United States Patent [191
`Sehgal et al.
`
`[11] Patent Number:
`[45] Date of Patent:
`
`5,066,493
`Nov. 19, 1991
`
`[75)
`
`[54] RAP AMYCIN IN TREATMENT OF TUMORS
`Inventors: Surendra N. Sehgal, Princeton, N.J.;
`Claude Vezina, Oka, Canada
`[73] Assignee: American Home Products
`Corporation, New York, N.Y.
`
`[21] Appl. No.: 682,813
`[22] Filed:
`Apr. 9, 1991
`
`Related U.S. Application Data
`[60] Continuation-in-part of Ser. No. 391,334, Aug. 9, 1989,
`abandoned, which is a division of Ser. No. 592,193,
`Mar. 22, 1984, Pat. No. 4,885,171, which is a continua(cid:173)
`tion of Ser. No. 126,276, Mar. 3, 1980, abandoned,
`which is a continuation of Ser. No. 957,626, Nov. 3,
`1978, abandoned.
`
`[51]
`
`Int. Cl.5 ................... H61K 31/66; H61K 31/505;
`H61K 31/415; H61K 35/74
`[52] U.S. CI •.................................... 424/122; 514/110;
`514/274; 514/291
`
`[58] Field of Search ....................... 514/291, 110, 274;
`424/122
`
`[56]
`
`References Cited
`U.S. PATENT DOCUMENTS
`3,929,992 12/1975 Sehgal et al ........................ 424/122
`
`OTHER PUBLICATIONS
`C. P. Eng et al., J. Antibiotics, 37, No. 10, 1231-1237
`(1984).
`C. Vezina et al., J. Antibiotics, 28, 721 (1975).
`S. N. Sehgal et al., J. Antibiotics, 28, 727 (1975).
`
`Primary Examiner-Jerome D. Goldberg
`Attorney, Agent, or Firm-Walter Patton
`
`[57]
`ABSTRACT
`Methods of using rapamycin in the treatment of cancers
`or tumors are disclosed.
`
`6 Claims, No Drawings
`
`West-Ward Exhibit 1051
`Sehgal USP '493
`Page 001
`
`

`

`1
`
`5,066,493
`
`RAPAMYCIN IN TREATMENT OF TUMORS
`
`2
`plastic agent commonly used in cancer therapy and
`with pharmaceutically acceptable carriers, the propor(cid:173)
`tion of which is determined by the solubility and chemi-
`cal nature of the compounds, chosen route of adminis-
`BACKGROUND OF THE INVENTION
`This is a continuation-in-part application of co-pend- S tration and standard biological practice. For example,
`an anticancer effective amount of the antibiotic may be
`ing application Ser. No. 07/391,334, filed Aug. 9, 1989,
`now abandoned, which in tum is a divisional applica-
`administered alone or in combination orally in solid
`tion of co-pending application Ser. No. 06/592,193,
`form containing such excipients as starch, sugar, certain
`filed on Mar. 22, 1984, now issued as U.S. Pat. No.
`types of clay and so forth. Similarly, such an amount
`4,885,171, on Dec. 5, 1989, which in tum is a continua- 10 may also be administered orally in the form of solutions
`or suspensions, or the antibiotic may be injected paren-
`tion application of co-pending application Ser. No.
`06/126,276, filed on Mar. 3, 1980, now abandoned,
`terally alone or in combination. For parenteral adminis-
`which in tum is a continuation application of co-pend-
`tration the antibiotic may be used alone or in combina-
`ing application Ser. No. 05/957,626, filed Nov. 3, 1978,
`tion in the form of a sterile solution or suspension con-
`15 taining other solutes or suspending agents, for example,
`now abandoned.
`1. Field of the Invention
`enough saline or glucose to make the solution isotonic,
`This invention relates to the use of rapamycin as an
`bile salts, acacia, gelatin, sorbitan monooleate, polysor-
`bate 80 (oleate esters of sorbitol and its anhydrides co-
`anti-cancer or anti-tumor agent.
`2. Description of the Prior Art
`polymerized with ethylene oxide) and the like.
`R~pamycin is an ~~ifungal antibiotic described by C. 20 When utilizing rapamycin alone or in combination
`Vezma et~·.' J. Anttb1ot., 28, 721 (1975), S. N. Sehgal et
`with a therapeutically effective amount of an antineo-
`al., J. Anttb1ot., 28, 727 _(1975) and S. N. Sehgal et al.,
`plastic agent commonly used in cancer therapy for the
`U.S. Pat. No. 3,929;99.2, issued Dec. 30, 1975, filed Apr.
`treatment of tumors, the total dose of active agent can
`12, 1974. Rapamycm 1s ex~racte.d from a streptomycete
`range from 0.5 to 500 mg per kg of body weight per day
`25
`(Strepto:a:iyces hygrosc<;>p1cus~ isolated fro~ an ~ter with a preferred dosage range from 10 to 250 mg per kg
`Islan~ sml ~ple and .1s i:art1cular~y e!fect1ve agamst
`of body weight per day. However, as the dosage of
`Candida ~.b1cans both m vitro and m vivo.
`rapamycin to be administered by the method of this
`ill f
`In add1t1on, a recent report by R. R. Martel et al.,
`·
`·
`· h h
`·bes h
`· l 55 48 (1977) d
`tnventmn w o course vary wit
`t e tumor or cancer
`f
`Can J Ph
`. .
`ys10 .,
`,
`escn
`t e use o rapa-
`d 1
`f h
`d
`· h h
`al
`f h
`·
`,.
`th
`t'
`f th d
`t
`f t
`30 an to erance o t e mamm
`, an wtt t e nature o t e
`mycm 1or
`e preven 10n o
`e eve opmen o wo
`h
`.
`.
`·
`·
`·
`·
`·
`1
`1
`ot er antmeo~ ~tic agents used m combmat1on, 1~ 1s
`experimental immunopathies [(experimental allergic
`preferred ~o trut1ate t.reatment of the t~mor beanng
`encephalomyelitis (EAE) and adjuvant arthritis (AA)].
`The latter report also describes the inhibitory effect of mammal ~th a low daily dose of ~apam~cm and the~ to
`~radually ~er~ t~e dosag~ until a d~1rable reduction
`rapamycin on the formation of humoral (lgE-like) anti-
`body. This report concludes that immunosuppressant 35 m tumor ~1ze 1s. achieved without causing any ~armful
`or deletenous side effect~. The schedule of ~osmg can
`activity of rapamycin appears to be related to inhibition
`of the lymphatic system.
`range from one to five trmes per day to a smgle dose
`given every two to ten days. Such dosages and schedul(cid:173)
`ing of administration must be determined on an individ(cid:173)
`ual basis, depending upon the tumor or cancer, nutri(cid:173)
`tional state of the mammal, age of the mammal, toxicity
`in each individual, and with the nature of the other
`antineoplastic agents used in combination, etc.
`Rapamycin reduces tumor size in and prolongs the
`survival time of tumorbearing mammals. More specifi(cid:173)
`cally, rapamycin is useful for controlling the following
`carcinogenic tumors in a mammal: lymphatic leukemia,
`colon, mammary, melanocarcinoma and ependymoblas(cid:173)
`toma. The effectiveness ofrapamycin in this respect can
`be demonstrated in the laboratory with rodents having
`transplanted tumors. Details of methods used to evalu(cid:173)
`ate this effect are described in various publications; for
`example, R. I. Geran et al., Cancer Chemother. Rep.,
`Part 3, 3, (No. 2) 1-103 (1972) and references therein. In
`addition, the protocols for the antitumor tests are avail(cid:173)
`able from the National Cancer Institute, Bethesda, Md.,
`U.S.A.
`Tables 1 to 6 show the effects of therapy with rapa(cid:173)
`mycin on various tumors or cancers in rodents.
`More specifically, Table 1 shows the prolongation of
`survival time of female CDF1 mice implanted with
`lymphatic leukemia P338 by administering rapamycin;
`Table 2 shows the reduction in size of colon 38 tumors
`in female BDF1 mice by administering rapamycin;
`Table 3 shows the prolongation of survival time of male
`CDF1 mice implanted with colon 26 tumors by adminis(cid:173)
`tering rapamycin; Table 4 shows the reduction in size of
`CD8F1 mammary tumors in male CD8F1 rats by admin-
`
`DETAILS OF THE INVENTION
`According to the present method, rapamycin is em(cid:173)
`ployed as the active agent. The isolation and description
`of rapamycin is given in U.S. Pat. No. 3,929,992, cited 50
`above, herein incorporated by reference.
`Rapamycin is administered to a carcinogenic tumor
`bearing mammal for the purpose of reducing the tumor
`size and prolonging the survival time of the tumor bear-
`ing mammal, either orally or parenterally.
`While rapamycin can be administered above, e.g. as a
`sole component of a filled capsule, it is preferred to
`formulate the compound in various dosage forms for
`oral or parenteral administration, e.g. tablets or sterile
`solutions. Such formulations are described in U.S. Pat. 60
`No. 3,929,992, cited above. Rapamycin may also be
`administered in combination with a therapeutically ef(cid:173)
`fective amount of an antineoplastic agent commonly
`used in cancer therapy.
`When the antifungal antibiotic of this invention is 6S
`employed as an anticancer agent in warm-blooded ani(cid:173)
`mals, e.g. rats, it may be used alone or in combination
`with a therapeutically effective amount of an antineo-
`
`SUMMARY OF THE INVENTION
`According to this invention a method is provided for 40
`treating carcinogenic tumors in a mammal which com(cid:173)
`prises administering to the mammal an antitumor effec(cid:173)
`tive amount of rapamycin. More specifically, rapamy(cid:173)
`cin reduces tumor size in and prolongs the survival time
`of tumor bearing mammals.
`
`45
`
`55
`
`West-Ward Exhibit 1051
`Sehgal USP '493
`Page 002
`
`

`

`3
`istering rapamycin; Table S shows the prolongation of
`survival time offemale BDF1 mice implanted with B16
`melonocarcinoma by administering rapamycin; and
`Table 6 shows the prolongation of survival time of male
`Swiss mice implanted with ependymoblastoma by ad- 5
`ministering rapamycin.
`TABLE 1
`Effect of Rapamycin on Survival Time of CDF1 Mice
`lml?lanted with L~l?hatic Leukemia P-338 (ascetic2
`Ave. Wt. Difference
`MST
`Sur-
`of Animals
`Dosc/Inj.
`da;i:s
`TIC%
`vivors
`c MST
`mg/kg
`(T - C, g)
`on Day S T
`14.l
`10.2
`138
`-1.9
`6/6
`400
`-2.4
`J3.J
`J0.2
`J28
`6/6
`200
`-1.6
`13.7
`10.2
`J34
`6/6
`JOO
`so
`14.3
`J0.2
`J40
`-1.9
`6/6
`-1.6
`6/6
`J3.9
`10.2
`136
`25
`-0.6
`J3.9
`136
`6/6
`10.2
`12.5
`Treatment: Nine intraperitonul injections starting on day one in a vehicle of saline
`with Twecn-80 {Trade Mark for a derivative of Z-sorbitan mono-9.oOCtadcccnoate
`poly(oxy-1.2-ethanediyl)].
`Evaluation: TIC % = Median Survival Time (MSTI in days of treated animals
`(T)/control animals (C) x 100. A T /C % of 125 or greater is considered as a
`significant prolongation of host survival. Evaluation done on day 30.
`
`5,066,493
`
`4
`TABLE 4-continued
`Effect ofRa2!!!!:r:cin on CD8F1 Mammary Tumors in CD8F1 Rats
`Average
`Net Wt. Difference
`Sur-
`Dosc/Inj.
`of Animals
`TIC%
`vivo rs
`c MTW
`on Day 5 T
`(T- C, g)
`mg/kg
`3200
`29
`928
`10/JO
`-0.8
`J2.5
`Treatment: Single intraperitoneal injection on days 1, 8. 1,, 22 and 29 in a vehicle of
`saline with Tween-80.
`10 Evaluation: TIC % = Median tumor weight (MTW) estimated from tumor diame(cid:173)
`ter of treated animals (T)lcontrol animals (C) X JOO. A TIC % of 42 or less is
`considered as a significant inhibitor of tumor growth. Evaluation done on day 30.
`
`MTW
`mg
`
`.15
`
`20
`
`TABLES
`Effect ofRa!!!!.!!:r:cin on BJ6 Melanocarcinoma in BDF1 Mice
`Average
`MST
`Sur-
`Net Wt. Difference
`da;i:s
`of Animals
`Dosc/Inj.
`TIC%
`vivors
`c MST
`(T-C,g)
`mg/kg
`on Day 5 T
`20.1
`109
`10/10
`22.0
`-3.3
`400
`20.I
`IJO
`22.3
`10/JO
`200
`-1.S
`28.0 20.l
`139
`JO/JO
`JOO
`-1.2
`25.3
`20.I
`125
`J0/10
`-0.7
`50
`28.0
`20.1
`139
`JO/JO
`O.J
`25
`29.0 20.J
`144
`10/10
`0:1
`12.5
`2S Treatment: Single intrapcritoneaJ injection on each of days I through 9 in a vehicle
`of saline with Tween-80.
`Evaluation: TIC % = Median Survival Time (MST) in days of treated animals (T)
`control animals (C) X 100. AT IC % of 12S or greater is considered as a significant
`prolongation of host survival. Evaluation done on day 60.
`
`Dosc/Inj.
`mg/kg
`
`TABLE 2
`Effect of Ral?am:r:cin on Colon 38 Tumor Weight in Mice
`Ave. Wt. Difference
`MTW
`Sur-
`of Animals
`mg
`TIC%
`vivors
`(T - c, g)
`c MTW
`on Day S T
`23
`810
`-3.4
`188
`10/10
`400
`25
`810
`-2.0
`10/JO
`209
`200
`33
`810
`-0.8
`272
`100
`10/10
`so
`810
`39
`-0.8
`320
`9/10
`45
`-0.4
`810
`10/10
`368
`25
`45
`0.4
`368
`12.5
`810
`10/10
`Treatment: Single intraperitoneal injection on days 2, 9 and 16 in a vehicle of saline
`with Tween-80.
`Evaluation: TIC % = Median tumor weight (MTW) estimated from tumor diame(cid:173)
`ter of treated animals m1control animals (C) x JOO. A TIC % of 42 or less is
`considered as a significant inhibitor of tumor growth. Evaluation done on day 20.
`
`30
`
`35
`
`40
`
`TABLE 6
`Effect of Ral?am;i:cin on El!!;nd;i:moblastoma in Swiss Mice
`Average
`MST
`Net Wt. Difference
`Sur-
`Dosc/Inj.
`da:zcs
`of Animals
`TIC%
`vivors
`on Day 5 T
`mg/kg
`MST
`(T - c. g)
`c
`243
`18.l
`44.0
`10/JO
`-3.3
`200
`143
`18.J
`26.0
`JO/JO
`-2.2
`JOO
`187
`18.J
`34.0
`9/10
`50
`-1.3
`187
`34.0 18.l
`10/10
`-2.0
`25
`178
`32.3
`18.l
`10/10
`-1.0
`12.5
`Treatment: Single intraperitoneal injection on each of days I through 9 in a vehicle
`of saline with Tween-80.
`Evaluation: TIC % = Median Survival Time (MSTI in days of treated animals m
`control animals (C) x 100. A TIC % or 12' or greater is considered as a significant
`prolongation of host survival. Evaluation done on day 60.
`
`Rapamycin also can be used to produce beneficial
`effects in the treatment of malignant tumors when com(cid:173)
`bined with a therapeutically effective amount of an
`antineoplastic agent commonly used in cancer therapy.
`so Such antineoplastic agents
`include
`the alkylating
`agents, for example, busulfan, chlorambucil, cyclophos(cid:173)
`phamide, mechlorethamine hydrochloride, melphalan,
`pipobroman, thiotepa and uracil mustard; antimetabo-
`lites, for example, cytarabine, fluorouracil, floxuridine,
`mercaptopurine, methotrexate and thioguanine; miscel(cid:173)
`laneous anticancer agents, for example, dacarbazine,
`hydroxyurea, mitotane, procarbazine hydrochloride,
`quinacrine hydrochloride, vinblastine sulfate and vin(cid:173)
`cristine sulfate; estogens, for example, chlorotrianisene,
`60 conjugate estogens (e.g. PREMARIN @), diethylstil(cid:173)
`bestrol and the like; androgens, for example, methyltes(cid:173)
`tosterone, testosterone and the like; adrenal corticoster(cid:173)
`oids, for example, prednisone and the like; progesta(cid:173)
`gens, for example, megestrol, hydroxyprogesterone
`65 caproate and the like; radioactive isotopes; and antibiot(cid:173)
`ics, for example, bleomycin sulfate, doxorubicin hydro(cid:173)
`chloride and the like. Suitable methods of administra(cid:173)
`tion, compositions and dosages of the antineoplastic
`
`45
`
`TABLE 3
`Effect of Rapamycin on Survival Time of CDF1 Mice
`Im!!lanted with Colon 26 Tumor
`Ave. Wt. Difference
`Sur-
`MST
`Dosc/Inj.
`of Animals
`da;i:s
`TIC%
`vivers
`c MST
`mg/kg
`(T-C,g)
`on Day 5 T
`-2.4
`J37
`26.3
`J9.I
`400
`J0/10
`200
`J0/10
`2S.8
`J9.J
`J3S
`-1.8
`-1.4
`29.0
`100
`J0/10
`19.1
`ISi
`so
`-0.8
`30.6
`19.1
`160
`10/10
`-0.3
`30.3
`1S8
`25
`10/10
`19.1
`0.3
`J2.S
`JO/JO
`30.4
`19.1
`1S9
`Treatment: Single intnperitoneal injection on days I, ' and 9 in a vehicle of saline
`with Tween-80.
`Evaluation: TIC % = Median survival time (MST) in days of treated animals
`mlcontrol animals (C) x 100. A TIC % or 12' or greater is considered u a 55
`significant prolonption of hoot survival. Evaluation done on day 60.
`
`TABLE4
`Effect of Ra2!!!!:r:cin on CD8F1 Mamm!D'. Tumors in CD8F1 Rats
`Average
`Net Wt. Difference
`of Animals
`(T - c. g)
`-6.6
`-6.S
`-4.8
`-4.J
`-2.4
`
`MTW
`mg
`
`Sur-
`TIC%
`vivers
`c MTW
`on Day 5 T
`4/JO
`0 3200
`JO/JO
`323
`3200
`448
`10/10
`3200
`10/10
`3200
`15S
`825
`10/JO
`3200
`
`10
`14
`23
`25
`
`Dosc/Inj.
`mg/kg
`400
`200
`100
`so
`25
`
`West-Ward Exhibit 1051
`Sehgal USP '493
`Page 003
`
`

`

`5,066,493
`
`5
`agents are described in medical textbooks; for instance,
`"PHYSICIANS' DESK REFERENCE'', 32nd ed.,
`Medical Economics Co., Oradell, N.J., U.S.A., 1978
`and "AMA DRUG EVALUATIONS", 3rd ed. PSG
`Publishing Company, Inc., Littleton, Mass., U.S.A. pp S
`1106-1151, 1977. When used in combination, rapamycin
`is administered as described previously; however, a
`lower dose can be used for efficacious results.
`We claim:
`1. A method of treating colon tumors in a mammal, 10
`which comprises administering to said mammal an anti(cid:173)
`tumor effective amount of rapamycin in combination
`with an antitumor effective amount of the antineoplastic
`agents 5-fluorouracil and cyclophosphamide.
`2. The method of claim 1 wherein rapamycin and said lS
`antineoplastic agents are administered sequentially.
`3. The method of reducing tumor size in a colon
`tumor bearing mammal, comprising administering to
`
`6
`said mammal an anti-colon tumor effective amount of
`rapamycin in combination with an antitumor effective
`amount of the antineoplastic agents 5-fluorouracil and
`cyclophosphamide.
`4. The method of prolonging the survival time of a
`colon tumor bearing mammal, which comprises admin(cid:173)
`istering to said mammal an anti-colon tumor effective
`amount of rapamycin in combination with an antitumor
`effective amount of the antineoplastic agents 5-
`fluorouracil and cyclophosphamide.
`5. The method of claim 1 wherein rapamycin is ad(cid:173)
`ministered at a dose of 0.5 to 500 mg per kg of body
`weight.
`6. The method of claim 1 wherein rapamycin is ad(cid:173)
`ministered at a dose of 10 to 250 mg per kg of body
`weight.
`• • • • •
`
`20
`
`25
`
`30
`
`35
`
`40
`
`4S
`
`SS
`
`6S
`
`West-Ward Exhibit 1051
`Sehgal USP '493
`Page 004
`
`

`

`PATENT NO.
`
`:
`
`UNITED STATES PATENT AND TRADEMARK OFFICE
`CERTIFICATE OF CORRECTION
`5,066,493
`November 19, 1991
`Surendra N. Sehgal
`Claude Vezina
`It is certified that error appears in the above-identified patent and that said Letters Patent
`is hereby corrected as shown below:
`
`DATED
`
`INVENTOR(S) :
`
`On title page, item [73]
`
`Should read:
`
`[73] Assignee: Ayerst, McKenna & Harrison, Inc.
`St. Laurent, Quebec, Canada
`
`Signed and Sealed this
`
`Ninth Day of March, 1993
`
`Attest:
`
`Attesting Officer
`
`Acting Commissioner of Patents and Trademarks
`
`STEPHEN G. KUNIN
`
`West-Ward Exhibit 1051
`Sehgal USP '493
`Page 005
`
`