-dichloro-N-2 -methyl-6-pyrrolidin-1-yl- pyridine-2 , 4-diamine;
39) 3 , 5-Dichloro-2-morpholin-4-yl-6-pyrrolidin-l-yl-pyridin-4- ylamine; 40)3, 5-Dichloro-6-pyrrolidin-l-yl-N-2- (tetrahydro-furan-2- ylmethyl) -pyridine-2 , 4-diamine;
41)3, 5-Dichloro-N-2- (4-phenyl-butyl) -6-pyrrolidin-l-yl- pyridine-2 , 4-diamine;
42) 3 , 5-Dichloro-N-2-phenethyl-6-pyrrolidin-1-yl-pyridine-2 , 4- diamine;
43) 3 , 5-Dichloro-2- (4-pyrimidin-2-yl-piperazin-l-yl) -6- pyrrolidin-1-yl-pyridin-4-ylamine;
44) 3 , 5-Dichloro-N-2-methyl-N-2-octadecyl-6-pyrrolidin-l-yl- pyridine-2 , 4-diamine; 45) 3 ' , 5 ' -Dichloro-4-methyl-6 ' -pyrrolidin-l-yl-3 , 4, 5,6- tetrahydro-2H- [1,2'] bipyridinyl-4 ' -ylamine;
46)3, 5-Dichloro-2 , 6-di-pyrrolidin-l-yl-pyridin-4-ylamine;
47) 3 ' , 5 ' -Dichloro-6 ' -pyrrolidin-l-yl-3 , 4 , 5, 6-tetrahydro-2H- [1,2'] bipyridinyl-4 ' -ylamine; 48)3, 5-Dichloro-2-fluoro-6-pyrrolidin-l-yl-pyridin-4-ylamine;
49) (4-Amino-3 , 5-dichloro-6-pyrrolidin-l-yl-pyridin-2- ylamino) -acetic acid ethyl ester;
50)3, 5-dichloro-2-pyrrolidin-l-yl-6- (1,3,4, 9-tetrahydro-beta- carbolin-2-yl) -pyridin-4-ylamine; 51) [ (4-Amino-3, 5-dichloro-6-pyrrolidin-l-yl-pyridin-2-yl) - methyl-amino] -acetic acid ethyl ester;
52) 3 , 5-dichloro-2- (4-Methyl-piperazin-l-yl) -6-pyrrolidin-1- yl-pyridin-4-ylamine;
53 ) N-2-adamantan-1-yl-3 , 5-dichloro-6-pyrrolidin-1-yl- pyridine-2 , 4-diamine; 54) -2-adamantan-2-ylmethyl-3,5-dichloro-6-pyrrolidin-1-yl- py idine-2,4-diamine;
55) 3, 5-dichloro-N-2- (3-Methoxy-phenyl) -6-pyrrolidin-1-yl- pyridine-2 , 4-diamine; 56) 3 , 5-dichloro-N-2- (2-Methoxy-ethyl) -6-pyrrolidin-l-yl- pyridine-2 , 4-diamine;
57) 3, 5-dichloro-N-2- (4-Methoxy-phenyl) -6-pyrrolidin-l-yl- pyridine-2 , 4-diamine; or
58)3, 5-dichloro-N-2-octadecyl-6-pyrrolidin-1-yl-pyridine-2 , 4- diamine .
4. A pharmaceutical composition, which comprises a therapeutically effective amount of a compound of formula (I) as defined in claim 1 or a pharmaceutically acceptable salt thereof as an active ingredient and a pharmaceutically acceptable excipient or carrier.
5. A compound of formula (I) as defined in claim 1 or a pharmaceutically acceptable salt thereof for use in the prevention, amelioration, and/or treatment of benign and malignant tumors or neoplasias.
6. A compound of formula (III)
(HI) wherein
Hal is Cl, Br, I or F,
R3 represents NR4R5 wherein:
R4 represents hydrogen or C1-C4 alkyl and R5 represents an unsubstituted or substituted C1-C18 n-alkyl or C3-C6 cycloalkyl; or
R4 and R5 taken together with N to which they are linked represent an unsubstituted or substituted saturated 5 to 7 membered heterocyclic ring.
EP03794969A 2002-09-10 2003-08-26 Substituted pyridine derivatives as antitumor agent Withdrawn EP1546129A1 (en)

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