Antitumor activity of Ge-132, a new organogermanium compound, in mice is expressed through the functions of macrophages and T lymphocytes

The antitumor activity of Ge-132 against a variety of allogeneic and syngeneic murine ascites tumors was first evaluated. The antitumor effects of Ge-132 were observed when mice inoculated with Ehrlich carcinoma (allogeneic) or RL male 1 leukemia (syngeneic) cells were treated orally. However, Ge-132 had no activity on a T-cell lymphoma (EL 4, syngeneic) or a methylcholanthrene-induced fibrosarcoma (Meth-A, syngeneic). The antitumor effect of Ge-132 in mice was related to the dose administered as well as the administration schedule. The antitumor activity of Ge-132 was next studied in mice pretreated with some blockers against immunocompetent cells. The antitumor efficacy of Ge-132 was not observed when tumor-bearing mice were treated with trypan blue and carrageenan or monoclonal anti-Thy 1.2 antibody. However, when natural killer cells were eliminated from mice bearing RL male 1 or Ehrlich ascites tumors by treatment with anti-asialo GM 1 antiserum, the antitumor efficacy of the compound was unchanged. These results suggest that Ge-132 is effective against certain ascites tumors regardless of whether the tumor is syngeneic or allogeneic. Further, its effect might be expressed through host defense mechanisms, including macrophages and/or T lymphocytes.     

Suppression of tumor growth by peritoneal macrophages isolated from mice treated with carboxyethylgermanium sesquioxide (Ge-132)

In a murine model it has been shown that the antitumor activity of carboxyethylgermanium sesquioxide (Ge-132) can be depleted by administration of macrophage (M phi) blockers. In the present study, the role that M phi play in the antitumor activity of the compound was investigated. Oral administration of Ge-132 in mice was demonstrated to be effective in activating M phi (Ge-132-cytotoxic M phi), and the cytotoxic activity of these M phi appeared in the peritoneal cavity of mice 48 hours after the oral administration of the compound. Co-cultivation of RL male-1 leukemia or Ehrlich carcinoma cells with Ge-132-cytotoxic M phi in vitro resulted in marked suppression of the growth of tumor cells. The transfer of peritoneal exudate cells (PEC), or purified M phi fractions of PEC from Ge-132-treated mice to mice bearing Ehrlich or RL male-1 ascites tumors resulted in significant protection. However, when the cytotoxic M phi were depleted by carbonyl-iron treatment in vitro, no antitumor effect was demonstrated in mice bearing Ehrlich or RL male-1 ascites tumors. Macrophage fractions obtained from PEC of Ge-132-treated mice exhibited an inhibitory effect against certain tumors both in vivo and in vitro suggesting that the antitumor effect of Ge-132 observed in vivo resulted from the activation of M phi.     

Ability of sera from mice treated with Ge-132, an organo-germanium compound, to inhibit experimental murine ascites tumors

Serum specimens from mice treated orally with Ge-132 (100 mg/kg) exhibited antitumor activity against Ehrlich (allogeneic) and RL 1 (syngeneic) ascites tumors in BALB/c mice. Sera obtained from mice 24 hours after Ge-132 administration displayed the highest antitumor effect and the antitumor activity was dose-dependent. Sera prepared from mice 12, 36 or 48 hours after Ge-132 treatment had no protective effect. Circulating interferon (IFN) was induced at 24 hours after administration. The antiviral activity of serum from Ge-132-treated mice was inactivated by treatment with trypsin, low pH, and anti-IFN-gamma antiserum. The inactivated preparations of serum IFN induced by Ge-132 did not show antitumor activity when administered to mice bearing Ehrlich ascites tumors. These results suggest that the antitumor activity in the sera of Ge-132-treated mice may have been expressed through IFN-gamma which was induced by Ge-132.     

Importance of T-cells and macrophages in the antitumor activity of carboxyethylgermanium sesquioxide (Ge-132)

The purpose of this study was to investigate the effective mechanisms of Ge-132, an organogermanium compound with immunomodulatory activity, on experimental murine ascites tumors. The antitumor effects of Ge-132 were observed when mice inoculated with Ehrlich carcinoma (allogeneic) or RL male 1 leukemia (syngeneic) cells were treated orally. However, Ge-132 had no activity on EL-4 lymphoma (syngeneic) or Meth A fibrosarcoma (syngeneic). The antitumor activity of Ge-132 was not observed when tumor-bearing mice were treated with trypan blue, carrageenan, or monoclonal anti-Thy 1.2 antibody. However, when natural killer (NK) cells were eliminated from mice bearing RL male 1 or Ehrlich ascites tumors by treatment with anti-asialo GM1 antiserum, the antitumor activity of the compound was unchanged. This suggests that Ge-132 was effective against certain ascites tumors regardless of whether the tumor was syngeneic or allogeneic. Furthermore, its effect might be expressed through host defense mechanisms, including macrophages and/or T-cells.     

Induction of interferon and activation of NK cells and macrophages in mice by oral administration of Ge-132, an organic germanium compound

After oral administration of an organic germanium compound, Ge-132 (300 mg/kg), a significant level of interferon (IFN) activity was detected in the sera of mice at 20 hr and it reached a maximum of 320 U/ml at 24 hr. This IFN activity was lost after heat- or acid-treatment, suggesting that the induced IFN is of gamma nature. The molecular weight of this IFN was estimated to be 19,000 daltons by gel filtration. After the oral administration of Ge-132, NK activity of spleen cells had increased at 24 hr and cytotoxic macrophages were induced in the peritoneal cavity at 48 hr. In mice, receiving intraperitoneal (i.p.) injections of trypan blue or carrageenan 2 days before oral administration of Ge-132, neither induction of IFN nor augmentation of NK activity followed. X ray irradiation of mice also rendered the mice incapable of producing IFN, all suggesting that both macrophages and lymphocytes are required for this IFN induction. After i.p. administration of induced IFN, both NK and cytotoxic macrophage activities appeared at 48 hr with as little as 20 U/ml titered IFN. These facts may indicate that the augmentation of NK and activation of macrophages in mice after oral administration of Ge-132 is mediated by induced IFN.     

Antitumor effect of a synthetic cord factor, 6,6'-di-O-decanoyl-alpha, alpha-trehalose (SS 554) in mice

The antitumor effect of a synthetic cord factor (6, 6'-Di-O-decanoyl-alpha, alpha-trehalose) (SS 554) on the growth of Meth-A fibrosarcoma in BALB/c mice was examined. With regard to administration routes, only intratumoral (i.t.) injection showed a curative effect; subcutaneous (s.c.), per oral (p.o.) or intravenous (i.v.) routes has no such effect. To show the antitumor effect of known natural and synthetic cord factors, the co-presence of oily vehicles has been shown to be necessary. Accordingly, compound SS 554 examined in suspensions of sesame oil, squalane (SQA), squalene (SQE) or sesame oil and water emulsion had a curative effect with a 60% survival rate. However, no such effect was obtained with a suspension in PBS or in HCO-60 solution. In this regard, it should be noted that sequential but independent administration of SS 554 and oil was found to be equally as effective as simultaneous administration of oil with SS 554. Thus the effect of the oil should be reconsidered through an examination of the sequential appearance of effector cells. In the case of sesame oil, the amount of oil necessary was over 10%, or 0.01 mg absolutely. When the dose effect of SS 554 was examined in the presence of 10% sesame oil, doses over 1 mg exhibited a dose dependent curative effect. In tumor-bearing mice, the effect of the time of administration was also examined; the best result was obtained when intratumor injection was performed on day 3 after tumor implantation. Mice that recovered after SS 554 treatment exhibited growth inhibition and rejection of rechallenged Meth-A cells. However, this immunity was specific as it did not extend to a rechallenge with RL male-1 leukemia cells.     

Ability of sera from mice treated with Ge-132, an organic germanium compound, to inhibit experimental murine ascites tumours

Sera from C57Bl/6 mice treated orally with Ge-132 exhibited antitumour activity against Ehrlich (allogeneic) and RL male 1 (syngeneic) ascites tumours in BALB/c mice. Sera obtained from mice 24 h after Ge-132 administration displayed the greatest antitumour effect and this was dose dependent. Sera prepared from mice 12, 36, or 48 h after Ge-132 treatment had no protective effect. Circulating interferon (IFN) was induced at 24 h after administration of Ge-132 but was not detected in the sera at 12, 36, or 48 h after administration. The antiviral activity of sera from Ge-132-treated mice was inactivated by treatments with trypsin, low pH, and anti-IFN gamma antiserum. The inactivated preparations of serum IFN induced by Ge-132 did not exhibit antitumour activity when administered to tumour-bearing mice. These results suggest that antitumour activity in the sera of Ge-132-treated mice may be expressed through activities of Ge-132-induced lymphokine(s), such as IFN gamma.     

双β-羧乙基锗倍半氧化物对小鼠和裸鼠移植性肿瘤生长的影响

我们研究了双β-羧乙基锗倍半氧化物(Ge—132)的抗瘤作用,结果表明对小鼠Harding—Passey黑色素瘤(M—HP)、艾氏腹水癌实体瘤(ESC)和裸鼠移植人肺鳞癌的生长有明显的抑制作用,并发现Ge—132与DDP合并应用对抗裸鼠移植人肺鳞癌有协同作用。     

Effect of combination immunochemotherapy with an organogermanium compound, Ge-132, and antitumor agents on C57BL/6 mice bearing Lewis lung carcinoma (3LL)

The antitumor effect of combination immunochemotherapy with Ge-132 and antitumor agent was studied using C57BL/6 mice bearing Lewis lung carcinoma (3LL). Ge-132 was administered orally at a daily dose of 100 mg/kg. Antitumor agents were administered intraperitoneally once a week. Initially, the effect of combination immunochemotherapy with Ge-132 and 5-fluorouracil (5-FU) was studied on 3LL local tumor growth, pulmonary metastases, survival, delayed type hypersensitivity (DTH) and body weight in tumor-bearing mice, and the following results were obtained: Inhibition of tumor growth in the combined group; Enhanced anti-metastatic effect; Prolonged survival time, and; Recovery of loss of both DTH and body weight as a result of combination therapy. These antitumor effects were also obtained by adoptive transfer of Ge-132-stimulated splenocytes in 5-FU-treated mice bearing 3LL. These results therefore suggest that the effects of Ge-132 were expressed through modification of immunocytes. Furthermore, Ge-132 enhanced the antitumor activity of bleomycin as well as that of 5-FU. These facts suggested that Ge-132 is useful for antitumor combination immunochemotherapy.     

Enhancement of cytogenetic damage and of antineoplastic effect by caffeine in Ehrlich ascites tumor cells treated with cyclophosphamide in vivo

Enhanced cytogenetic damage by cyclophosphamide (CP) was observed when Ehrlich ascites tumor cells were exposed in vivo to nontoxic concentrations of caffeine. One h before i.p. injection of 5-bromodeoxyuridine adsorbed to activated charcoal Ehrlich ascites tumor-bearing mice treated i.p. with CP appear to have a dose-dependent increase in sister chromatid exchange rates and cell division delays. Caffeine increased the survival time of the Ehrlich ascites tumor-bearing mice treated with CP and markedly reduced the ascitic volume. Therefore, the in vivo antitumor effect by CP in conjunction with caffeine appears to correlate well with the in vivo synergistic effect on cytogenetic damage caused by the combined CP plus caffeine treatment.     

Antitumor activity and its properties of Eubacterium lentum

In the present study, some basic effects of Eubacterium lentum (TYH-11), isolated from normal intestinal flora, upon the immune system and various experimental tumor cell lines were investigated. E. lentum showed no direct cytotoxicity against Ehrlich ascites tumor, while Serratia marcescens (TY-142) did show direct cytotoxicity. E. lentum presented striking antitumor activity which differed according to the injection route and time. This strain showed antitumor activity against 11 experimental tumor cell lines including Ehrlich ascites tumor, Meth-A, etc., but not against EL4 or Lewis lung carcinoma. The antitumor activity in this strain was recognized to lie in the cell wall and granular fractions. The effect of this strain on the immune system was studied by using plaque formation and the footpad reaction. When mice received 5 injections of E. lentum, the plaque number increased to treble the control level. Spleen weight was also increased following administration of E. lentum. In normal and tumor-bearing mice immunized with SRBC alone, the footpad reaction was increased significantly to the control level by administration of E. lentum.     

Antitumor mechanisms of carboxyethyl-germanium sesquioxide (Ge-132) in mice bearing Ehrlich ascites tumors

The administration of IFN-containing sera (Ge-sera) obtained from Ge-132-treated mice (Ge-mice) or the passive transfer of macrophages (M phi) to mice bearing ascites tumors resulted in the inhibition of tumor growth. The cooperative role of Ge-sera and Ge-M phi in the display of Ge-132-antitumor activity was studied. When mice were pretreated with antimouse IFN gamma antiserum, no IFN-inducing or antitumor activities of the compound were detected. Cytotoxic activities were detected in peritoneal M phi of mice treated with Ge-sera, and passive transfer of these M phi to tumor-bearing mice resulted in the inhibition of tumor growth. When tumor-bearing mice were pretreated with substances toxic to M phi, no antitumor activity of Ge-sera was observed. However, Ge-132 antitumor activity was observed in mice depleted of T-cells, even though the antitumor effects of the compound itself were not demonstrable in T-cell-depleted mice. Therefore, a part of the antitumor activity of Ge-132 appears to be expressed as follows: Ge-132 stimulates T-cells to produce circulating lymphokine(s) which are inactivated by anti-IFN gamma treatment; activated M phi are generated from resting M phi by these lymphokine(s); the transplanted tumors are inhibited by these M phi.     

Antitumor Activity and Its Properties of Eubacterium lentum

In the present study, some basic effects of Eubacterium lentum (TYH-11), isolated from normal intestinal flora, upon the immune system and various experimental tumor cell lines were investigated. E. lentum showed no direct cytotoxicity against Ehrlich ascites tumor, while Serratia marcescens (TY-142) did show direct cytotoxicity. E. lentum presented striking antitumor activity which differed according to the injection route and time. This strain showed antitumor activity against 11 experimental tumor cell lines including Ehrlich ascites tumor, Meth-A, etc., but not against EL4 or Lewis lung carcinoma. The antitumor activity in this strain was recognized to lie in the cell wall and granular fractions. The effect of this strain on the immune system was studied by using plaque formation and the footpad reaction. When mice received 5 injections of E. lentum , the plaque number increased to treble the control level. Spleen weight was also increased following administration of E. lentum . In normal and tumor-bearing mice immunized with SRBC alone, the footpad reaction was increased significantly to the control level by administration of E. lentum .     

Antitumor effect of Cepharanthin in the double grafted tumor system

The antitumor effect of Cepharanthin (CR) in a new experimental mouse model was studied. Intratumoral administration of CR strongly inhibited the growth of Meth-A solid tumors in male BALB/c mice and led to a complete regression of tumors and resistance to reinoculated tumor. Subsequently, the antimetastatic effect of CR was examined in the double grafted tumor system, in which mice first received simultaneous intradermal inoculations of Meth-A in both right (10(6) cells) and left (2 x 10(5) cells) flanks and were then injected with 0.5 mg of CR in the right tumor on days 3, 4 and 5. CR inhibited the growth of not only the right but also the left, nontreated tumor. Immunized spleen cells were taken from mice which had been cured with the intratumoral administration of CR. Adoptive transfer of CR immunized spleen cells caused the complete regression of Meth-A tumors. The effector cell activity was lost only after treatment with anti-Lyt-1 antibody. These results suggest that intratumoral administration of CR might induce Lyt-1 positive cytotoxic cells in the spleen and the left, non-treated tumor. In BALB/c nude mice, CR inhibited the growth of the right tumor but did not the left tumor. Therefore, the antitumor activity of CR on the left tumor in the double grafted tumor system is associated with a sequential immune mechanism in which T cells may play an important role. The antitumor effect of CR on the right tumor is direct cytotoxic effect. TILs (tumor infiltrating lymphocytes) obtained from left and right sides tumors treated with CR were examined by Winn assay for their antitumor activity against Meth-A sarcoma in BALB/c mice. TILs from both sides clearly inhibited the growth of admixed Meth-A cells and seems to play an important role on antitumor effect in both tumors.     

Inhibitory effect of Capsella bursa-pastoris extract on growth of Ehrlich solid tumor in mice.

The treatment of ICR mice with i.p. injections (0.14 g/kg/day) of the extract of Capsella bursa-pastoris herb (Cruciferae) caused 50 to 80% inhibition of the solid growth of Ehrlich tumor cells that had been inoculated into the s.c. tissue of the animals. The tumor lumps in the treated mice showed multifocal necroses and the infiltartion of host fibrous tissue cells. Experiments were also performed to isolate and identify the active component for the antitumor action, and an acidic substance was isolated in crystalline form from the herb extract. This acidic substance was identified as fumaric acid and was effective in inhibiting the growth of Ehrlich solid tumor at a dose of 10 mg/kg/day. The 50% lethal dose (i.p.) of this acid was 266 mg/kg.     

Cooperation of lymphokine(s) and macrophages in expression of antitumor activity of carboxyethylgermanium sesquioxide (Ge-132)

The administration of IFN containing sera (Ge-sera) obtained from Ge-132-treated mice (Ge-mice) or the passive transfer of macrophages (M phi) to mice bearing ascites tumors resulted in the inhibition of tumor growth. The cooperative role of Ge-sera and Ge-M phi in the display of Ge-132-antitumor activity was studied. When mice were pretreated with antimouse IFN gamma antiserum, no IFN-inducing and antitumor activities of the compound were detected. Cytotoxic activities were detected on peritoneal M phi of mice treated with Ge-sera, and passive transfer of these M phi to tumor-bearing mice resulted in the inhibition of tumor growth. When tumor-bearing mice were pretreated with substances toxic to M phi, there was no antitumor activity of Ge-sera observed. However, there was antitumor activity of Ge-sera in mice depleted of T-cells, even though the antitumor effects of the compound itself were not demonstrable in T-cell depleted mice. Therefore, a part of the antitumor activity of Ge-132 may appear to be expressed as follows: (1) Ge-132 stimulated T-cells to produce circulating lymphokine(s) which were inactivated by anti-IFN gamma treatment; (2) activated M phi were generated from resting M phi by such lymphokine(s); (3) the transplanted tumors were inhibited by these M phi.     

Experimental study of antitumor effect of methyl-B12

We examined the antitumor effect of vitamin B12 (methyl-B12) using C3H/He, C57BL/6 and BALB/C mice for animals and MH134 hepatoma ascites cells, Lewis lung cancer cells and Ehrlich ascites tumor cells for tumor cells. At 1.0-10 micrograms/ml, methyl-B12 enhanced PHA- and Con-A-induced lymphocyte blastoformation of C3H/He mice. The growth of MH134 tumors on the backs of C3H/He mice were suppressed by the 7-day administration of 50 or 100 micrograms/day i.p. and their survival was longer than that of untreated mice. However, methyl-B12 administration did not positively affect the survival of C3H/He mice that had been irradiated with 60Co 300 R on the day before tumor cell inoculation. The growth of Ehrlich ascites tumor cells inoculated into BALB/C mice was also reduced at 17 and 19 days after tumor inoculation by administration of methyl-B12 50 micrograms/day i.p. and the mice survived longer than the untreated mice.     

Combined therapy of mice bearing a lymphokine-activated killer-resistant tumor with recombinant interleukin 2 and an antitumor monoclonal antibody capable of inducing antibody-dependent cellular cytotoxicity

The ability of lymphokine-activated killer (LAK) cells to mediate antibody-dependent cellular cytotoxicity and its efficacy against a LAK-resistant tumor were investigated. Cells of the MH134 murine hepatoma line are scarcely lysed by LAK cells generated in vitro by incubation of C3H/HeN mouse spleen cells with human recombinant interleukin 2 (rIL 2). However, the splenic LAK cells potently lysed the LAK-resistant tumor cells in the presence of 11G2, a monoclonal antibody (MAb) of the IgG1 isotype reactive with a part of MM antigen. Peritoneal cells induced by daily i.p. injections of rIL 2 not only exhibited LAK activity but also mediated antibody-dependent cellular cytotoxicity against MH134 tumor cells in the presence of 11G2. The peritoneal cells exhibiting these cytotoxic activities were found to be nonadherent and nonphagocytic mononuclear cells possessing a similar cell surface phenotype as that of splenic LAK cells, that is Thy-1.2+ approximately -, Lyt-1.1-, Lyt-2.1-, and asialo GM1+. Treatment of spleen cells with antibodies and complement before culture with rIL 2 revealed that the phenotype of splenic LAK precursors is Thy-1.2- and asialo GM1+. The in vivo induction of peritoneal LAK cells in response to i.p. injections of rIL 2 was markedly depressed in C57BL/6 beige mice but was normally accomplished in BALB/c nude mice. Combined therapy of C3H/HeN mice bearing MH134 ascitic tumor with i.p. injection of rIL 2 and 11G2 brought about potent suppression of the tumor growth, resulting in the significant increase in the number of tumor-free mice, whereas neither rIL 2 nor the MAb could exhibit such a potent antitumor effect when used alone. Injection (i.v.) of anti-asialo GM1 antibody not only blocked the induction of peritoneal LAK cells by rIL 2 but also abrogated the development of the antitumor effect of the combined therapy. These results strongly suggest that combination of antitumor MAbs capable of inducing antibody-dependent cellular cytotoxicity with rIL 2 therapy could result in the generation of potent antitumor effects against LAK-resistant tumors and that asialo GM1-positive non-T-cell populations including cells of the natural killer cell lineage are essential, at least in part, for development of the antitumor effects of the combined therapy with rIL 2 and MAbs.     

Antitumor effect of a novel antitumor compound, NC-190, in the double grafted tumor system

The antitumor effect of NC-190, N-beta-dimethylaminoethyl-9-carboxy-5-hydroxy-10-methoxybenzo [alpha] phenazine-6-carboxamide sodium salt, with a new experimental mouse model was studied. Intratumoral administration of NC-190 strongly inhibited the growth of Meth-A solid tumors in male BALB/c mice and led to a complete tumor regression and also resistance to the reinoculated tumor. Subsequently, the anti-metastatic effect of NC-190 was examined in the double grafted tumor system, in which mice first received simultaneous intradermal inoculations of Meth-A in both right (10(6) cells) and left (2 X 10(5) cells) flanks and were then injected with 25 micrograms of NC-190 in the right tumor on days 3, 4 and 5. NC-190 inhibited the growth of not only the right but also the left, non-treated tumor. Immunized spleen cells were taken from mice which had been cured by the intratumoral administration of NC-190. Adoptive transfer of NC-190 immunized spleen cells caused a complete regression of Meth-A tumors. The effector cell activity was lost only after treatment with anti-Lyt-2 antibody. These results suggest that intratumoral administration of NC-190 might induce Lyt-2 positive cytotoxic cells in the spleen and the left, non-treated tumor. In BALB/c nude mice, NC-190 inhibited slightly the growth of the right tumor but did not that of the left tumor. Therefore, the antitumor activity of NC-190 in the double grafted tumor system was judged as associated with a sequential immune mechanism in which T cells may play an important role. TILs (tumor infiltrating lymphocytes) obtained from the left and right sides tumors treated with NC-190 were examined by Winn assay for their antitumor activity against Meth-A sarcoma in BALB/c mice. TILs obtained from neither sides inhibited the growth of admixed Meth-A cells. Different from immunopotentiators such PSK or IL-1, NC-190 enhanced neither concomitant immunity nor sinecomitant immunity.     

In vivo antitumor activity of the bitter melon (Momordica charantia)

The in vivo antitumor activity of a crude extract from the bitter melon (Momordica charantia) was determined. The extract inhibited tumor formation in CBA/H mice which had been given i.p. injections of 1.0 X 10(5) CBA/Dl tumor cells (77% of the untreated mice with tumors versus 33% of the treated mice with tumors after 6 weeks). The extract also inhibited tumor formation in DBA/2 mice which had been given i.p. injections of either 1 X 10(5) P388 tumor cells (0% of untreated mice survived after 30 days versus 40% survival of the treated mice) or 1 X 10(5) L1210 tumor cells (0% survival of untreated mice versus 100% of treated mice after 30 days). The in vivo antitumor effect required both the prior exposure of tumor cells to the extract (2 hr) in vitro and i.p., biweekly injections of the extract into the mice. The optimum dose for tumor inhibition (8 micrograms protein, biweekly, i.p.) was not toxic to mice for at least 45 days of treatment. This same treatment caused a marked enhancement of C3H mouse thymic cell response to concanavalin A in vitro. When compared to the untreated control mice, the bitter melon-injected animals exhibited a 4-fold-higher incorporation of tritiated thymidine into trichloroacetic acid-precipitable material after 48 hr of exposure to 50 micrograms of concanavalin A. Nylon wool-purified spleen cells from these same bitter melon-treated mice exhibited an enhanced mixed lymphocyte reaction when exposed to irradiated P388 stimulator cells (186% of the untreated control mice). These data indicate that in vivo enhancement of immune functions may contribute to the antitumor effects of the bitter melon extract.