JPH02188526A - Drug composition for curing of tumor - Google Patents

Abstract

PURPOSE: To prepare a pharmaceutical composition for treating tumor, comprising a tumor necrosis factor(TNF) and dipyridamole or its salt and having enhanced antitumor actions of the TNF. CONSTITUTION: This pharmaceutical composition comprises a TNF composed of TNF-α, TNF-β or a combination thereof and dipyridamole: 2,6-bis(diethanolamino)-4,8-piperidino-pyrimido[5,4-d]pyrimidine or its salt. The ingredients are substantially administered together to a patient suffering from tumor. The preferred amounts thereof in blood plasma for treating the tumor are within the following ranges: 1-5,000U/mL TNF and 0.1-200μM dipyridamole. The TNF is administered as an intamuscular or a subcutaneous injection in preferably about 5×10<6> to 5×10<8> U, more preferably 7×10<6> to 3.5×10<7> U daily dose once and dipyridamole is orally and daily administered in 25-125mg, preferably 30-60mg dose twice.

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates generally to the treatment of tumors, and in particular to tumor necrosis factor (TNF).
) regarding the treatment of tumors using More particularly, the present invention describes the use of dipyridamole to enhance the antitumor effects of TNF, methods of treating patients suffering from tumors with combination therapy of dipyridamole and TNF, and methods of such treatment for treating tumors. The present invention relates to the use of dipyridamole and TNF for the preparation of pharmaceutical compositions and to the pharmaceutical compositions so prepared.

Traditionally, TNF was first disclosed as a tumor cell degrading factor found in the serum of animals injected (smeared) with a reticular endothelial stimulant and then administered ribopolysaccharide (Carswell et al.
Proc, Natl, Acad, Sci,, U
S.A.

72:3666-3670 (1975)).

Tryponosoma blues (TryponosoIWa)
Other researchers studying the biochemical mechanisms of cachexia in rabbits injected with C. brucei observed increased concentrations of very low density lipoproteins in the plasma resulting from inhibition of riboprotein lipase activity. (Rouser and Ce
rami +Mo1. Biochem, Para
sitol+ 2: 31-38 (1980), subsequently showing that macrophages and macrophage cell lines assimilate monotons produced in response to CPS that can suppress the activity of lipoprotein lipase in adipose field cell lines. suggested. This group subsequently isolated, purified, and cloned the gene for the monopotent protein exerted by cachectin, suggesting that cachectin and TNF are the same protein. This TNF was given the name TNFα.

Lymphotoxin (designated tumor necrosis factor-β) is activated T
It is a product of cells that is involved in the defense against tumors and virus-infected cells, and was initially identified as “Ruddle and -a
ksman+J, Exp, Med, 128
:1267 (1968), and “Grange
r and -i11iams, Nat, (Lond,
) 218: 1253 (196B)”.

Human TNF-α and TNF-β have been found to have the same biological activity, but both have only 28
% amino acid sequence homology (Penn
ica, etc., Proc, Natl, Acad, Sc
i.

USA, 82:6060 (1985)).

The production of each TNF by recombinant methods was first carried out by Gray et al. and Penn1ca et al.
(Lond,) 312 Near 21 724 (1984
) and 317:724-729 (1984).

Various studies suggest that each TNF has potential antitumor activity in vivo, producing tumor regression through both direct and indirect effects.

However, its toxicity is a serious limitation. These situations, for example, 'Tumor Necros
isFactor and Re1ated C
ytotoxing, C1baFoundation
n Symposium (3L John Wtle
y & 5ons, 1987”.

In general, administration of TNF, which causes tumor necrosis and regression, approximates toxic administration.

Systemic toxicity of TNF, including fever, chills, anorexia, and nausea, is seen in most patients and is not clearly related to administration. However, other toxicities include abnormal liver R function, hypotension, -temporal neurological changes and hematological abnormalities.

The first clinical trial of TNF was initiated in Japan, where Kimte et al. reported the results of 75 patients treated with rHuTNF (Proc, Int, Cancer Con
gr.

Abstr, 1490. p, 389.1986).

In this study, I x 10' to 16 x 10' units/d was administered as an intravenous porous. Early thrombocytopenia, along with systemic side effects not clearly related to treatment, was seen in 15% of patients, leading to hepatotoxic dose limitations.

Taguchi et al., Proc, Int.

Cancer Congr, Abstr, 445
5+ p, 1.151° 1986, describes another early clinical trial with recombinant TNF. Patients received TNF as a 30-minute infusion daily or twice a week.
treated with 1 to 20 x 10' unit doses of. Apart from increased systemic toxicity, increased activity of serum transaminases was seen in 56% of treated patients, hypotension was seen in 46% of patients, and clinical shock was seen in 2 patients treated with TNF. Ta.

Creaven et al., Prod, Int, Cance
r-Congr, Adstr.

rHu TNF by 1106, p. 289.1986
In clinical trials, lXl0' ~ 4.8 x 10
Dosing of 5' units/d was reported as a 1 hour intravenous infusion, which was often accompanied by changes in blood pressure, and symptomatic hypotension was the limiting dose toxicity in this study.

Khan et al., Proc+^m, ^5soc, Canc
er Res, 27:320.1986, general systemic toxicity was the limit of administration.

It has been pointed out that on a molecular scale TNF is one of the most toxic substances used in chemotherapy.

Some studies are investigating combination treatments with TNF and other substances. TNF and endotoxin or IFN-
The combination of γ increased both antitumor efficacy and toxicity. B
loksma etc. are detoxified, endotoxin and poly(A
:U) were used; these two drugs have little or no antitumor activity on their own, but are effective in inducing tumor necrosis and regression when combined with unknown doses of TNF. Met.

A synergistic effect was observed in vitro in a cytotoxicity test with the combination of TNF and α-IFN.

The objective of the present invention is to provide a novel combination therapy capable of enhancing the antitumor activity of TNF.

The compound 2,6-bis(jetanolamine)4,8-dipiperidinopyrimide (5,4-d)pyrimidine has a single name, dipyridamole, and its preparation is described, for example, in U.S. Pat. No. 3,031 , No. 450.

Dipyridamole is a well-known vasodilator and also has anti-platelet aggregation properties; these properties have made it useful for many years in the treatment of chronic coronary insufficiency and in the prevention and treatment of cardiac infarction and in the prevention of arterial thrombosis. It has found widespread use.

More recently, the potential use of dipyridamole in tumor treatment has been investigated. The interest here is that although dipyridamole does not degrade DNA, it is an effective blocker of the salvage pathway of DNA synthesis and that dipyridamole suppresses nucleoside transport across the cell membrane and blocks the restoration of internal cellular nucleoside content, thus inhibiting tumor cell synthesis. (Btochem, Biophy
s, Acta.

158: 435-447.1968, and Bio
chem, Biophys, Acta+ 211
: 8 B-94, 1970).

Because of this property, the effects of dipyridamole alone and in combination with various anticancer agents have been investigated. That is, C
han et al. (Cancer Treat, Rep.

69:425-430.1985), dipyridamole is N-phosphoracetyl-aspartate (PAL).
It has been found that the activity of A) can be enhanced in vitro and in vivo. PALA is a pyrimidine antimetabolite that is taught to suppress the early steps of new pyrimidine synthesis, causing a depletion of intracellular pyrimidine nucleotides.

In another study, Chan et al. (Cancer Re5
45.3598-3604. August 1985) reported that dipyridamole increases PALA activity against human ovarian carcinoma cell lines but exhibits no cytotoxicity of its own.

Zhen et al., Cancer Research 43
+ 1616-1619. In April 1983, the addition of cytidine, deoxycytidine, and guanosine in combination at an optimal concentration of 8 μM each stimulated rat hepatoma 3924A cells with the antiglutamine drug adipine (Aci).
vin) and that this protection was blocked by dipyridamole at a concentration of 6 μM.

Nelson et al.
44, 24932496. In June 1984, we reported that dipyridamole enhanced the toxicity of methotrexate (MTX) to Chinese hamster ovary cells both in vitro and in vivo;
The antitumor activity of TX against certain other cancers, namely Ridgway osteogenic sarcoma and LIZIO leukemia, was not dramatically improved.

Various other publications discuss dipyridamole and anticancer drugs, such as cytarapine (Cancer Treat, Rep.
, 68:361-366.1984) or 2'-
Deoxyadenosine (Cancer Re5search
45: 64186424. December 1985) on tumor cells.

Additionally, some studies have been conducted on the use of dipyridamole alone as an anticancer drug.

Namely, in a study on the combined activity of dipyridamole/adipine, Weber (Cancer Re5e
arch73.3466-3492.August 1983)
also reported that dipyridamole was effective in killing Hepatoma 3924A cells. Ba5tida(Can
cer Research 45. 4048-405
2゜September 1985) is probably involved in the inhibition of tumor cell metabolism or the ability of tumor cells to activate platelets.
reported a potential anti-metastatic effect of dipyridamole derived from inhibition of more than one mechanism.

In the literature, dipyridamole has been used in antitumor therapy.
There is nothing that teaches or suggests its use in combination with NF, or that any valuable results can be expected from such combination therapy.

Generally, the invention includes dipyridamole or a salt thereof for use in the preparation of a pharmaceutical composition to enhance the antitumor activity of TNF.

Another aspect of the invention comprises administering dipyridamole or a salt thereof to a patient suffering from a tumor in an amount sufficient to enhance the antitumor activity of TNF also administered to the same patient. Including treatment.

Another aspect of the invention includes administering TNF and dipyridamole or a pharmaceutically acceptable salt thereof to a tumor patient, wherein the dipyridamole or salt thereof is administered in an amount sufficient to enhance the antitumor effects of TNF. The method includes a method for treating a patient with a tumor characterized by the following.

Another aspect of the invention includes dipyridamole or a pharmaceutically acceptable salt thereof for the manufacture of a pharmaceutical composition suitable for enhancing the antitumor effects of TNF when administered to a tumor patient.

Yet another aspect of the invention includes TNF for use in the manufacture of a pharmaceutical composition for use with dipyridamole or a pharmaceutically acceptable salt thereof in the treatment of tumors.

Another aspect of the invention includes a pharmaceutical composition containing TNF and dipyridamole or a pharmaceutically acceptable salt thereof together with conventional pharmaceutical excipients.

The term TNF as used herein refers to TNFα, TNF
-β or a combination thereof.

The amino acid sequences of each TNF, their recombinant preparation and the preparation of pharmaceutical compositions containing these TNFs are described in the literature, as mentioned above.

Isolation of TNF-β substantially free of impurities is described in European Patent Publication No. 0.100.641, and its preparation by recombinant techniques is described in European Patent Publication No. 164,
No. 965. Recombinant preparation of TNF-α is described in European Patent Nos. 155.549 and 168.214.

Dipyridamole or its salt and TNF are preferably administered such that both are present at the tumor site at the same time. Therefore, both are preferably administered simultaneously or substantially simultaneously.

Currently, the preferred effective plasma dipyridamole amount for antitumor therapy is about 0.1-200 μM, and the effective amount of TNF for treatment is about 1-5000 U/M.
- is considered to be

Recommended doses of dipyridamole or its salts and each TNF are based on the plasma levels described above. It will be appreciated that the actual dosage may be varied by the attending physician depending on the circumstances and conditions of the individual patient.

For use in the practice of this invention, TNF can be administered by parenteral route. The dosage and rate of administration is preferably from about 5x106 to once a day as an intramuscular or subcutaneous injection.
5X10' more preferably 7X106 to 3.5X10'
It is U.

The preparation of suitable dosage forms of human TNF is well known.

To prepare a dosage form convenient for parenteral administration, an appropriate amount of TNF is added to a 5% solution containing an appropriate buffer as required.
Soluble in human serum albumin.

The resulting solution is then passed through a microbiological filter and the filtered solution is aliquoted under sterile conditions into vials, each vial containing the appropriate amount of TNF and, if necessary, lyophilized. Each glass vial is kept cold (-20°C) before use.
It is preferable to store it in TNF can be formulated in a known manner to obtain pharmaceutically usable compositions, in which the TNF is mixed with pharmaceutically acceptable carrier materials: common carriers and their preparation are described in Reming by E. H. Martin.
tonsPharmaceutical 5cienc
It is described in es.

Dipyridamole or a salt thereof can be administered by any conventional route of administration, eg, orally or parenterally. At present, the preferred route of administration is oral. The recommended dosage is 25-125 cm twice a day, preferably 30-60
■It is.

Dipyridamole comes in many forms, for example as an injection containing 10 μl of dipyridamole under the trademark “Persanthin” (Pe
rsantin)'' and each Drazier (#M coated tablet) containing 25■ and 75■ dipyridamole is manufactured in West Germany, Frankfurt A, M,
D-6000 bundesverband derP
Harmazeutishen Industry
Rote Li5te published by e, V,
1987. Furthermore, a number of special galesus forms have been described in the literature, which aim to give accelerated or delayed (sustained) release of dipyridamole, for example European Patent Publication No. 0.032.56
There is a delayed capsule form as described in European Patent No. 2 and an immediate release form as described in European Patent No. 0.068.191. In addition, the delayed-release Galleth form is available in British Patent No. 2,025.
, No. 227.

The present invention is based on the observation that dipyridamole has shown surprising properties in promoting the antitumor effects of human TNF in a variety of established human cell lines. Briefly, several tests were performed as described below: Mouse LM cells (ATCC) used in each test.

CCLl,2) was kindly provided by Dainipong Pharmaceutical Co., Osaka, Japan and BENDER+COGes mb)l, Bienne, Austria.

Three established human cell lines were also used. These are MM I B C (Jpn) derived from the composition of a malignant melanoma (level IVpT4 NOMO) of a 70 year old man.

J. Dermatol, 96:947. 198
6), HEC-1 (intrauterine carcinoma cell line) and Y (squamous cell carcinoma cell line). Each cell was grown in Eagle's minimum essential medium containing 10% calf serum and antibiotics (100 μg streptomycin/- and 100 units penicillin G/Wd) for 37 days in a humidified atmosphere containing 5% CO□.
Cultured at ℃. Each cell was used in each test after subculture (
Approximately 1X10' cells/100 m*plastic dish).

Recombinant TNF-β formulation (2,9X10'μ/■) and recombinant TNF-α formulation (6X10'μ/■) were manufactured by Dainirapon Pharmaceutical Co. and BENDER+COGe, respectively.
Provided by smbH.

TNI? The activity was determined by measuring the erysipelas activity against LM cells.
It was confirmed that it corresponds to F-α. Dipyridamole was obtained from Karl Thomae GmbH, Biberach and der R15s, West Germany.

The effects of TNF and/or dipyridamole on the proliferation of the cells tested were described in 5uzuki et al., J. gen.

ν1vo1.67: Evaluated by cell proliferation test under continuous exposure conditions according to the procedure described in 651-661.1986. The result of the inhibitory effect on cell proliferation is
Expressed as a percentage of each subject (control) between each cell line as described in the above literature. As a general rule, all tests were performed in dark light or under yellow lamps.

FIG. 1A is a series of graphs showing percent cell survival after 11 consecutive days of exposure of LM cells to various concentrations of TNF-β and dipyridamole.

FIG. 1B is a series of photographs of culture plates after 11 days of incubation with the concentrations of each active substance indicated in the figure.

It is well known that mouse LM cells are highly sensitive to the antiproliferative effect of TNFβ, as shown in FIG. 1st
Figures A and B show that LM cells are also unexpectedly sensitive to dipyridamole. However, L
- Cell proliferation of M cells was inhibited more by the combination treatment of the two drugs than by each drug alone. This synergistic effect was also found in human tumor cell lines, as described below.

Figure 2A shows TNF-β, dipyridamole and TNF-.
2 is a series of graphs showing the cell survival percentage of Y cells after continuous exposure for 14 days and 11 days to various concentrations of dipyridamole and dipyridamole, respectively. FIG. 2B is a series of photographs of culture plates after 11 days of incubation at the concentrations of each active substance indicated in the figure. As shown in Figures 2A and B, 2A and 2B proliferation of Y cells was inhibited by each drug alone (1-100μ/■ TNF-β and 0.1μM-10μM dipyridamole).
However, treatment with a combination of the two drugs inhibited it more markedly. Synergistic effects on Y cells were observed regardless of the type of TNF formulation. The synergistic effect is
For example, it is evident in Y cells treated with 50μ/- or more of TNF-α and 0.1 μM or more of dipyridamole.

FIG. 3A is a series of graphs showing percent cell survival of MM-I CB cells after 8 consecutive days of exposure to various concentrations of TNF-α and dipyridamole.

FIG. 3B is a series of photographs of culture plates after 8 days of incubation with the concentrations of each active substance indicated in the figure. As shown in Figure 3, MM-I CB cells are also sensitive to the inhibitory effects of dipyridamole and TNF-α alone, but more sensitive to the combination.

FIG. 4A is a series of graphs showing percent HEC-1 cell survival after 11 days of continuous exposure to various concentrations of TNF-β and dipyridamole. FIG. 4B is a series of photographs of culture plates after 11 days of incubation with the concentrations of active substance indicated in the figure.

It can be seen from FIG. 4 that HEC-1 cells are sensitive to TNF-β and dipyridamole, but more markedly sensitive to the combination.

The dose required for a 50% suppressive effect on TNF sensitivity (rD,
. ), the maximum sensitivity is below 3μ/- ID,. was observed in LM cells with .

Among the three human cell lines tested, MM-ICB had the lowest ID. (50μ/-). Despite the different degrees of sensitivity among the various cell lines, combination treatment with dipyridamole was able to rapidly inhibit the proliferation of all cell lines tested.

Each human cell line used in the above studies was divided into three groups based on its sensitivity to Hu IFH. Hu
Despite the level of IFN susceptibility, the combination of TNF and dipyridamole had a significant inhibitory effect on cell proliferation.

Kaya-ichi' - Table 1 Human cell lines used and their characteristics Sensitivity to anti-proliferative strains of flu IFN
Squamous cell carcinoma
Malignant melanoma Susceptible DEC-1 Intrauterine tumor Resistant After further testing, TNF-
The enhancement of the antiproliferative effect of alpha by dipyridamole on other tumor cell lines, namely melasma melanoma cell lines, HMV I
(Gann, 73:952.1982), and the glioblast cell line, T-98 (Jpn, Cancer C
hemoLher,, 9; 1400, 198
2) Additional confirmation was made inside.

Both cell lines were sensitive to TNF-α alone, but TNF-α and dipyridamole (1 μM to 10 μM)
Proliferation inhibition of both cell lines was substantially enhanced when subjected to combination treatment with Dipyridamole; however, dipyridamole itself had only a slight inhibitory effect even at 10 IM.

Normal fibroblast cell line (J, Biochem, +10
4:570.1988) were also sensitive to each of TNF-α and dipyridamole, but surprisingly, they were not sensitive to the stimulatory effect of TNF-α on dipyridamole.

The results are shown in Figure 5. Figure 5 shows melanoma HMV I
Percent cell survival after 8 days of continuous exposure to various concentrations of TNF-α and dipyridamole in cells (A), original cells (B) and fibroblasts (C) is shown.

Therefore, it appears that dipyridamole-induced enhancement of TNF activity is enhanced only against tumor cell lines and not against normal fibroblasts.

The effect of dipyridamole in promoting the antiproliferative effect of TNF-α in intact animals was tested as follows: Nude mice (BALB/cA, JcL-nu.

MM-1cB (5X10' cells/mouse) at 4 weeks of age)
injection followed by TNF-α (2X105μ/mouse)
was administered S, C, and dipyridamole (0,1 /
mice) were administered orally every second day starting 3 days after cell implantation for a total of 62 days.

Black tumors appeared at the injection site in all treated nude mice; only slight necrosis in the tumor was found in the TNF-α-treated mice, and no changes in the tumors in dipyridamole-treated mice, but rapid onset of necrosis and ulceration. A significant reduction was detected in the combination-treated mice (Figure 6).Example 1 1.07X10' ltJ Ampoule component containing TNF-α: TNF-ot 1,07X10'I
II (0,218mg) (1) HS A
10.00■(2) Na1lzPO4X 2Hz
0 0.78 Nf3) NaJPO4X12+1
20 1.79 #(4) NaC1,1,17a
ir (5) Injection water 1.0 Ingredients 2.3
and 4 were mixed intimately and dissolved in 1/3 of the final volume of water.

The resulting solution contains a desired amount of TNF-α and TNF-α.
One volume of TNF solution taken from the alpha bulk solution was added. Water was then added to the resulting solution to the desired final volume as above.

The solution was subjected to sterile filtration and aliquots were filled into ampoules and lyophilized.

Example 2 Example 1 was repeated to obtain 5.35 x 106 I U (0,
An ampoule containing 108■) of TNF-α was obtained.

Example 3 Ampoule components with 4 0.96X10' U TNF-β: (1) TNFβ 0.96X10' U (0,5
5 ■) (21H3A 33 mg (3) Injection water 1.1 d (0.9% Na
C12) TNF-β bulk material (NaCj!TR in solution
IS) was diluted to a predetermined concentration with 20% HSA solution and water, and the 1yd portion thereof was packed into ampoules and lyophilized. The remainder from the bulk material is 16.7mM NaCj
! and 3.3mM TRl5.

Example 4 Composition of dragee containing 30■ dipyridamole: (1) Dipyridamole 30■ (2)
Lactic acid 30 mg (3) Potato starch 17.5 Akebono (4 N Aerosil)
1.5■(5) Magnesium stearate 1. A wet mass was prepared by mixing 0 mgl, 2 and 3 and adding water to the resulting mixture.

The wet mass was passed through a sieve with a 1.6 m opening and dried at 45°C in a drying room. The dry particles were mixed with 4 and 5 through a sieve with 1 + u clearance.

The final mixture was pressed to prepare a dragee.

Core weight N: 80 ■ The prepared dragee core was coated with a surface coating in a known manner; the coating had a woody appearance from sugar and talc. The coating may also contain approved colorants. The final Drazier was polished with wax.

Drazier weight 120■ Example 5 10■ dipyridamole 4 ampoule dipyridamole 10■ tartaric acid 4■ polyethylene glycol)
100 I! 1g Hydrochloric acid (IN) adjusted to pH 2.7
q, a, (0,018m) injection water
2 ml dipyridamole and each other component were dissolved in water.

After sterile filtration, the solution was filled into ampoules and sterilized by heating.

Example 6 Ampules from Examples 1-3 were formulated for injection using water for injection or isotonic saline to obtain an injection solution of 2-.

[Brief explanation of the drawing]

FIG. 1A is a series of graphs showing percent cell survival after 11 consecutive days of exposure of LM cells to various concentrations of TNF-D and dipyridamole. FIG. 1B is a series of photographs of the morphology of the organisms (culture plate showing survival of LM cells) after 11 days of culture with the concentrations of each active substance indicated in the figure. Figure 2A shows TNF-D, dipyridamole and TNF-α;
2 is a series of graphs showing the cell survival percentage of Y cells after continuous exposure for 14 days and 11 days to various concentrations of dipyridamole and dipyridamole, respectively. FIG. 2B is a series of photographs of the morphology of the organisms (culture plate showing survival of Y cells) after 11 days of culture at the concentrations of each active substance indicated in the figure. FIG. 3A is a series of graphs showing percent cell survival of MM-1cB cells after 8 consecutive days of exposure to various concentrations of TNF-α and dipyridamole. Figure 'J3B is a series of photographs of the morphology of the organisms (culture plate showing survival of MM-1cB cells) after 8 days of culture with the concentrations of each active substance indicated in the figure. Figure 4A is a series of graphs showing percent cell survival of HEC-1 cells after 11 days of continuous exposure to various concentrations of TNF-D and dipyridamole. FIG. 4B is a series of photographs of the morphology of the organisms (culture plate showing survival of HEC-1 cells) after 11 days of culture with the concentrations of active substances indicated in the figure. Figure 5 shows melanoma HMV I cells (A), original cells (
Percent cell survival after 8 days of continuous exposure to various concentrations of TNF-α and dipyridamole in B) and fibroblasts (C). TNF β (U/ml Flg, TB-1 Flg, IB-3 5U/ml TNF-β Fig, 18-2 + opM dipyridamole Fig, 1B-4 5LJ/ml TNF β Ratio of 10 μM dipyridamole to IK4 (壬) RoloFlo, 28-1 Fl (1, 2B-3 500 U/m [TNFa vs. control) Flg, 2B-2 FIG, 2B-4 500U/mt TNF-a and +PM dipyridamole T
NF-α (U/To) Flg, 3B-1 F19.38-3 500 U/ml TNF a Nobiligmol (μM) Fig, 3B-2 1 μM dipyridamole F19.3B-4 500 U/ml TNF-a and IμM dipyridamole'
I" NF-β (U/1rL1.) i g・A- Flq, 4B-1 FIG, 4B-3 10 Ll/ml TNF-β dipyridamole (μM) i g. Fl(], 4B-2 1 μM dipyridamole Flg , 4B-4 10U/ml TNF-β and IPM dipyridamole 1,
Indication of the case 29 Title of the invention 3. Person making the amendment Relationship with the case 4. Substitution (method) % formula % (1) As shown in the appendix of the engraving of the specification and drawings originally attached to the application (2) In accordance with the directive Based on this, we have corrected the brief explanation of drawings in the specification as follows. (a) Lines 2 and 3 of page 29 of the specification were corrected as follows. ``These are a series of photographs of the morphology of organisms (culture plate showing survival of LM cells) after interculturing.'' iii) Lines 9 and 10 on page 29 of the same were corrected as follows. ``This is a series of photographs of the morphology of the organism (culture plate showing the survival of Y cells) after 11 days of culture.'' (C) Lines 15 and 16 of page 29 were corrected as follows. "These are a series of photographs of the shape of the organism FW (culture plate showing survival of MM-1cB cells) after interculturing." (d) Lines 1 and 2 of page 30 of the same page were corrected as follows (
e) I did. ``This is a series of photographs of the morphology of the organism (culture plate showing survival of HEC-1 cells) after interculturing.'' Page 30, line 7 and following sentences have been deleted. (Figure 6 has been deleted because it is impossible to draw it on the drawing and will be submitted separately as a reference photo in the property submission form.)

Claims (1)

Claims: 1. Dipyridamole or a pharmaceutically acceptable salt thereof for the preparation of a medical composition suitable for enhancing the antitumor action of tumor necrosis factor. 2. Tumor necrosis factor for the preparation of a pharmaceutical composition for use with dipyridamole or a pharmaceutically acceptable salt thereof in the treatment of tumors. 3. A tumor patient comprising administering tumor necrosis factor and dipyridamole or a pharmaceutically acceptable salt thereof to the tumor patient, wherein dipyridazole is administered in an amount that enhances the antitumor effect of tumor necrosis factor. treatment methods. 4. A method for treating a tumor patient, which comprises administering dipyridamole or a pharmaceutically acceptable salt thereof to the patient in an amount that enhances the antitumor effect of tumor necrosis factor administered to the patient. 5. Dipyridamole or a pharmaceutically acceptable salt thereof, which is used for the treatment of a patient suffering from a tumor and who has been administered tumor necrosis factor, and which is administered so that dipyridamole or its salt enhances the antitumor effect of tumor necrosis factor. 6. A pharmaceutical composition comprising tumor necrosis factor and dipyridamole or a pharmaceutically acceptable salt thereof.