JP2003532675A - A method for inhibiting cancer cell growth and inducing its apoptosis - Google Patents

Abstract

  (57) [Summary] Disclosed are a method for reducing the proliferation of adenocarcinoma cells, a method for inducing apoptosis of adenocarcinoma cells, and a method for inducing adenocarcinoma cells to differentiate into non-cancerous cells. One such method involves contacting adenocarcinoma cells with a compound under conditions effective to inhibit binding of leukotriene B4 to the leukotriene B4 receptor. Another such method is to treat adenocarcinoma cells with 2- (2-propyl-3- (3- (2-ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy) phenoxy) benzoic acid Or contacting it with a pharmaceutically acceptable salt, solvate, or congener thereof. Also disclosed is a method of treating adenocarcinoma in a subject. One method involves administering to a subject an amount of a compound effective to inhibit binding of leukotriene B4 to the leukotriene B4 receptor.

Description

Detailed Description of the Invention

The present invention relates to the National Cancer Institute Contract No.
Made with the support of P50CA72712. The federal government may have certain rights in the invention.

FIELD OF THE INVENTION The present invention relates generally to methods of reducing the growth of cancer cells, or inducing apoptosis of cancer cells, or inducing the differentiation of cancer cells into non-cancerous cells, and the gland of a subject. It relates to a method of treating cancer.

[0003] Pancreatic cancer of the invention is one of the most baffling and aggressive malignant disease faced by oncologists (
Parker et al., "Cancer Statistics, 1996" CA Cancer J. Clin., 46: 5-27 (1996) ("Parker"). It is now the fourth leading cause of cancer death in both men and women in the United States, and the incidence of this disease has increased significantly over the last 20 years (Parker, Tolede et al., "Survival Rates After Pancreaticoduodenectomy". : 11 without death due to surgery
Surg., 211: 447-458 (1990), Cameron et al., 145 consecutive pancreaticoduodenectomy without death Ann. Surg., 217: 430-438 (1993), Holward. , "Adenocarcinoma of the pancreas" Curr. Prob. In Cancer, 20: 286-293 (1996) ("Hollward"), Poston et al., Gut. Biology of Pancreatic Cancer, 32: 800-812 (
1991) ("Poston"), and Black et al., "Treatment of pancreatic cancer: current constraints, future potential", Oncology, 10: 301-307 (1996) ("Black"). Pancreatic cancer is responsible for 27,000 deaths annually in the United States. Due to lack of early diagnosis of pancreatic cancer and poor response to treatment, less than 2% of patients survive more than 5 years and expected life expectancy after diagnosis of pancreatic cancer is less than 6 months (( , Poston, and Black).

[0004]   Colon cancer is the second most common form of cancer in the United States (Dol et al.
Mortality in Japan: A 20-Year Observation of Male British Physicians, ”BMJ, 2: 1525-1536 (1
976), Fulvan et al., "Molecular Diagnosis of Metastases in Cancer and Microcancer Tissues" Adv. Anat. Pathol.
, 5: 175-178 (1998) ("Furuvan"), Fig-Eled et al., "The second post-complete resection.
Adjuvant Treatment of Tage Colon Cancer, Rural Gastrointestinal Site Group "Cancer Prev.
 Control, 1: 379-92 (1997) ("Figueredo"), Ness et al. "Final colorectal cancer.
Status: Confirmation and description using patient focus groups ”, Am, J. Gastroenterol
., 93: 1491-7 (1998) ("Ness"), Trehue et al., "Screening colorectal cancer.
Costs: Results of Community Mass Screening Programs and Literature Review ”So
uth Med. J., 85: 248-253 (1992), and Wingo et al., Cancer Statistics, CA Cancer J.
 Clin., 45: 8-30 (1995) ("Wingo"). Colon cancer occurs in the United States every year at 138,0.
More than 00 patients have occurred, causing over 55,000 deaths (
Wingo). Up to 70% of colon cancer patients have liver metastases by the time of death
There is. This suggests that there are undetectable micrometastases at the time of surgery
(Furuban, Figuered, and Nes). Furthermore, metastatic cancers are often standardized
It does not respond to the chemotherapeutic regimen, resulting in treatment failure (figured and nes).
The overall response to chemotherapeutic agents in patients with advanced or unresectable colorectal cancer is 26
% To 44%. For example, one-third of colorectal cancer patients with liver metastases
Less than responds to treatment with agents such as 5-FU and leucovorin ( Id ． ).

Breast cancer is diagnosed in over 275,000 people each year in the United States,
Having the highest incidence of cancer in women (Richards et al., “The effect of delay on survival in patients with breast cancer: a systematic review” Lancet, 353: 1119-26 (1999), Norton, “ Adjuvant treatment: current and future strategies---Breast cancer growth kinetics and improved drug treatment "Semin. Oncol., 26: 1-4 (1999), Morrow et al.,"Today's Controversy in Breast Cancer Treatment, "Curr. Probl. Surg., 36: 163-216 (1999), and Rupert et al., "
Breast Cancer Res. Treatment, 44: 93-114 (1
997)). Even though the 5-year survival rate has increased to over 80%, over 77,000 women are still dying of this disease each year ( Id .).

Thus, another dimension of chemotherapeutic agents for pancreatic, colon, and breast cancer would be of great benefit, especially for controlling metastatic or unresectable disease.

SUMMARY OF THE INVENTION The present invention relates to a method of reducing proliferation of adenocarcinoma cells, or of inducing apoptosis of adenocarcinoma cells, or of inducing differentiation of adenocarcinoma cells into non-cancerous cells. A sample containing adenocarcinoma cells is contacted with a compound that is under conditions effective to inhibit the binding of leukotriene B4 to the leukotriene B4 receptor.

The invention also relates to a method of treating adenocarcinoma in a subject. The method comprises the step of administering to the subject an amount of a compound effective to inhibit the binding of leukotriene B4 to the leukotriene B4 receptor.

The present invention also relates to a method of reducing proliferation of adenocarcinoma cells, or of inducing apoptosis of adenocarcinoma cells, or of inducing differentiation of adenocarcinoma cells into non-cancerous cells. This method involves converting a sample containing adenocarcinoma cells to the following formula (“Formula I”):

[0010]

[Chemical 13]

Wherein R ₁ is C ₁ -C ₅ alkyl, C ₂ -C ₅ alkenyl, C ₂ -C ₅ alkynyl, C ₁ -C ₄ alkoxy, (C ₁ -C ₄ alkyl) ) Thio, halo, or R ₂ -substituted phenyl, wherein each of R ₂ and R ₃ is independently hydrogen, halo, hydroxy, C ₁ -C ₄ alkyl, C ₁ -C ₄ alkoxy, (C ₁ -C 2 ₄ alkyl) -S (O) _q -, trifluoromethyl, or di - (a C ₁ -C ₃ alkyl) amino, X is -O -, - S -, - C (= O), or —CH ₂ — and Y is —O
- or -CH ₂ -, or when taken together -X-Y- is -CH = CH- or -
C≡C—, Z is a linear or branched C _{1 to} C ₁₀ alkylidenyl, A is a single bond, —O—, —S—, —CH═CH—, or —CR _a R _b −,
R _a and R _b are each independently hydrogen, C ₁ -C ₅ alkyl, or R ₇ -substituted phenyl, or R _a and R _b together with the carbon atom to which they are attached are C _{4 to} C ₈ cycloalkyl ring is formed, R ₄ is R ₆ or R ₄ is the following formula:

[0012]

[Chemical 14]

[0013]

[Chemical 15]

Wherein R ₆ is independently —COOH, 5-tetrazolyl, —CON (R ₉ ) ₂ or —CONHSO ₂ R ₁₀ , and R ₇ is each is hydrogen, alkyl of C ₁ -C _4, alkenyl C ₂ -C _5, alkynyl C ₂ -C _5, benzyl, _{methoxy, -W-R 6, -T-} G-R 6, (
C ₁ -C ₄ alkyl) -T- (C ₁ -C ₄ alkylidenyl) -O-, or hydroxy, R ₈ is hydrogen or halo, and each R ₉ is independently hydrogen, phenyl, or C ₁ -C ₄ alkyl,
Or R ₉ is taken together with the nitrogen atom to which it is attached to form a morpholino group, a piperidino group, a piperazino group, or a pyrrolidino group, and R ₁₀ is a C ₁ -C ₄ alkyl or phenyl , R ₁₁ is R ₂ , —W—R ₆ , or —T—G—R ₆ , and each W is a single bond or a linear or branched divalent hydrocarbyl radical having 1 to 8 carbon atoms. And each G is a linear or branched divalent hydrocarbyl radical having 1 to 8 carbon atoms, and each T is a single bond, —CH ₂ —, —O—, —NH—, —NHCO—, -C
(= O)-, or -S (O) _q- , K is -C (= O)-, or -CH (OH)-, and each q is 0, 1, or 2 independently. , P is 0 or 1, and t is 0 or 1, provided that when X is -O- or -S-, then Y is not -O-, and A is -O-. Or -S-, R ₄ is not R ₆ , and when A is -O- or -S- and Z is a single bond, Y is not -O-, and When p is 0, W is not a single bond, and the step of contacting with a compound having the formula: or a pharmaceutically acceptable salt or solvate thereof.

The present invention also relates to a method of treating adenocarcinoma in a subject. The method comprises the step of administering to the subject a therapeutically effective amount of a compound of Formula 1, or a pharmaceutically acceptable salt or solvate thereof.

[0016] METHOD The invention relates to reducing the growth of an adenocarcinoma cell or a method of inducing apoptosis of adenocarcinoma cells, or to a method of inducing differentiation into non-cancerous cells of adenocarcinoma cells. The method comprises contacting a sample containing adenocarcinoma cells with a compound that inhibits binding of leukotriene B4 to leukotriene B4 receptors.

“Inhibit” and its alternative forms (eg, “inhibiting”) means complete block (1
To any extent (eg, greater than about 5%, greater than about 10%, greater than about 20%, greater than about 30%, greater than about 40%, greater than about 50%, about 100% inhibition). Greater than about 60%, greater than about 70%, greater than about 80%, greater than about 90%, greater than about 95%, greater than about 98%, greater than about 99% inhibition), and completely blocked (100). % Inhibition) is also included.

The mechanism by which the binding of leukotriene B4 to the leukotriene B4 receptor is inhibited is not particularly important for the practice of the present invention. This inhibition occurs when the compound is leukotriene B.
4 can be direct, as in the case of directly impeding the binding of 4 to the leukotriene B4 receptor (eg, by directly binding to the leukotriene B4 receptor, as described in more detail below), or Inhibition can also be indirect, such as where the compound interferes with the production of either leukotriene B4 and / or leukotriene B4 receptors, as described further below.

For example, the compound can inhibit the binding of leukotriene B4 to the leukotriene B4 receptor by binding to the leukotriene B4 receptor.
The binding of the compound to the leukotriene B4 receptor is preferably specific, ie the compound does not substantially bind to other biological substances present in the sample. For example, the IC ₅₀ of the compound for the leukotriene B4 receptor is preferably greater than about 1.1 times, and greater than about 1.5 times the IC ₅₀ of the compound for all other biological substances in the sample. More preferably, it is more preferably greater than about 2 times, even more preferably greater than about 5 times, even more preferably greater than about 10 times. The degree of binding of the compound to the leukotriene B4 receptor is preferably greater than the binding of leukotriene B4 to the leukotriene B4 receptor (ie, has a higher IC ₅₀ ), but the compound to the leukotriene B4 receptor is preferred. If the extent of binding of less than binding to leukotriene B4 receptor leukotriene B4 (i.e., if with lower IC ₅₀₎ even, under certain circumstances be achieved the desired effect.

“Binding” as used herein is meant to include irreversible and reversible binding. The bond can be the result of thermodynamic forces (eg, the preferred equilibrium constant) or kinetic reasons (eg, the preferred on / off ratio constant), and the bond is a chemical interaction (eg, a covalent bond). , Ionic bonds, hydrogen bonds, van der Waals forces, chelation, π-σ bonds, and / or π-π bonds) or physical interactions (eg, surface phenomena, enclosures, etc.). When used in the context of “binding” of a binding partner and the compound (eg, binding of the compound to the leukotriene B4 receptor), “binding” results in the ability of the binding partner to play its physiological role. It is also meant to include any interaction of the compound with its binding partner that reduces. For example, when the binding partner is the leukotriene B4 receptor, "binding" of the compound to the leukotriene B4 receptor results in a physical, chemical and / or other change in the leukotriene B4 receptor. , Which in turn is meant to include any interaction or combination of interactions between the compound and the leukotriene B4 receptor that reduces the ability of the leukotriene B4 receptor to bind to leukotriene B4.

Various compounds can be used to inhibit the binding of leukotriene B4 to the leukotriene B4 receptor by binding to the leukotriene B4 receptor. Examples of such compounds include those of the following formula (“formula I”)

[0022]

[Chemical 16]

And a pharmaceutically acceptable salt or solvate thereof. In the above formula, R ₁ is C ₁ -C ₅ alkyl, C ₂ -C ₅ alkenyl, C ₂ -C ₅ alkynyl, C ₁ -C ₄ alkoxy, (C ₁ -C ₄ alkyl) thio. , Halo, or R ₂ -substituted phenyl. R ₂ and R ₃ are each independently hydrogen, halo, hydroxy, C ₁ -C ₄ alkyl, C ₁ -C ₄ alkoxy, (C ₁ -C ₄ alkyl) -S (O) _q- , Trifluoromethyl or di- (C ₁ -C ₃ alkyl) amino. X is, -O -, - S -, - C (= O) -, or -CH ₂ -, and Y is, -O- or -CH ₂ -, or when taken together -X-
Y- is -CH = CH- or -C≡C-. Z is a linear or branched C _{1 to} C ₁₀ alkylidenyl. A is a single bond, -O-, -S-, -CH = CH-,
Or —CR _a R _b —, each of R _a and R _b is independently hydrogen, C ₁ -C ₅ alkyl, or R ₇ -substituted phenyl, or R _a and R _b are Forming a C ₄ -C ₈ cycloalkyl ring. R ₄ is R ₆ (as defined below) or R ₄ is of the formula

[0024]

[Chemical 17]

[0025]

[Chemical 18]

It is a part having one of In the above formula, each of R _6, independently -COOH, 5-tetrazolyl, -CON (
R ₉ ) ₂ or —CONHSO ₂ R ₁₀ . Each of R ₇ is hydrogen, C ₁ ~
Alkyl C _4, alkenyl C ₂ -C _5, alkynyl C ₂ -C _5, benzyl, _{methoxy, -W-R 6, -T-} G-R 6, ( alkyl C ₁ ~C ₄₎ -T -(
C alkylidenyl of _{1 ~C 4)} -O-, or hydroxy. R ₈ is hydrogen or halo. Each of R ₉ is independently hydrogen, phenyl, or C ₁ -C ₄ alkyl, or R ₉ together with the nitrogen atom to which it is attached is a morpholino group, piperidino group, piperazino group, or Form a pyrrolidino group. R ₁₀ is C ₁ -C ₄ alkyl or phenyl. R ₁₁ is R ₂ , -W-R ₆ , or -T-G
-R _6. Each W is a single bond or a linear or branched divalent hydrocarbyl radical having 1 to 8 carbon atoms. Each G is a linear or branched divalent hydrocarbyl radical having 1 to 8 carbon atoms. Single bond each _{T, -CH 2 -, - O} -, - NH -, - NHCO -, - C (= O) -, or -S
(O) _q −. K is -C (= O)-or -CH (OH)-. Each of q is independently 0, 1, or 2, p is 0 or 1, and t is 0 or 1. Preferably, when X is -O- or -S-, Y is not -O- and when A is -O- or -S-, R ₄ is not R ₆ and A is-. When O- or -S-, Z is a single bond, Y is not -O-, and when p is 0, W is not a single bond.

The term “C ₁ -C ₆ alkyl” means methyl, ethyl, propyl, isopropyl, n-butyl, sec-butyl, tert-butyl, n-pentyl, 2,2-
It means linear and branched aliphatic radicals having 1 to 6 carbon atoms such as dimethylpropyl and hexyl. Some of this definition, "alkyl C ₁ -C _3" refers "alkyl of C ₁ -C _4", and "alkyl of C ₁ -C _5" are also included. The term "alkenyl C ₂ -C _{5" is, -CH = CH 2, -CH 2} -CH = CH 2, -CH 2 -CH 2 -CH = CH 2, -CH-C (CH 3) = CH _2, -CH ₂ -CH =
By straight and branched chain aliphatic radicals of 2 to 5 carbon atoms containing one double bond, such as C (CH ₃ ) ₂ .

The term “C ₂ -C ₅ alkynyl” means —C≡CH, —CH ₂ —C≡CH, —
_{CH 2 -CH 2 -C≡CH, -CH 2} -CH (CH 3) -C≡CH, -CH 2 -
It refers to straight and branched chain aliphatic residues of 2 to 5 carbon atoms containing one triple bond such as C≡C—CH ₃ . The term "C ₁ -C ₄ alkoxy" refers to, for example, methoxy, ethoxy, propoxy, isopropoxy, butoxy, sec-butoxy,
And tert-butoxy. The term "halo" means, for example, fluoro, chloro, bromo, and iodo.

The term “C ₁ -C ₁₀ alkylidenyl” refers to —CH ₂ —, —CH (CH ₃ ) —.
_{, -C (CH 3) 2 -} , - CH (C 2 H 5) -, - CH 2 -CH 2 -, - CH 2 -CH (CH 3) -, - CH (CH 3) -CH 2 -, -CH (CH ₃₎ -CH (
_{CH 3) -, - CH 2} -CH (C 2 H 5) -, - CH 2 -CH 2 -CH 2 -, -
_{CH (CH 3) -CH 2 -CH} 2 -, - CH 2 -CH (CH 3) -CH 2 -, -
_{CH 2 -CH (C 2 H 5} ) -CH 2 -, - CH 2 -CH 2 -CH (C 2 H 5) -
_{, -C (CH 3) 2 -CH} 2 -CH 2 -, - CH (CH 3) -CH 2 -CH (C
_{H 3) -, - CH 2} -CH 2 -CH 2 -CH 2 -, - CH 2 -C (CH 3) 2 -
_{CH 2 -CH 2 -, - CH} 2 -C (CH 3) 2 -CH 2 -, - CH 2 -CH 2 -
_{CH (C 2 H 5) -CH} 2 -, - CH 2 -CH 2 -CH 2 -CH 2 -CH 2 -,
_{-CH (CH 3) -CH 2 -CH} 2 -CH 2 -CH 2 -, - CH 2 -CH 2 -C
_{H 2 -CH 2 -CH 2 -CH 2} -, - (CH 2) 10 - means a divalent radical derived from an alkane of C ₁ -C _10, such as. Within this definition, the term "C ₁ ~
C ₄ alkylidene "and includes" alkylidene of C ₂ -C ₄ ".

The term “C ₄ -C ₈ cycloalkyl” includes cyclobutyl, cyclopentyl,
It means a cycloalkyl ring of 4 to 8 carbon atoms such as cyclohexyl, 4,4-dimethylcyclohexyl, cycloheptyl, cyclooctyl and the like. The term "linear or branched divalent hydrocarbyl residue of 1 to 8 carbon atoms" means a divalent radical derived from a linear or branched alkane, alkene, or alkyne of 1 to 8 carbon atoms. .

As will be appreciated by organic chemists, depending on the number of carbon atoms and their branching, such moieties may contain one, two, or three double or triple bonds or a combination of both. it can. As such, the term contains from 1 to 8 carbon atoms as defined above and is optionally limited to 1 to 3 double bonds or as defined in the preceding sentence. It can be considered as an alkylidene group containing a triple bond or a combination of both.

As mentioned above, it is also possible to use pharmaceutically acceptable base addition salts of the compounds of formula I. Such salts include salts derived from inorganic bases such as ammonium, alkali and alkaline earth metal hydroxides, carbonates, bicarbonates, as well as aliphatic or aromatic amines, aliphatic diamines, hydroxys. Included are salts derived from basic organic amines such as alkylamines. Bases useful in preparing salts useful in the practice of the present invention include ammonium hydroxide, potassium carbonate, sodium bicarbonate,
It includes calcium hydroxide, methylamine, diethylamine, ethylenediamine, cyclohexylamine, ethanolamine and the like. The potassium and sodium salt forms are particularly preferred.

The present invention relates to the monosalt form, that is to say a 1: 1 ratio of the compound of the formula I to the base mentioned above, as well as the disalt form when the compound of the formula I has two acidic groups. Including the use of both. Further, the methods of the present invention can be practiced with solvated forms of the compounds of formula I or salts thereof, such as ethanol solvation, hydration and the like.

It will be appreciated that various stereoisomeric products may exist in compounds with branched alkyl, alkylidenyl, or hydrocarbyl functional groups, and in compounds with double or triple bonds. The process of the present invention is not limited to any particular stereoisomer and includes all possible individual isomers and mixtures thereof. The term "5-tetrazolyl" refers to both tautomers, namely (1H) -5-tetrazolyl and (2
H) -5-tetrazolyl.

Examples of compounds that can be used to practice the method of the present invention include 2- (2-propyl-
3- (3- (2-Ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy) phenoxy) benzoic acid, 3- (2- (3- (2-ethyl-
4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy) -6
-(4-Carboxy-phenoxy) phenyl) propionic acid, 1- (4- (carboxy-methoxy) phenyl) -1- (1H-tetrazol-5-yl) -6-
(2-Ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) hexane, 3- (4- (7-carboxy-9-oxo-3- (3- (2-ethyl-
4- (4-fluorophenyl) -5-hydroxyphenoxy) -propoxy)-
9H-xanthene)) propanoic acid, 5- (3- (2- (1-carboxy) -ethyl) -4- (3- (2-ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy. ) Phenyl) -4-pentynoic acid,
Included are pharmaceutically acceptable salts or solvates thereof, and combinations of these compounds, salts, and / or solvates. The present invention provides 2- (2-propyl-3-
Preference is given to working with (3- (2-ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy) phenoxy) benzoic acid or a pharmaceutically acceptable salt or solvate thereof.

The compounds of formula I may be synthesized by established methods such as those described by Baker et al., US Pat. No. 5,462,954, and Fleish et al., US Pat. No. 5,910,505. You can The above patents are incorporated herein by reference.

By binding itself to the leukotriene B4 receptor, leukotriene B4
Other compounds that can be used to inhibit the binding of Escherichia coli to the leukotriene B4 receptor include Fleish et al., US Pat. No. 5,998,454, Fleish et al., US Pat. U.S. Pat. No. 5,817,864, Anderskewitz et al., U.S. Pat. No. 6,037,377, Parchonock et al. Structure-Activity Relationship Study: A Novel Class of Leukotriene Antagonists "
J. Med. Chem., 29: 1442-1452 (1986), Gleason et al., "Letter to the Editor," J. M.
ed. Chem. 30: 959-961 (1987), Konno et al., "Synthesis and Structure-Activity Relationships of a Series of Substituted Phenylpropionic Acids as a Novel Class of Leukotriene B4 Antagonists," Adv. Prostaglandin Thromboxane Leukot. Res., 21A: 411. -414 (1991), Danes et al., "(E) -3-[[[[6- (2-carboxyethenyl) -5-[[8- (4-methoxyphenyl
) Octyl] oxy] -2-pyridinyl] -methyl] thio] methyl] benzoic acid: a novel high affinity leukotriene B4 receptor antagonist ”J. Med. Chem., 36 (18): 2
703-2705 (1993) (Error correction, J. Med. Chem., 36 (25): 4130 (1993), Schauer et al. "Preclinical pharmacological profile of the potent and selective leukotriene B4 antagonist CP-195543". J. Pharmacol. Exp. Ther., 285 (3): 346-954 (1998)
And Flot et al., "New Leukotriene B4 Receptor Antagonist (1S ^* , 3S ^* )-1-Hydroxy-3-[(3R ^* S ^* , E) -3-Hydroxy-7-phenyl-1-heptane-1"
-Ill] -1-Synthesis and Biochemical / Pharmacological Profiles of Sodium Cyclohexane Acetate, Arzneimittelforschung, 47 (1): 51-58 (1997), are other leukotriene B4 receptor antagonists such as those described in included. The above publications are incorporated herein by reference.

Still other compounds that can be used to inhibit the binding of leukotriene B4 to the leukotriene B4 receptor by binding to the leukotriene B4 receptor include antibodies (monoclonal or polyclonal) as well as leukotriene B4 receptor. Examples include peptide drugs such as mimotopes that recognize the body and bind to it. Methods for preparing and using such antibodies and mimotopes are well known to those skilled in the art, and the preparation of antibodies and mimotopes that recognize and bind to leukotriene A4 hydrolase will be described below.

Also, the binding of leukotriene B4 to the leukotriene B4 receptor can be inhibited by using a compound that binds to leukotriene B4 and forms, for example, a complex that cannot bind to the leukotriene B4 receptor. it can. It is preferred that the binding of this compound to leukotriene B4 is specific, ie the compound does not substantially bind to other biological substances present in the sample. For example, the IC ₅₀ of the compound for leukotriene B4 is preferably greater than about 1.1 times the IC ₅₀ of the compound for all other biomaterials in the sample, about 1.
More preferably, it is greater than 5 times, more preferably greater than about 2 times, even more preferably greater than about 5 times, even more preferably greater than about 10 times. The degree of binding of the compound to leukotriene B4 is preferably greater than the binding of leukotriene B4 to the leukotriene B4 receptor (ie, has a higher IC ₅₀ ), but the degree of binding of the compound to leukotriene B4 is In some circumstances, the desired effect can be achieved even when the binding of leukotriene B4 to the leukotriene B4 receptor is less than (ie, has a lower IC ₅₀ ).

Moreover, the binding of leukotriene B4 to the leukotriene B4 receptor can be inhibited by inhibiting the production of leukotriene B4. Production of leukotriene B4 can be inhibited, for example, by inhibiting one or more of the steps in the leukotriene B4 synthetic pathway.

For example, the compound can be one that inhibits the conversion of leukotriene A4 to leukotriene B4.

Illustratively, the conversion of leukotriene A4 to leukotriene B4 binds to leukotriene A4, forming a complex that is less susceptible to (eg, less than leukotriene A4) hydrolysis by, for example, leukotriene A4 hydrolase. It can be inhibited by contacting the sample with the compound.

Also illustratively, the conversion of leukotriene A4 to leukotriene B4 can be inhibited by contacting a sample with a compound that inhibits the activity of leukotriene A4 hydrolase.

As will be appreciated by those in the art, the activity of leukotriene A4 hydrolase can be inhibited by contacting the sample with a compound that binds to leukotriene A4 hydrolase. Such leukotriene A4 hydrolase inhibitors may traditionally be small molecules, or pharmaceutically acceptable salts thereof. Examples of such leukotriene A4 hydrolase inhibitors include 3-oxiranyl benzoic acid and their derivatives, Lone-Poulan Roller RP-64996, and (2S) -2-amino-3-phenylpropyl mercaptan, 2-amino. -2- (4'-benzyloxy) phenylpropyl mercaptan, and 2- such as (2S) -3- (4- (2-naphthylmethyloxy) phenyl) -1,2-diamino-propane HCl.
Amino-3-phenylpropane derivatives and their corresponding 4'-benzyloxy derivatives are mentioned. Other leukotriene A4 hydrolase inhibitors are, for example:
Julic et al., US Pat. No. 5,990,326, Isaxon et al., US Pat. No. 5,990,148, Yuan et al., US Pat. No. 5,455,271, International Patent Publication Nos. WO96 / 11192 and WO96 /. 1099, Evans et al., Prostaglandins, Leukotrienes and Drugs, 23: 167-171 (1986), Yuan et al., "Isolation and Identification of Metabolites of the Leukotriene A4 Hydrolase Inhibitor SC-57461 in Rats" Drug Metab. Dispos., 24 (10): 1124-1133 (1996), Tsuji et al., "Effect of SA6541 which is a leukotriene A4 hydrolase inhibitor and indomethacin on carrageenan-induced murine dermatitis," Eur. J. Pharmacol., 346 (1). : 81
-85 (1998), and Penning et al., “1- [2- (4-phenylphenoxy) ethyl] pyrrolidin (SC-22716), a potent inhibitor of leukotriene A (4) (LTA (4)) hydrolase. Structure-Activity Relationship Studies ", J. Med. Chem., 43 (4): 721-735 (2000). The above references are incorporated herein by reference.

The leukotriene A4 hydrolase inhibitor may also be present in the form of a peptide drug product. In general, see below. Beban et al., Trends in Biotechnology, 13 (3).
: 115-121 (1995), Sepetov et al., Proc. Nat'l Acad. Sci. USA, 92: 5426-5430.
(1995), O'Connor et al., Cancer Chemother. Pharmacol., 34: 225-229 (1994), and Webber et al., J. Med. Chem., 36: 733-746 (1993). These documents are incorporated herein by reference.

Drugs such as peptide drugs can be prepared by a variety of methods known in the art. One such method utilizes the development of epitope libraries and the biopanning of bacteriophage libraries.
Briefly, attempts to establish binding sites for various monoclonal antibodies have led to the development of epitope libraries. Palmley et al., Gene, 73
: 305-318 (1988) ("Permley") describes a bacteriophage expression vector capable of displaying foreign epitopes on its surface. This document is incorporated herein by reference. This vector is short (eg 6 amino acids)
It could be used to construct a large assembly of bacteriophages that would contain practically all possible sequences of peptides. Permley, incorporated herein by reference, also describes biopanning, a method of affinity purification of phage displaying foreign epitopes using specific antibodies. For example,
Palmley, Willa et al., Proc. Nat'l Acad. Sci. USA, 87: 6378-6382 (1990),
Scott et al., Science, 249: 386-390 (1990), Christian et al., J. Mol. Biol.,
227: 711-718 (1992), and Smith et al., Methods in Enzymology, 217: 228-257.
(1993) ("Smith"). The above references are incorporated herein by reference.

After developing the epitope library, Smith (incorporated herein by reference), in order to identify the epitope from all possible sequences of a given length, bacteriophage expression vectors and It has been suggested that it should be possible to use the biopanning technology of Palm Lee (incorporated herein by reference). This has led to the idea of identifying peptide ligands for antibodies by biopanning the epitope library. This could then be used for vaccine design, epitope mapping, gene identification, and many other applications (Palmley and Scott, Trends in Biochem. Sci.
., 17: 241-245 (1992), which is incorporated herein by reference.
).

Researchers searching for epitope sequences using epitope libraries and biopanning have found that they are not epitopes, but peptide epitopes that mimic epitopes, ie, sequences that are contiguous with linear, natural sequences. We have found sequences that did not occur or sequences that did not necessarily appear within the native protein sequence. These mimicking peptides are called mimotopes. In this way, mimotopes / proteins with different binding sites have been discovered. Larocca et al., Hybridoma, 11: 191-201 (1992) describe mimotope expression of human mammary epithelial mucin tandem repeats in E. coli. This document is incorporated herein by reference. Barras et al., Proc. Nat'l
Acad. Sci. USA, 90: 10638-10642 (1993) ("Varas") identifies a hexapeptide that mimics the conformation-dependent binding site of the acetylcholine receptor. This document is incorporated herein by reference. Hobart et al., Proc
R. Soc. London. B, 252: 157-162 (1993) discloses the isolation of mimotopes that mimic the C6 epitope, the epitope for the sixth component of complement. This document is incorporated herein by reference.

The sequences of these and other mimotopes are, by definition, inconsistent with the contiguous, linear, native sequence, or necessarily in any form in a naturally occurring molecule, eg, a naturally occurring protein. It does not appear. The mimotope sequence only forms a peptide that functionally mimics the binding site on the naturally occurring protein.
For example, the mimotope described in Barras mimics the binding site of the acetylcholine receptor.

Many of these mimotopes are short peptides. The availability of short peptides that can be easily synthesized in large quantities and mimic the naturally-occurring sequence (ie, the binding site) is
It can be utilized in the method of the present invention.

For example, this technique can be used to identify mimotopes to monoclonal antibodies that recognize leukotriene A4 hydrolase. The sequences of these mimotopes represent short peptides, which can then be used in various ways, for example as peptide drugs that bind to leukotriene A4 hydrolase and reduce the activity of leukotriene A4 hydrolase. Once the mimotope sequence is determined, the peptide drug can be chemically synthesized.

Antibodies to leukotriene A4 hydrolase represent another class of compounds useful in inhibiting the conversion of leukotriene A4 to leukotriene B4 by inhibiting the activity of leukotriene A4 hydrolase. "Antibody" as used herein is meant to include antibody fragments such as Fab, Fab2, and Fd fragments, as well as humanized versions. Humanized versions of the antibodies can be made using one of the procedures known in the art such as chimerization. The antibody is leukotriene A
It binds to 4-hydrolase and reduces the activity of leukotriene A4 hydrolase. Suitable antibodies include polyclonal and monoclonal antibodies.

Monoclonal antibodies that bind leukotriene A4 hydrolase can be produced by hybridomas. Hybridomas are immortal cell lines capable of secreting specific monoclonal antibodies.

Generally, techniques for preparing polyclonal and monoclonal antibodies as well as hybridomas capable of producing the desired antibody are well known in the art. For example, Campbell “Monoclonal Antibody Technology: Laboratory Technology in Biochemistry and Molecular Biology, Amsterdam, The Netherlands: Elswil Science Publishers (198).
4) ("Campbell"), and St. Gross et al. J. Immunol. Methods, 35: 1-21.
See (1980). These documents are incorporated herein by reference. Any animal known to produce antibodies (mouse, rabbit, etc.) can be immunized with the antigenic leukotriene A4 hydrolase (or antigenic fragment thereof). Methods of immunization are well known in the art. Such methods include subcutaneous or intraperitoneal injection of the enzyme. Those skilled in the art will appreciate that the amount of the enzyme used for immunization will vary depending on the animal immunized, the antigenicity of the enzyme, and the site of injection.

The enzyme used as an immunogen may be modified to enhance the antigenicity of the enzyme or administered in admixture with an adjuvant. Methods of increasing the antigenicity of an enzyme are well known in the art and include heterologous proteins (such as globulin or beta-galactosidase).
With, but not limited to, a method of binding an antigen with an antigen, or a method of including an adjuvant during immunization.

For monoclonal antibodies, splenocytes were removed from the immunized animal and S
A hybridoma cell producing a monoclonal antibody is obtained by fusing with a myeloma cell such as P2 / O-Ag 15 myeloma cell.

Any one of several methods well known in the art can be used to identify hybridoma cells that produce antibodies with the desired properties. These are, for example, E
Screening hybridomas using LISA assay, Western blot analysis, or radioimmunoassay. For example, Lutz et al., Exp. Cell Res.,
175: 109-124 (1988). It is incorporated herein by reference.

Hybridomas secreting the desired antibody are cloned and their class and subclasses are determined using procedures known in the art, such as those described by Campbell. This procedure is incorporated herein by reference.

For polyclonal antibodies, antisera containing antibody is isolated from the immunized animal and screened for the presence of antibodies with the desired specificity using one of the procedures described above.

Although this is also an example, conversion of leukotriene A4 to leukotriene B4 is
The leukotriene A4 hydrolase can be inhibited by contacting the sample with a nucleic acid molecule that binds to the nucleic acid molecule that encodes the leukotriene A4 hydrolase, thus
The expression of hydrolases can be reduced.

Suitable nucleic acid molecules include, for example, antisense nucleic acid molecules and ribozymes. Antisense nucleic acid molecule is a mR encoding leukotriene A4 hydrolase
Complementary to at least a portion of NA. The nucleic acid and amino acid sequences of leukotriene A4 hydrolase are known. For example, Medina et al., "Molecular cloning and expression of mouse leukotriene A4 hydrolase cDNA," Biochem.
Biophys. Res. Commun., 176: 1516-1524 (1991), Minami et al., "Molecular cloning of cDNA encoding human leukotriene A4 hydrolase: complete primary structure of enzyme involved in eicosanoid synthesis", J. Biol. Chem., 262: 13873-13876 (
1987), Funk et al., "Molecular cloning and amino acid sequence of leukotriene hydrolase," Proc. Nat'l Acad. Sci. USA, 84: 6677-6681 (1987), Mantini et al.
Cloning and Characterization of the Human Leukotriene A4 Hydrolase Gene "Eur.
J. Biochem., 231 (1): 65-71 (1995), Makita et al. "Molecular cloning and functional expression of rat leukotriene A4 hydrolase using the polymerase chain reaction" FEBS Lett., 299 (3): 273-277. (1992), Genbank Accession No.M63848 (mouse mRNA sequence and amino acid sequence), GenBank Accession Nos. J03459, J02959, and
U27293 (human mRNA sequence and amino acid sequence), and Genbank Accession No. S8752
2 (rat mRNA sequence and amino acid sequence), which are incorporated herein by reference. The antisense nucleic acid molecule is RNA or single-stranded D
A full-length mR that can be NA and encodes a leukotriene A4 hydrolase
It can also be complementary to an NA molecule (ie, of the same nucleotide length as the full length molecule). However, it may be desirable to work with shorter molecules. In this case, the antisense molecule can be complementary to a portion of the full length mRNA molecule that encodes a leukotriene A4 hydrolase. These shorter antisense molecules are capable of hybridizing to the mRNA encoding the full length molecule and preferably consist of at least 15 nucleotides and up to about 100 nucleotides. Leukotriene A4 hydrolase mRNA
Levels of leukotriene A4 hydrolase by contacting cells with RNA or single-stranded DNA molecules complementary to at least a portion of the antisense molecules (eg, by introducing antisense molecules into a sample). It can be used to reduce. The antisense molecule can base pair with the mRNA of leukotriene A4 hydrolase and inhibits translation of the mRNA into protein. Thus, an antisense molecule to leukotriene A4 hydrolase can inhibit the translation of the mRNA encoding leukotriene A4 hydrolase into a functional leukotriene A4 hydrolase, which results in leukotriene A in the cell.
The activity of 4 hydrolase can be reduced.

The antisense molecule is contacted with the cell by any suitable means. According to the method of the invention, the cells are adenocarcinoma cells such as prostate cancer cells, lung cancer cells, gastric cancer cells, breast cancer cells, colon cancer cells and pancreatic cancer cells. In one embodiment, the antisense R
The antisense RNA molecule is contacted with the cell by injecting the NA molecule directly into the cytoplasm, and in the cytoplasm the RNA interferes with translation. Vectors can also be used to introduce antisense molecules into cells. Such vectors include, for example, various plasmids and viral vectors. The antisense molecule can also be introduced into cells using liposomes. For a general discussion of antisense molecules and their use, see Han et al., Proc. Nat'l Acad. Sci. USA, 88.
: 4313-4317 (1991) ("Han"), and Rossi, British Medical Bulletin 51 (
1): 217-225 (1995). These are incorporated herein by reference.

MR, known as ribozyme, which encodes leukotriene A4 hydrolase
A special category of antisense RNA molecules with recognition sequences complementary to specific regions of NA can also be used to inhibit the activity of leukotriene A4 hydrolases. Ribozymes not only form a complex with the target sequence via the complementary antisense sequence, but also catalyze the hydrolysis or cleavage of the template mRNA molecule.

Ribozyme expression in cells can inhibit gene expression (such as expression of leukotriene A4 hydrolase). More specifically, a ribozyme having a recognition sequence complementary to a region of mRNA encoding leukotriene A4 hydrolase is
It can be used to reduce the expression of leukotriene A4 hydrolase. Select cells with a first level of leukotriene A4 hydrolase expression,
This ribozyme is then introduced into the cells. This ribozyme in the cell reduces the expression of leukotriene A4 hydrolase in the cell since the mRNA encoding leukotriene A4 hydrolase is normally cleaved and cannot be translated.

Ribozymes can be contacted with the cancer cells of the methods of the invention by any suitable means. For example, the ribozyme can be directly injected into the cytoplasm, where it interferes with translation by cleaving the mRNA. In addition, a vector can be used to introduce the ribozyme into cells. Such vectors include various plasmids and viral vectors. In this respect, the DNA encoding the ribozyme need not be "integrated" in the genome of the host cell, but for example, the DNA molecule encoding the ribozyme may be a viral vector that expresses the ribozyme. It should be noted that it could be expressed in infected host cells. The ribozyme molecule could be introduced into cells using liposomes. For a general discussion of ribozymes and their use, see
For example, Server et al., Science, 247: 1222-1225 (1990), Chrysei et al., Antisens.
e Research and Development, 1 (1): 57-63 (1991), Rossi et al., AIDS Reseacrh.
and Human Retroviruses, 8 (2): 183-189 (1992), and Christophersen et al.
See Journal of Medicinal Chemistry, 38 (12): 2023-2037 (1995). These are incorporated herein by reference.

As explained, some of these methods of transforming cells are best performed with intermediate plasmid vectors. Cohen et al., U.S. Pat. No. 4,2
37,224 describes the production of expression systems in the form of recombinant plasmids using restriction enzyme cleavage and ligation with DNA ligase. This document is incorporated herein by reference. These recombinant plasmids are then introduced into single cell cultures such as prokaryotes and eukaryotic cells grown in tissue culture by means of transformation,
Duplicate. The DNA sequence was prepared according to Sambrook et al., Molecular Cloning: A Laborator.
y Manual, 2nd Edition, Cold Spring Harbor, New York: Cold Spring Harbor
Clone into a plasmid vector using standard cloning procedures known in the art as described in Laboratory Press (1989). This text is incorporated herein by reference.

As explained above, the level of leukotriene A4 hydrolase in adenocarcinoma cells, such as prostate, lung, gastric, breast, colon, or pancreatic cancer cells, can be increased by the antisense or ribozyme constructs in the cells. Can be reduced by introducing. The antisense construct blocks translation of the mRNA encoding leukotriene A4 hydrolase into leukotriene A4 hydrolase. The ribozyme construct also cleaves the mRNA encoding leukotriene A4 hydrolase, thus blocking the expression of functional leukotriene A4 hydrolase. In
Various gene therapy techniques can be utilized to introduce antisense or ribozyme constructs into desired cells for the purpose of reducing leukotriene A4 hydrolase expression in vivo. The construct needs to be targeted to the desired cells (eg, cancer cells such as prostate, lung, stomach, pancreas, colon or breast) by known methods. This is because it is not desired to reduce the expression of leukotriene A4 hydrolase in other cells of the subject.

As explained above, the binding of leukotriene B4 to the leukotriene B4 receptor can be inhibited by inhibiting the production of leukotriene B4. Also, as pointed out above, the production of leukotriene B4 can be inhibited, for example, by inhibiting one or more steps of the leukotriene B4 synthetic pathway. Another method of inhibiting the leukotriene B4 synthetic pathway comprises inhibiting leukotriene A4 production, for example by contacting a sample of adenocarcinoma cells with a 5-lipoxygenase inhibitor. As used herein, a 5-lipoxygenase inhibitor is 5
-Means any compound which directly or indirectly inhibits the activity of lipoxygenase. Examples of such 5-lipoxygenase inhibitors include inhibitors of 5-lipoxygenase activating protein (for example, 3- (1- (4-chlorobenzyl) -3
-T-butyl-thio-5-isopropyl-2-yl) -2,2-dimethylpropanoic acid and / or nordihydroguaiaretic acid) and nucleic acid molecules that reduce the expression of 5-lipoxygenase. Further details regarding these compounds are discussed in Applicant's co-pending US patent application Ser. No. 09 / 111,343. This application is incorporated herein by reference.

The method of the present invention can also be carried out using a compound that inhibits the binding of leukotriene B4 to the leukotriene B4 receptor, but does not inhibit the production of leukotriene A4. For example, the method of the present invention comprises a compound which is not a 5-lipoxygenase inhibitor, a compound which is not an inhibitor of 5-lipoxygenase activating protein, 3- (1
-(4-chlorobenzyl) -3-t-butyl-thio-5-isopropyl-2-yl) -2,2-dimethylpropanoic acid and / or nordihydroguaiaretic acid can do.

As pointed out above, the method of the present invention comprises leukotriene B4 of leukotriene B4.
It can be carried out using a compound that inhibits binding to the 4 receptor. This can be done by reducing the expression of the leukotriene B4 receptor instead of or in addition to the various methods described above. The methods discussed above with respect to reducing leukotriene A4 hydrolase expression are equally applicable here. More specifically, the compound is a nucleic acid molecule that binds to a nucleic acid molecule that encodes a leukotriene B4 receptor, such as a ribozyme targeted to an antisense nucleic acid molecule and a nucleic acid molecule that encodes a leukotriene B4 receptor. Can be Nucleic acid and amino acid sequences of leukotriene B4 receptors that can be used to design suitable antisense nucleic acid molecules and ribozymes include, for example:
Martin et al., Leukotriene binding, signaling, and analysis of HIV co-receptor function in cells transfected with mouse and human leukotriene B4 receptors, J. Biol. Chem., 274 (13): 8597-8603. (1999), Yokomizo et al., “G-protein coupled receptor of leukotriene B4 mediating chemotaxis”, Nature, 387 (6633).
): 620-624 (1997), Toda et al., "Cloning and Characterization of the Rat Leukotriene B (4) Receptor," Biochem. Biophys. Res. Commun., 262 (3): 806-812 (1999).
Genbank accession number AF077673 (mouse gene sequence and amino acid sequence), Genbank accession numbers D89078 and D89079 (human gene sequence and amino acid sequence), and Genbank accession number AB025230 (rat gene sequence and amino acid sequence). These documents are incorporated herein by reference.

Although various compounds have been identified herein that can be used to inhibit the binding of leukotriene B4 to the leukotriene B4 receptor, the method of the present invention indicates that these compounds reduce the growth of adenocarcinoma cells. And / or induce apoptosis of adenocarcinoma cells and / or induce differentiation of adenocarcinoma cells into non-cancerous cells. The meanings of the terms "proliferation", "apoptosis", and "differentiation" are readily understood in the art. Examples of methods of assaying proliferation, apoptosis, or differentiation are provided in the Examples below and in Applicant's co-pending US patent application Ser. No. 09/11.
No. 1,343. It is incorporated herein by reference. These methods inhibit the binding of leukotriene B4 to the leukotriene B4 receptor and “reduce” proliferation, “induce” apoptosis, and / or
Alternatively, it can be used to identify other compounds that “induce” differentiation.

The above discussion shows that leukotriene B4 to leukotriene B4 receptors on cancerous epithelial cells such as adenocarcinoma cells, especially prostate cancer cells, lung cancer cells, gastric cancer cells, breast cancer cells, pancreatic cancer cells, and colon cancer cells. The concept of the present invention regarding the effect of a compound that inhibits the binding of E. coli is explained. The method of the present invention can be performed in vitro or in vivo.

For example, the methods of the present invention may be used to screen glandular compounds to screen for compounds that are potentially useful in treating adenocarcinomas, such as prostate, lung, stomach, breast, colon, and / or pancreatic cancers. To assess the efficacy of a compound in treating cancer, or the mechanism by which the compound combats adenocarcinoma (eg, by inducing apoptosis, inducing differentiation, thereby reducing proliferation, and so on). Can be used in vitro to study (whether, etc.). For example, once a compound is identified as a compound that inhibits the binding of leukotriene B4 to the leukotriene B4 receptor, one of skill in the art will appreciate the extent to which the compound induces apoptosis, differentiation, and / or differentiation of cancer cells. The method of the invention can be applied in vitro to assess the extent to which proliferation is reduced. Alternatively, one skilled in the art will use the methods of the invention to determine whether the compound acts by inducing apoptosis, inducing differentiation, reducing proliferation, or a combination of these methods. Can be applied.

The methods of the invention can also be used in vivo to treat adenocarcinomas such as prostate cancer, lung cancer, gastric cancer, pancreatic cancer, breast cancer, and colon cancer. When the method of the present invention is carried out in vivo, for example, when adenocarcinoma cells are present in a human subject,
Contacting can be performed by administering to the subject a therapeutically effective amount of the compound, eg, by directly injecting the compound into the tumor. Details regarding the steps of administering a compound according to the methods of the present invention are described below.

The invention in another of its aspects relates to a method of treating an adenocarcinoma, such as prostate cancer, lung cancer, gastric cancer, breast cancer, pancreatic cancer, colon cancer or other cancer involving epithelial cells of a subject. The method comprises administering to the subject an amount of a compound effective to inhibit the binding of leukotriene B4 to the leukotriene B4 receptor.

Suitable subjects include mammals, such as rats, mice, cats, dogs, monkeys and humans. Suitable human subjects include, for example, prostate cancer, lung cancer, gastric cancer,
Patients previously determined to be at risk of developing pancreatic cancer, colon cancer, and / or breast cancer, and patients diagnosed with prostate cancer, lung cancer, gastric cancer, pancreatic cancer, colon cancer, and / or breast cancer Is mentioned. Subject does not have oral squamous cell carcinoma,
Alternatively, it is preferably not pointed out as a candidate for the treatment of oral squamous cell carcinoma by another method. In subjects determined to be at risk of having an adenocarcinoma (and preferably not having oral squamous cell carcinoma), a compound that inhibits the binding of leukotriene B4 to the leukotriene B4 receptor is preferably by any chance. If it develops, it is administered to the subject under conditions effective to reduce the proliferation of adenocarcinoma cells and / or induce apoptosis and / or induce differentiation. Such preventative (although not absolutely 100%) treatment is based on leukotriene B4.
It may be useful in high-risk individuals, as long as the potential benefit of prevention outweighs the harmful effects of administration of a compound that inhibits the binding of L. to leukotriene B4 receptors.

Any of the compounds described above can be used in the therapeutic methods of the invention. For example, a compound that inhibits the binding of leukotriene B4 to the leukotriene B4 receptor, for example, a conventional chemical substance or peptide drug directly binding to the leukotriene B4 receptor as disclosed herein is used alone. Can be administered in or as a composition in admixture with carriers that are not incompatible. Non-compounding carriers include suitable pharmaceutical carriers or diluents. The diluent or carrier component should be chosen so that they do not diminish the therapeutic effect of the compounds used in the invention.

The compositions herein are prepared in any suitable dosage form suitable for the desired application. Examples of suitable dosage forms include oral dosage forms, parenteral dosage forms, or topical dosage forms.

Suitable dosage forms for oral use include tablets, dispersible powders, granules, capsules,
Suspensions, syrups, and elixirs are included. Inert diluents and carriers for tablets include, for example, calcium carbonate, sodium carbonate, lactose, and talc. Tablets may contain granulating or disintegrating agents such as starch, alginic acid, binders such as starch, gelatin, and acacia, and lubricating agents such as magnesium stearate, stearic acid, and talc. The tablets may or may not be coated by known techniques to delay disintegration and absorption. Inert diluents and carriers, such as calcium carbonate, calcium phosphate, and kaolin, may be used in capsules. Suspensions, syrups and elixirs contain, for example, conventional excipients such as methylcellulose, tragacanth, sodium alginate, moisturizers such as lecithin and polyoxyethylene stearate, and preservatives such as ethyl p-hydroxybenzoate. May be included.

Suitable dosage forms for parenteral administration include solutions, suspensions, dispersions, emulsions and the like. They may be manufactured in the form of sterile solid compositions which can be dissolved or suspended in sterile injectable medium immediately before use. These may include suspending agents or dispersing agents known in the art. Examples of parenteral administration include intracerebroventricular administration, intracerebral administration, intramuscular administration, intravenous administration, intraperitoneal administration, rectal administration, and subcutaneous administration.

In addition to the generally inactive components of the formulations described above, these formulations are useful for treating other active agents, especially prostate, lung, stomach, breast, colon, pancreatic cancer and / or other adenocarcinomas. Active agents identified as being useful in therapy can be included. These active agents are also useful in treating other types of cancer (ie, in addition to adenocarcinoma), or
For example, there may be a wide range of anti-cancer agents in which the other active agent is more specific in that it is useful in treating adenocarcinoma but not in treating oral squamous cell carcinoma. The other active substances may also have non-anti-cancer pharmacological properties in addition to their anti-adenocarcinogenic properties. For example, the other active substance can also have anti-inflammatory properties,
Alternatively, it may not have such anti-inflammatory properties.

It should be understood that the actual preferred amount of the compound administered according to the present invention will vary depending on the particular compound, the particular composition formulated and the mode of administration. One of ordinary skill in the art will consider many factors that may modify the action of the compound, such as weight, sex, diet, time of administration, route of administration, excretion rate, condition of the subject, drug combination, and reaction susceptibility and severity. Can be put in. Administration can be carried out continuously or periodically within the maximum tolerated dose. Optimal dosing rates for a given set of conditions can be ascertained by one of ordinary skill in the art using routine dose dosing tests.

[0083]   The invention is further described using the following examples.

[0084] In Example Example 1 materials and in the following examples the method, materials and methods described below were used.

Materials DMEM, MEM, and McCoy's 5A medium, penicillin-streptomycin solution, trypsin-EDTA solution, proteinase K, propidium iodide, RNase A, and phenol / chloroform were purchased from Sigma Chemicals (St. Louis, MO). did. Fetal bovine serum ("FBS") was obtained from Atlanta Biologicals (Norcross, GA). The APO-BRDU kit for the Tuner assay was purchased from Pharmingen (San Diego, CA). Leukotriene B4 (“LTB4”) was obtained from Cayman Chemicals (Ann Arbor, Michigan). 2- (2-propyl-3- (3- (2-ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy) -phenoxy) benzoic acid, an LTB4 antagonist (
Hereinafter "LY293111") was provided by Eli Lilly and Company (Indianapolis, IN). 1- (2-hydroxy-3-propyl-4- (4- (1H-tetrazole-5-
(Il) butoxy) phenyl) ethanone (hereinafter "LY171883") was purchased from Biomol Research Laboratories (Plymouth Meeting, PA).

Cell Culture The following pancreatic cancer cell lines were used. That is, MiaPaCa-2 and PANC-
1 (poorly differentiated), Capan-1 and Capan-2 (well differentiated), and HPAF and AsPC-1 (multifunctional). These were purchased from the American Type Culture Collection (Rockville, MD). PANC-1 and MiaPaCa-2 were grown in DMEM medium. HPAF and AsPC-1 were grown in MEM medium. And Capan
-1 and Capan-2 were grown in McCoy's medium. 1 for these media
Supplemented with 0% FBS and cells were grown as a monolayer at 37 ° C in a humidified 95% O ₂ and 5% CO ₂ atmosphere. The cells were regularly seeded in a 75 cm ² flask and the medium was replaced every other day. For experiments, cells were grown to 80% confluence, digested with trypsin-EDTA and seeded in either 12- or 24-well plates as appropriate at a concentration of 50,000 / ml.

DNA synthesis by [ ³ H] thymidine incorporation Cells were seeded in 12 or 24 well plates. After reaching 70% confluence, they are incubated in serum-free medium for 24 hours and then leukotriene B4 receptor antagonist LY293111 (62.5-10) at appropriate concentration.
00 nM), LTB4 (100-400 nM), or fresh serum-free medium with or without both. After culturing for an appropriate period, 0.5 μCi / well
Cellular DNA synthesis was assayed by adding [ ³ H] -methyl-thymidine and incubating for an additional 2 hours. The cells were then washed twice with PBS, 10
Fix with trichloroacetic acid ("TCA") for 30 minutes and add 0.4 M Na to each well.
It was solubilized by adding OH. Radioactivity, indicating incorporation of ³ H-thymidine into DNA, was measured by adding scintillation cocktail and counting in a scintillation counter (ElKby Rack Beta, Wallas, Turku, Finland).

Cell Proliferation Assay Pancreatic cancer cells were seeded in 24-well microplates and incubated at 37 ° C. After 24 hours, cells were cultured for 24, 48, and 72 hours in serum-free medium with or without 250 nM LY293111. At the end of each time, the cells were trypsinized into a single cell suspension and the number of cells in each well was determined using a Z1-Coulter counter (Luton, Bedfordshire, UK).

Morphological Changes Using Light Microscopy Pancreatic cancer cells grown in 75 cm ² flasks were treated with different concentrations of LY293111.
At different times. It was then viewed with a phase contrast microscope and photographed.

DNA Fragmentation Assay For analysis of DNA fragmentation by agarose gel electrophoresis, treated pancreatic cancer cells were detached from a 75 cm ² flask with trypsin-EDTA, washed twice with PBS, and washed with 1 ml of lysis buffer ( 25 mM EDTA, 10 mM Tris, 0.5%
Triton X-100 and 1% SDS) and freeze-thaw overnight at -80 ° C. An equal volume of PEG / NaCl stock solution was then added to the lysate and added to 0.
The tubes were centrifuged at 13,200 g for 30 minutes at 4 ° C. to a final concentration of 25% PEG and 1 M NaCl. The supernatant containing the fragmented small molecular weight DNA was then incubated with RNase A (0.3 mg / ml) for 1 hour at 37 ° C. and then with 10 ml of proteinase K (10 mg / ml) for 3 hours. The sample was then extracted with phenol-chloroform and precipitated with 70% ethanol overnight. The DNA-containing pellet was then resuspended in 50 ml TE buffer, electrophoresed on a 1.8% agarose gel containing ethidium bromide (1 mg / ml) and visualized by UV irradiation, Molecular Analyst. ·gel·
Analysis was performed using a documentation system (Bio-Rad Laboratories, Hercules, CA).

Tunel Assay Apoptosis was measured by flow cytometry using the terminal deoxynucleotidyl transferase mediated deoxyuridine triphosphate nick-endo labeling (“tunnel”) assay. LY293 cells
Treatment with 111 for an appropriate time, then digestion with trypsin-EDTA, ice-cold PBS
It was washed twice with and fixed in 10% paraformaldehyde on ice for 20 minutes. The cells were then washed with PBS and permeabilized with 70% ethanol for at least 4 hours,
Wash again with PBS, 2.5 units of terminal deoxynucleotidyl transferase (TdT enzyme) and 100 pM Br in DNA labeling solution.
-Incubated with dUTP at 37 ° C for 1 hour. The cells were then rinsed twice and resuspended in 0.1 ml of fluorescein-labeled anti-BrdU antibody solution for 30 minutes in the dark. Then 0.5 ml of Propidium iodide / RNase A
Solution was added and the cells were analyzed by flow cytometry with excitation at 488 nm.

Statistical analysis Data were analyzed by analysis of variance (“ANOVA”) with Bonferroni's corrected Dunnet for multivariate comparisons where appropriate. This analysis was performed using the Prism software package (Graph Pad, San Diego, CA). Data are expressed as mean ± SEM.

Example 2 Effect of LY293111 on Pancreatic Cancer Cells LY293111, a leukotriene B4 receptor antagonist, is Mia
Concentration dependence of thymidine incorporation and cell number (FIG. 2) in the PaCa-2 pancreatic cancer cell line (FIG. 1A) [F (5,30) = 77.30, P <0.0001] and time dependence (FIG. 1B). ) [F (3,20) = 331.1, P <0.0001] is caused. 5 other pancreatic cancer cell lines (HPAF, Capan-1, Cap
Experiments performed with an-2, AsPC-1, and PANC-1) also showed LY293.
111 was shown to inhibit proliferation after 24 hours in a concentration-dependent manner. The results of these experiments are shown in Table I. In the table, the results are percent of control (mean ± SEM)
, ^* Is P <0.05, ^** is P <0.01, and ^*** is P <0.00
It is 1.

[0094]

[Table 1]

In addition, the transient effect of LY293111 on cell proliferation was studied in five other pancreatic cancer cell lines (HPAF, Capan-1, Capan-2, AsPC-1, and PANC-1). The results, shown in Table II, show that 500 nM LY2931.
11 shows that it inhibits pancreatic cancer cell growth of each of these cell lines in a time-dependent manner. In Table II, the results are expressed as percent of control (mean ± SEM), where ^* is P <0.05, ^** is P <0.01, and ^*** is P <0.001.

[0096]

[Table 2]

LY293111 inhibits growth at a concentration of 250 nM by at least 50%,
Inhibition was over 1000% at 1000 nM. 250 nM by phase contrast microscope
MiaPaC treated in about 4 hours after treatment with LY293111 of
The a-2 pancreatic cancer cells (FIG. 3B) showed a marked morphological change as compared to the untreated control (FIG. 3A). Over time, the treated cells became rounded, exhibiting membrane vesicle formation, chromatin condensation, nuclear fragmentation, and eventually detached from the microplate. These morphological changes have been previously interpreted as reflecting apoptosis. Similar results are shown in Figure 3C (LY293111).
Treatment) and the colon cell line LoVo, as shown in FIG. 3D (control).

Example 3 Effect of LY171883 on Pancreatic Cancer Cells In contrast to the effect of the LTB4 antagonist LY293111, the specific LTD4 receptor antagonist LY171883 was identified as MiaPaCa-2.
[F (5,6) = 1.875, P = 0.316] and HPAF [F (5,6)
= 1.940, P = 0.2217] In pancreatic cancer cells, the highest concentration used (10 m
M) only affected proliferation. However, this may reflect toxic effects. The results for MiaPaCa-2 and HPAF pancreatic cancer cells are shown in Figures 4A and 4B, respectively.

Example 4 Effect of leukotriene B4 on proliferation of pancreatic cancer cells Leukotriene B4 is MiaPaCa-2 [F (3,8) = 64.02 and F.
(3,8) = 50.31, P <0.0001] and HPAF [F (3,8) = 3
The proliferation of the 0.06, P <0.0001] pancreatic cancer cell line was stimulated in both a concentration-dependent and a time-dependent manner. FIG. 5A shows the effect of different concentrations of LTB4 on thymidine incorporation in MiaPaCa-2 cells after 24 hours. Figures 5B and 5C show 24 of MiaPaCa-2 cells (Figure 5B) and HPAF cells (Figure 5C).
10 shows the effect of 100 nM LTB4 on thymidine incorporation for 48, 48 and 72 hours. At a concentration of 100 nM at 72 hours, LTB4 more than doubled thymidine incorporation compared to controls.

Example 5 Effect of LY293111 on the stimulatory effect of LTB4 on pancreatic cancer cell proliferation LTB4 is a leukotriene B4 receptor antagonist on the proliferation of MiaPaCa-2 (FIG. 6A) and HPAF (FIG. 6B) pancreatic cancer cell lines. LY29
The inhibitory effect of 3111 was abolished. After treatment with 100 nM LTB4 for 24 hours, there was a 25% increase in proliferation compared to controls. And LY29311 at 250 nM
1 reduced proliferation by 50%. LTB4 and its antagonist LY293
When treated with 111 combinations, proliferation was restored to near control levels.

Example 6 DNA Fragmentation Assay DN to study apoptosis of pancreatic cancer cells treated with LY293111.
The A fragmentation assay was used. MiaPaCa-2 cells at 250, 500, and 10
Electrophoresis of cellular DNA on a 1.8% agarose gel after treatment with 00 nM LY293111 for 48 hours showed characteristic DNA ladders, whereas control cells did not. (Fig. 7). Similar results were obtained with other pancreatic cancer cell lines tested.

Example 7 Tunel Assay LY293111-induced apoptosis of pancreatic cancer cell lines was also evaluated by the Tunel assay. MiaPaCa-2 cells at 250 nM and 500 nM
Treatment with LY293111 for 24 hours gives 12.6% and 18.6% respectively.
Until then the apoptosis was significantly increased. In contrast, the extent of apoptosis in the control cell population was very low (1.5%). The results are shown in Figures 8A-8C. In the figure, the increase in fluorescent events in the upper right part is due to the UTP labeling of fragmented DNA. Similar apoptosis was induced in the other pancreatic, colon, and breast cancer cell lines studied.

Example 8 Effect of LY293111 on proliferation of pancreatic and colon cancer cells The effect of LY293111 on proliferation of cancer cells was studied by measuring thymidine incorporation and cell number. LY293111 was found in all pancreas studied,
It caused a large concentration-dependent and time-dependent inhibition of the growth of colon and breast cancer cell lines. The results are shown in Figures 9A-9F. 9A and 9B relate to pancreatic cancer cell line MiaPaCa-2, FIG. 9C relates to pancreatic cancer cell line HPAF, and FIG. 9D relates to pancreatic cancer cell line Capan2. Figures 9A, 9C, and 9D each show the dose effect of LY293111 on these pancreatic cancer cell lines. Further, FIG. 9B shows LY29 against pancreatic cancer cell line MiaPaCa-2.
3 shows the time-dependent effect of 3111. 9E and 9F show pancreatic cancer cell line Mia.
The effect of 0.5 μM LY293111 on cell number for PaCa-2 (FIG. 9E) and colon cancer cell line LoVo (FIG. 9F) is shown as a function of time and compared to controls.

Example 9 Effect of LTB4 and LY293111 on Bcl-2 As shown in FIG. 10A, LTB4 (100 nM) showed MCF- as revealed by Western blotting with Bcl-2 specific antiserum. It was found to increase the expression of the pro-apoptotic protein Bcl-2 in 7 breast cancer cells. In contrast, as shown in FIG. 10B, LY293111 (500n
M) is a pro-apoptotic protein Bcl-2 in MCF-7 breast cancer cells
Expression was inhibited.

[0105] Effect athymic mice 及 boss LY293111 on growth of Example 10 Human pancreatic tumor cell lines AsPc-1 xenografted in athymic mice, 3 millions of cancer in serum-free medium 50μl Cells were injected subcutaneously. In this experiment, the extremely malignant and aggressive pancreatic tumor cell line AsPc-1 was used. After establishing the tumor and growing for 4 days, 0.01% LY29311
Drinking water containing 1 was given and treated. FIG. 11 shows the tumor size (in mm ³ ) as a function of treatment time. The results show that mice ingesting LY293111 from drinking water had smaller tumors than mice in the control group.

While the preferred embodiments have been described and described in detail herein, it will be apparent to those skilled in the art that various changes, additions, substitutions, etc. can be made without departing from the spirit of the invention. Therefore, they are considered to be within the scope of the invention as defined by the claims.

[Brief description of drawings]

1A and 1B are bar graphs showing concentration-dependent inhibition of the LTB4 receptor antagonist LY293111 (FIG. 1A) and time course effects (FIG. 1B) on thymidine uptake in MiaPaCa-2 pancreatic cancer cell lines. Is. Results are expressed as% of control compared to control, * = P <0.05, ** = P <0.01
, ** = P <0.001. Data represent results obtained in 4 separate experiments.

FIG. 2 is a graph showing the time course of the effect of LY293111 on the cell number of MiaPaCa-2 pancreatic cancer cells. The cells were 500 nM LY293111 24,
The cells were treated for 48 and 72 hours, and the results (solid black squares) are expressed as relative cell number per well. Control data (dashed lines, black triangles) are also shown. When compared to the control,
* = P <0.05, ** = P <0.01, *** = P <0.001. Data represent results obtained in 4 separate experiments.

3A-3D are images of a phase contrast microscope. FIG. 3B is a control MiaPaCa.
2 shows the morphological changes of MiaPaCa-2 pancreatic cancer cells induced after treatment with 250 nM LY293111 for 12 hours when compared to 2 pancreatic cancer cells (FIG. 3A). FIG. 3D shows 2 when compared to control LoVo colon cancer cells (FIG. 3C).
FIG. 4 is a phase contrast microscopy image showing morphological changes in LoVo colon cancer cells induced by treatment with 50 nM LY293111.

4A and 4B show pancreatic cancer cell lines MiaPaCa-2 (FIG. 4A) and HPA.
LTD4 antagonist LY17188 affects thymidine incorporation of F (FIG. 4B)
3 is a bar graph showing the effect of 3 different concentrations after 48 hours. Results are expressed as% of control. * = P <0.05, ** = P <0.01, ** when compared to control
* = P <0.001. Data represent results obtained in 4 separate experiments.

FIG. 5A: Different concentrations of L on thymidine incorporation in MiaPaCa-2 cells.
It is a bar graph which shows the effect of TB4 after 24 hours. 5B and 5C show MiaP.
24, 48, and 72 of aCa-2 cells (FIG. 5B) and HPAF cells (FIG. 5C).
Figure 3 is a bar graph showing the effect of 100 nM LTB4 on thymidine uptake over time. Results for thymidine incorporation are expressed as cpm and when compared to controls, * = P <
0.05, ** = P <0.01, *** = P <0.001. Data represent results obtained in 4 separate experiments.

6A and 6B show pancreatic cancer cells MiaPaCa-2 (FIG. 6A) and HPAF (FIG.
6B) 250 nM LY293 on the stimulatory effect of 100 nM LTB4.
11 is a bar graph showing the effect of 111 after 24 hours. Results are expressed as% of control. * = P <0.05, ** = P <0.01, *** = P <when compared to control
It is 0.001. Data represent results obtained in 4 separate experiments.

FIG. 7 is an image of agarose gel electrophoresis demonstrating DNA degradation reflecting LY293111-induced apoptosis in MiaPaCa-2 cells. Cells were treated with 250, 500, and 1000 nM LY293111 for 48 hours. Total cellular DNA was separated on 1.8% agarose and visualized by ethidium bromide staining. Results represent the results of 3 separate experiments.

8A-8C show 250 nM (FIG. 8B) and 500 nM (FIG. 8C) of LY29311.
8 is a dot plot showing the results of the Tuner assay of MiaPaCa-2 pancreatic cancer cells treated with 1 for 48 hours when compared to the control (FIG. 8A). The increase in fluorescent events in the upper right quadrant is due to UTP labeling of fragmented DNA, the results representing the results of 3 separate experiments.

9A-9F are bar graphs showing the effect of LY293111 on proliferation of pancreatic and colon cancer cell lines. 9A, 9C, and 9D show MiaPaC, respectively.
LY29311 on pancreatic cancer cell lines of a-2, HPAF, and Capan2
1 shows a dose effect of 1. FIG. 9B shows LY affecting pancreatic cancer cell line MiaPaCa-2.
3 shows the time-dependent effect of 293111. FIG. 9E shows pancreatic cancer cell line MiaPaCa.
2 shows the effect of 0.5 μM LY293111 on the number of cells as a function of time, and FIG. 9F shows the effect of 0.5 μM LY293111 on the number of cells as a function of time of the colon cancer cell line LoVo.

10A and 10B show MCF7 of LTB4 and LY293111, respectively.
Fig. 3 is an image of a Western blot showing the effect on Bcl-2 expression in breast cancer cells.

FIG. 11 is a graph of tumor volume as a function of treatment time in athymic mice xenografted with the human pancreatic cell line AsPc-1 using oral LY293111 (filled triangles) versus controls (filled circles). Is.

─────────────────────────────────────────────────── ─── Continuation of front page (51) Int.Cl. ⁷ Identification code FI theme code (reference) A61P 35/00 A61P 35/00 43/00 105 43/00 105 111 111 (81) Designated country EP (AT, BE, CH, CY, DE, DK, ES, FI, FR, GB, GR, IE, IT, LU, MC, NL, PT, SE, TR), OA (BF, BJ, CF, CG, CI, CM, GA, GN, GW, ML, MR, NE, SN, TD, TG), AP (GH, GM, KE, LS, MW, MZ, SD, SL, SZ, TZ, UG, ZW), EA (AM, AZ, BY, KG, KZ, MD, RU, TJ, TM), AE, AG, AL, AM, AT, AU, AZ, BA, BB, BG, BR, BY , BZ, CA, CH, CN, CO, CR, CU, CZ, DE, DK, DM, DZ, EE, ES, FI, GB, GD, GE, GH, GM, HR, HU, ID, IL, IN, IS, JP, KE, KG, KP, KR, KZ, LC, LK, LR, LS, LT, LU, LV, MA, MD, MG, MK, MN, MW, MX, MZ, NO, NZ , PL, PT, RO, RU, SD, SE, SG, SI, SK, SL, TJ, TM, TR, TT, TZ, UA, UG, US, UZ, VN, YU, ZA, ZW F terms ( Reference) 4C084 AA17 MA17 MA21 MA22 MA23 MA35 MA37 MA41 MA43 MA52 MA55 MA56 MA60 MA66 ZB212 ZB262 ZC202 4C086 AA01 AA02 BA08 BC62 MA01 MA04 NA14 ZB21 ZB26 ZC20 4C206 AA01 AA02 DA19 DA35 KA01 MA01 MA04 NA14 ZBZ

Claims (45)

[Claims] 1. A method for reducing the proliferation of adenocarcinoma cells, or for inducing apoptosis of adenocarcinoma cells, or for inducing the differentiation of adenocarcinoma cells into non-cancerous cells, the method comprising the steps of: Contacting the sample containing the compound with a compound under conditions effective to inhibit the binding of leukotriene B4 to the leukotriene B4 receptor. 2. The method according to claim 1, wherein the sample does not contain oral squamous cell carcinoma cells. 3. The method of claim 1, wherein the sample comprises prostate cancer cells, lung cancer cells, gastric cancer cells, breast cancer cells, pancreatic cancer cells, colon cancer cells, or a combination thereof. 4. The method according to claim 1, wherein the compound inhibits binding of leukotriene B4 to leukotriene B4 receptor by binding to leukotriene B4 receptor. 5. The compound is represented by the following formula: In the above formula, R ₁ is C ₁ -C ₅ alkyl, C ₂ -C ₅ alkenyl, C ₂ -C ₅ alkynyl, C ₁ -C ₄ alkoxy, (C ₁ -C ₄ alkyl) thio, Halo or R ₂ -substituted phenyl, each of R ₂ and R ₃ is independently hydrogen, halo, hydroxy, C ₁ -C ₄ alkyl, C ₁ -C ₄ alkoxy, (C ₁ -C ₄ alkyl) ) -S (O) _q - is (alkyl of C ₁ -C ₃₎ amino, X is -O - -, trifluoromethyl, or di-, - S -, - C ( = O), or -CH ₂ -And Y is -O
- or -CH ₂ -, or when taken together -X-Y- is -CH = CH- or -
C≡C—, Z is a linear or branched C _{1 to} C ₁₀ alkylidenyl, A is a single bond, —O—, —S—, —CH═CH—, or —CR _a R _b −,
R _a and R _b are each independently hydrogen, C ₁ -C ₅ alkyl, or R ₇ -substituted phenyl, or R _a and R _b together with the carbon atom to which they are attached are C is intended to form a cycloalkyl ring of ₄ -C _8, formula ## STR2 ## in R ₄ is R ₆ or R ₄ below [Chemical 3] Wherein each of R ₆ is independently —COOH, 5-tetrazolyl, —CON (R ₉ ) ₂ or —CONHSO ₂ R ₁₀ , and each R ₇ is hydrogen. , C ₁ -C ₄ alkyl, C ₂ -C ₅ alkenyl, C ₂ -C ₅ alkynyl, benzyl, methoxy, —W—R ₆ , —T—G—R ₆ , (
C ₁ -C ₄ alkyl) -T- (C ₁ -C ₄ alkylidenyl) -O-, or hydroxy, R ₈ is hydrogen or halo, and each R ₉ is independently hydrogen, phenyl, or C ₁ -C ₄ alkyl,
Or R ₉ is taken together with the nitrogen atom to which it is attached to form a morpholino group, a piperidino group, a piperazino group, or a pyrrolidino group, and R ₁₀ is a C ₁ -C ₄ alkyl or phenyl , R ₁₁ is R ₂ , —W—R ₆ , or —T—G—R ₆ , and each W is a single bond or a linear or branched divalent hydrocarbyl radical having 1 to 8 carbon atoms. And each G is a linear or branched divalent hydrocarbyl radical having 1 to 8 carbon atoms, and each T is a single bond, —CH ₂ —, —O—, —NH—, —NHCO—, -C
(= O)-, or -S (O) _q- , K is -C (= O)-, or -CH (OH)-, and each q is 0, 1, or 2 independently. , P is 0 or 1, and t is 0 or 1, provided that when X is -O- or -S-, then Y is not -O-, and A is -O-. Or -S-, R ₄ is not R ₆ , and when A is -O- or -S- and Z is a single bond, Y is not -O-, and The method according to claim 4, wherein when p is 0, W is not a single bond, and is a compound having the formula: or a pharmaceutically acceptable salt or solvate thereof. 6. The compound is 2- (2-propyl-3- (3- (2-ethyl-
4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy) phenoxy) benzoic acid, 3- (2- (3- (2-ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy) -6- (4-Carboxy-phenoxy) phenyl) propionic acid, 1- (4- (carboxy-methoxy) phenyl) -1- (1H-tetrazol-5-yl) -6- (2-ethyl-4- ( 4-
Fluorophenyl) -5-hydroxyphenoxy) hexane, 3- (4- (7-
Carboxy-9-oxo-3- (3- (2-ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) -propoxy) -9H-xanthene)) propanoic acid, 5- (3- (2- (1-Carboxy) -ethyl) -4- (3- (2-ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) -propoxy) phenyl) -4-pentynoic acid, pharmaceutical 5. The method according to claim 4, wherein the method is selected from the group consisting of those acceptable salts or solvates thereof, and combinations thereof. 7. The compound is 2- (2-propyl-3- (3- (2-ethyl-
4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy) phenoxy) benzoic acid or a pharmaceutically acceptable salt or solvate thereof.
The method described. 8. The method according to claim 1, wherein the compound inhibits the binding of leukotriene B4 to leukotriene B4 receptor by reducing the expression of leukotriene B4 receptor. 9. The method of claim 8, wherein the compound is a nucleic acid molecule that binds to a nucleic acid molecule encoding a leukotriene B4 receptor. 10. The method according to claim 1, wherein the compound inhibits binding of leukotriene B4 to leukotriene B4 receptor by binding to leukotriene B4. 11. The method according to claim 1, wherein the compound inhibits the binding of leukotriene B4 to the leukotriene B4 receptor by inhibiting the production of leukotriene B4. 12. The method according to claim 11, wherein the compound inhibits leukotriene B4 production by inhibiting conversion of leukotriene A4 to leukotriene B4. 13. The compound which inhibits conversion of leukotriene A4 to leukotriene B4 by binding to leukotriene A4.
2. The method described in 2. 14. The method according to claim 12, wherein the compound inhibits leukotriene A4 hydrolase activity, thereby inhibiting conversion of leukotriene A4 to leukotriene B4. 15. The method according to claim 14, wherein the compound inhibits the activity of leukotriene A4 hydrolase by binding to leukotriene A4 hydrolase. 16. The method according to claim 14, wherein the compound inhibits leukotriene A4 hydrolase activity by decreasing the expression of leukotriene A4 hydrolase. 17. The method of claim 16, wherein the compound is a nucleic acid molecule that binds to a nucleic acid molecule encoding a leukotriene A4 hydrolase. 18. The method according to claim 1, wherein the compound does not inhibit the production of leukotriene A4. 19. The method of claim 1, wherein the compound is not a 5-lipoxygenase inhibitor. 20. The method of claim 1, wherein the compound is not an inhibitor of 5-lipoxygenase activating protein. 21. The method of claim 1, wherein the compound is not a nucleic acid molecule that reduces the expression of 5-lipoxygenase. 22. The method of claim 1, wherein the adenocarcinoma cells are present in a human subject and said contacting comprises administering to said human subject a therapeutically effective amount of said compound. 23. A method of treating adenocarcinoma in a subject, comprising leukotriene B.
A method comprising administering to a subject an amount of a compound effective to inhibit the binding of 4 to the leukotriene B4 receptor. 24. The method of claim 23, wherein the subject is a human subject. 25. The subject is free of oral squamous cell carcinoma.
3. The method described in 3. 26. The method of claim 23, wherein the amount is an amount effective to reduce cancer cell proliferation in a subject. 27. The method of claim 23, wherein the amount is an amount effective to induce apoptosis of cancer cells in a subject. 28. The method of claim 23, wherein the amount is an amount effective to induce the differentiation of cancer cells into non-cancerous cells in a subject. 29. The method according to claim 23, wherein the compound inhibits binding of leukotriene B4 to leukotriene B4 receptor by binding to leukotriene B4 receptor. 30. The compound is represented by the following formula: In the above formula, R ₁ is C ₁ -C ₅ alkyl, C ₂ -C ₅ alkenyl, C ₂ -C ₅ alkynyl, C ₁ -C ₄ alkoxy, (C ₁ -C ₄ alkyl) thio, Halo or R ₂ -substituted phenyl, each of R ₂ and R ₃ is independently hydrogen, halo, hydroxy, C ₁ -C ₄ alkyl, C ₁ -C ₄ alkoxy, (C ₁ -C ₄ alkyl) ) -S (O) _q - is (alkyl of C ₁ -C ₃₎ amino, X is -O - -, trifluoromethyl, or di-, - S -, - C ( = O), or -CH ₂ -And Y is -O
- or -CH ₂ -, or when taken together -X-Y- is -CH = CH- or -
C≡C—, Z is a linear or branched C _{1 to} C ₁₀ alkylidenyl, A is a single bond, —O—, —S—, —CH═CH—, or —CR _a R _b −,
R _a and R _b are each independently hydrogen, C ₁ -C ₅ alkyl, or R ₇ -substituted phenyl, or R _a and R _b together with the carbon atom to which they are attached are C It is intended to form a cycloalkyl ring of ₄ -C _8, wherein R ₄ is R ₆ or R ₄ has the formula ## STR5] below [Chemical 6] Wherein each of R ₆ is independently —COOH, 5-tetrazolyl, —CON (R ₉ ) ₂ or —CONHSO ₂ R ₁₀ , and each R ₇ is hydrogen. , C ₁ -C ₄ alkyl, C ₂ -C ₅ alkenyl, C ₂ -C ₅ alkynyl, benzyl, methoxy, —W—R ₆ , —T—G—R ₆ , (
C ₁ -C ₄ alkyl) -T- (C ₁ -C ₄ alkylidenyl) -O-, or hydroxy, R ₈ is hydrogen or halo, and each R ₉ is independently hydrogen, phenyl, or C ₁ -C ₄ alkyl,
Or R ₉ is taken together with the nitrogen atom to which it is attached to form a morpholino group, a piperidino group, a piperazino group, or a pyrrolidino group, and R ₁₀ is a C ₁ -C ₄ alkyl or phenyl , R ₁₁ is R ₂ , —W—R ₆ , or —T—G—R ₆ , and each W is a single bond or a linear or branched divalent hydrocarbyl radical having 1 to 8 carbon atoms. And each G is a linear or branched divalent hydrocarbyl radical having 1 to 8 carbon atoms, and each T is a single bond, —CH ₂ —, —O—, —NH—, —NHCO—, -C
(= O)-, or -S (O) _q- , K is -C (= O)-, or -CH (OH)-, and each q is 0, 1, or 2 independently. , P is 0 or 1, and t is 0 or 1, provided that when X is -O- or -S-, then Y is not -O-, and A is -O-. Or -S-, R ₄ is not R ₆ , and when A is -O- or -S- and Z is a single bond, Y is not -O-, and 30. A compound having the formula, or a pharmaceutically acceptable salt or solvate thereof, provided that W is not a single bond when p is 0.
The method described. 31. The compound is 2- (2-propyl-3- (3- (2-ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy) phenoxy) benzoic acid, 3- (2 -(3- (2-Ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy) -6- (4-carboxy-phenoxy) phenyl) propionic acid, 1- (4- (carboxy-methoxy) Phenyl) -1- (1H-tetrazol-5-yl) -6- (2-ethyl-4- (4
-Fluorophenyl) -5-hydroxyphenoxy) hexane, 3- (4- (7
-Carboxy-9-oxo-3- (3- (2-ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) -propoxy) -9H-xanthene)) propanoic acid, 5- (3- (2 -(1-Carboxy) -ethyl) -4- (3- (2-
From ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) -propoxy) phenyl) -4-pentynoic acid, pharmaceutically acceptable salts or solvates thereof, and combinations thereof 30. The method of claim 29, which is selected from the group consisting of: 32. The compound is 2- (2-propyl-3- (3- (2-ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy) phenoxy) benzoic acid or pharmaceutically acceptable. 30. The method of claim 29, which is a possible salt or solvate thereof. 33. The method according to claim 23, wherein the compound does not inhibit the production of leukotriene A4. 34. The adenocarcinoma is prostate cancer, lung cancer, gastric cancer, breast cancer, colon cancer, pancreatic cancer,
24. The method of claim 23, which is selected from the group consisting of: and a combination thereof. 35. A method for reducing the proliferation of adenocarcinoma cells, or for inducing apoptosis of adenocarcinoma cells, or for inducing the differentiation of adenocarcinoma cells into non-cancerous cells, comprising the formula: Chemical 7] In the above formula, R ₁ is C ₁ -C ₅ alkyl, C ₂ -C ₅ alkenyl, C ₂ -C ₅ alkynyl, C ₁ -C ₄ alkoxy, (C ₁ -C ₄ alkyl) thio, Halo or R ₂ -substituted phenyl, each of R ₂ and R ₃ is independently hydrogen, halo, hydroxy, C ₁ -C ₄ alkyl, C ₁ -C ₄ alkoxy, (C ₁ -C ₄ alkyl) ) -S (O) _q - is (alkyl of C ₁ -C ₃₎ amino, X is -O - -, trifluoromethyl, or di-, - S -, - C ( = O), or -CH ₂ -And Y is -O
- or -CH ₂ -, or when taken together -X-Y- is -CH = CH- or -
C≡C—, Z is a linear or branched C _{1 to} C ₁₀ alkylidenyl, A is a single bond, —O—, —S—, —CH═CH—, or —CR _a R _b −,
R _a and R _b are each independently hydrogen, C ₁ -C ₅ alkyl, or R ₇ -substituted phenyl, or R _a and R _b together with the carbon atom to which they are attached are C is intended to form a cycloalkyl ring of ₄ -C _8, formula ## STR7 ## in R ₄ is R ₆ or R ₄ below [Chemical 9] Wherein each of R ₆ is independently —COOH, 5-tetrazolyl, —CON (R ₉ ) ₂ or —CONHSO ₂ R ₁₀ , and each R ₇ is hydrogen. , C ₁ -C ₄ alkyl, C ₂ -C ₅ alkenyl, C ₂ -C ₅ alkynyl, benzyl, methoxy, —W—R ₆ , —T—G—R ₆ , (
C ₁ -C ₄ alkyl) -T- (C ₁ -C ₄ alkylidenyl) -O-, or hydroxy, R ₈ is hydrogen or halo, and each R ₉ is independently hydrogen, phenyl, or C ₁ -C ₄ alkyl,
Or R ₉ is taken together with the nitrogen atom to which it is attached to form a morpholino group, a piperidino group, a piperazino group, or a pyrrolidino group, and R ₁₀ is a C ₁ -C ₄ alkyl or phenyl , R ₁₁ is R ₂ , —W—R ₆ , or —T—G—R ₆ , and each W is a single bond or a linear or branched divalent hydrocarbyl radical having 1 to 8 carbon atoms. And each G is a linear or branched divalent hydrocarbyl radical having 1 to 8 carbon atoms, and each T is a single bond, —CH ₂ —, —O—, —NH—, —NHCO—, -C
(= O)-, or -S (O) _q- , K is -C (= O)-, or -CH (OH)-, and each q is 0, 1, or 2 independently. , P is 0 or 1, and t is 0 or 1, provided that when X is -O- or -S-, then Y is not -O-, and A is -O-. Or -S-, R ₄ is not R ₆ , and when A is -O- or -S- and Z is a single bond, Y is not -O-, and When p is 0, W is not a single bond, and the compound having the formula: or a pharmaceutically acceptable salt or solvate thereof is contacted with a sample containing the adenocarcinoma cells. A method including steps. 36. The compound is 2- (2-propyl-3- (3- (2-ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy) phenoxy) benzoic acid, 3- (2 -(3- (2-Ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy) -6- (4-carboxy-phenoxy) phenyl) propionic acid, 1- (4- (carboxy-methoxy) Phenyl) -1- (1H-tetrazol-5-yl) -6- (2-ethyl-4- (4
-Fluorophenyl) -5-hydroxyphenoxy) hexane, 3- (4- (7
-Carboxy-9-oxo-3- (3- (2-ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) -propoxy) -9H-xanthene)) propanoic acid, 5- (3- (2 -(1-Carboxy) -ethyl) -4- (3- (2-
From ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) -propoxy) phenyl) -4-pentynoic acid, pharmaceutically acceptable salts or solvates thereof, and combinations thereof 36. The method of claim 35, which is selected from the group consisting of: 37. The compound is 2- (2-propyl-3- (3- (2-ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy) phenoxy) benzoic acid or a pharmaceutically acceptable compound. 36. The method of claim 35, which is a possible salt or solvate thereof. 38. The sample does not contain oral squamous cell carcinoma cells.
The method of claim 35. 39. The sample is a prostate cancer cell, a lung cancer cell, a gastric cancer cell, a breast cancer cell,
36. The method of claim 35, comprising pancreatic cancer cells, colon cancer cells, or a combination thereof. 40. A method of treating adenocarcinoma in a subject, the method comprising the formula: In the above formula, R ₁ is C ₁ -C ₅ alkyl, C ₂ -C ₅ alkenyl, C ₂ -C ₅ alkynyl, C ₁ -C ₄ alkoxy, (C ₁ -C ₄ alkyl) thio, Halo or R ₂ -substituted phenyl, each of R ₂ and R ₃ is independently hydrogen, halo, hydroxy, C ₁ -C ₄ alkyl, C ₁ -C ₄ alkoxy, (C ₁ -C ₄ alkyl) ) -S (O) _q - is (alkyl of C ₁ -C ₃₎ amino, X is -O - -, trifluoromethyl, or di-, - S -, - C ( = O), or -CH ₂ -And Y is -O
- or -CH ₂ -, or when taken together -X-Y- is -CH = CH- or -
C≡C—, Z is a linear or branched C _{1 to} C ₁₀ alkylidenyl, A is a single bond, —O—, —S—, —CH═CH—, or —CR _a R _b −,
R _a and R _b are each independently hydrogen, C ₁ -C ₅ alkyl, or R ₇ -substituted phenyl, or R _a and R _b together with the carbon atom to which they are attached are C Forming a _4- C ₈ cycloalkyl ring, wherein R ₄ is R ₆ or R ₄ is of the following formula: [Chemical 12] Wherein each of R ₆ is independently —COOH, 5-tetrazolyl, —CON (R ₉ ) ₂ or —CONHSO ₂ R ₁₀ , and each R ₇ is hydrogen. , C ₁ -C ₄ alkyl, C ₂ -C ₅ alkenyl, C ₂ -C ₅ alkynyl, benzyl, methoxy, —W—R ₆ , —T—G—R ₆ , (
C ₁ -C ₄ alkyl) -T- (C ₁ -C ₄ alkylidenyl) -O-, or hydroxy, R ₈ is hydrogen or halo, and each R ₉ is independently hydrogen, phenyl, or C ₁ -C ₄ alkyl,
Or R ₉ is taken together with the nitrogen atom to which it is attached to form a morpholino group, a piperidino group, a piperazino group, or a pyrrolidino group, and R ₁₀ is a C ₁ -C ₄ alkyl or phenyl , R ₁₁ is R ₂ , —W—R ₆ , or —T—G—R ₆ , and each W is a single bond or a linear or branched divalent hydrocarbyl radical having 1 to 8 carbon atoms. And each G is a linear or branched divalent hydrocarbyl radical having 1 to 8 carbon atoms, and each T is a single bond, —CH ₂ —, —O—, —NH—, —NHCO—, -C
(= O)-, or -S (O) _q- , K is -C (= O)-, or -CH (OH)-, and each of q is 0, 1, or 2 independently. , P is 0 or 1, and t is 0 or 1, provided that when X is -O- or -S-, then Y is not -O-, and A is -O-. Or -S-, R ₄ is not R ₆ , and when A is -O- or -S- and Z is a single bond, Y is not -O-, and administering to the subject a therapeutically effective amount of a compound having the formula, or a pharmaceutically acceptable salt or solvate thereof, provided that W is not a single bond when p is 0. Including the method. 41. The compound is 2- (2-propyl-3- (3- (2-ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy) phenoxy) benzoic acid, 3- (2 -(3- (2-Ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy) -6- (4-carboxy-phenoxy) phenyl) propionic acid, 1- (4- (carboxy-methoxy) Phenyl) -1- (1H-tetrazol-5-yl) -6- (2-ethyl-4- (4
-Fluorophenyl) -5-hydroxyphenoxy) hexane, 3- (4- (7
-Carboxy-9-oxo-3- (3- (2-ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) -propoxy) -9H-xanthene)) propanoic acid, 5- (3- (2 -(1-Carboxy) -ethyl) -4- (3- (2-
From ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) -propoxy) phenyl) -4-pentynoic acid, pharmaceutically acceptable salts or solvates thereof, and combinations thereof 41. The method of claim 40, which is selected from the group consisting of: 42. The compound is 2- (2-propyl-3- (3- (2-ethyl-4- (4-fluorophenyl) -5-hydroxyphenoxy) propoxy) phenoxy) benzoic acid or a pharmaceutically acceptable compound. 41. The method of claim 40, which is a possible salt or solvate thereof. 43. The method of claim 40, wherein the subject is a human subject. 44. The method of claim 40, wherein the subject does not have oral squamous cell carcinoma. 45. The adenocarcinoma is prostate cancer, lung cancer, gastric cancer, breast cancer, colon cancer, pancreatic cancer,
41. The method of claim 40, which is selected from the group consisting of: and a combination thereof.