Complete Inhibition Of Tumor Growth Research Articles

Novel Potent Orally Active Selective VEGFR-2 Tyrosine Kinase Inhibitors: Synthesis, Structure−Activity Relationships, and Antitumor Activities of N-Phenyl-N‘-{4-(4-quinolyloxy)phenyl}ureas

N-Phenyl-N'-{4-(4-quinolyloxy)phenyl}ureas were found to be a novel class of potent inhibitors for the vascular endothelial growth factor receptor 2 (VEGFR-2) tyrosine kinase through synthetic modifications of a lead compound and structure-activity relationship studies. A representative compound 6ab, termed Ki8751, inhibited VEGFR-2 phosphorylation at an IC(50) value of 0.90 nM, and also inhibited the PDGFR family members such as PDGFRalpha and c-Kit at 67 nM and 40 nM, respectively. However, 6ab did not have any inhibitory activity against other kinases such as EGFR, HGFR, InsulinR and others even at 10000 nM. 6ab suppressed the growth of the VEGF-stimulated human umbilical vein endothelial cell (HUVEC) on a nanomolar level. 6ab showed significant antitumor activity against five human tumor xenografts such as GL07 (glioma), St-4 (stomach carcinoma), LC6 (lung carcinoma), DLD-1 (colon carcinoma) and A375 (melanoma) in nude mice and also showed complete tumor growth inhibition with the LC-6 xenograft in nude rats following oral administration once a day for 14 days at 5 mg/kg without any body weight loss.

A recombinant humanized anti-insulin-like growth factor receptor type I antibody (h7C10) enhances the antitumor activity of vinorelbine and anti-epidermal growth factor receptor therapy against human cancer xenografts.

Interaction of insulin-like growth factor receptor I (IGF-IR) with its ligands has been reported to induce cell proliferation, transformation and blockade of cell apoptotic functions. IGF-IR is overexpressed on numerous tumor cell types and its blockade could be of importance for anti-cancer therapy. We have generated a humanized anti-IGF-IR antibody h7C10 that blocks in vitro IGF-I and IGF-II-induced cell proliferation of MCF-7 breast cancer cells. Analysis of the IGF-I transduction cascade demonstrated that the humanized anti-IGF-IR antibody and its murine parental form block IGF-I-induced tyrosine phosphorylation, both its beta-chain and IRS-1 tyrosine phosphorylation. This presumably leads to cell cycle arrest and, consequently, growth inhibition. Treatment of nude mice bearing either human breast cancer cells (MCF-7) or non small lung cancer cells (A549) with h7C10, or its murine parental form 7C10, inhibited significantly tumor growth. An almost complete inhibition of A549 tumor growth was observed when mice were treated with the anti-IGF-IR antibody combined with either a chemotherapeutic agent, Vinorelbine or an anti-epidermal growth factor receptor (EGFR) antibody, 225. Combined therapy prolonged significantly the life span of mice in an orthotopic in vivo model of A549; the combination of the anti-IGF-IR antibody with an anti-EGFR antibody was superior to the Vinorelbine combination. The present results indicate that the humanized anti-IGF-IR antibody h7C10 has a great potential for cancer therapy when combined with either a chemotherapeutic agent or an antibody that targets other growth factor receptors, such as the epidermal growth factor receptor.

Antiangiogenic versus cytotoxic therapeutic approaches to human pancreas cancer: an experimental study with a vascular endothelial growth factor receptor-2 tyrosine kinase inhibitor and gemcitabine

Pancreatic adenocarcinoma is a leading cause of cancer death in the United States and represents a challenging chemotherapeutic problem. The pharmacological control of angiogenesis might represent a novel approach to the management of pancreas cancer, since the pathological development of vascular supply is a critical step for tumor growth and may affect its prognosis. In order to test this hypothesis, SU5416 ([3-(3,5-dimethyl-1 H-pyrrol-2-ylmethylene)-1,3-dihydro-indol-2-one]) a selective inhibitor of the vascular endothelial growth factor receptor-2 tyrosine kinase, and gemcitabine (2′, 2′-difluorodeoxycytidine) were tested on endothelial (HUVEC) and pancreatic tumor cells (MIA PaCa-2) in vitro and in vivo alone and in simultaneous association. SU5416 inhibited HUVEC cells stimulated to proliferate by vascular endothelial growth factor but not MIA PaCa-2 cells; the drug concentration that decreased cell growth by 50% (IC 50) was 0.14 μM. Furthermore, SU5416 reduced the development of microvessels from placental explants (IC 50, 0.23 μM). Gemcitabine inhibited the growth of both HUVEC and MIA PaCa-2 cells with an IC 50 of 0.08 and 0.1 μM, respectively. A synergistic effect (combination index <1 and dose reduction index >1) on anti-proliferative and pro-apoptotic activity was calculated with the simultaneous combination of the two drugs on endothelial cells. A marked in vivo antitumor effect on MIA PaCa-2 xenografts was observed with SU5416 at a protracted schedules, as well as with gemcitabine; furthermore, the combination between the two drugs resulted in an almost complete suppression of tumor growth and relapse. In conclusion, the present results provide the evidence of an effective anti-endothelial/antitumor activity of protracted administration of SU5416 on human pancreas cancer xenografts, which is comparable with the one obtained by gemcitabine; moreover, the synergistic combination between these drugs on endothelial cells and the promising association in pancreatic cancer xenografts could be used in future studies and translated into the clinical setting.

Synthesis and Cytotoxic and Antitumor Activity of 1,2‐Dihydroxy‐1,2‐dihydrobenzo[b]acronycine Diacid Hemiesters and Carbamates.

AbstractFor Abstract see ChemInform Abstract in Full Text.

Addition of integrin binding sequence to a mutant human endostatin improves inhibition of tumor growth.

Tumor vasculatures express high levels of alphaVbeta3/alphaVbeta5 and alpha5beta1 integrins. Consequently, peptides containing the RGD (Arg-Gly-Asp) sequence, which is present in ligands of integrins, is effective in targeting therapeutic reagents to tumor vascular endothelium. In our study, we investigated whether the biologic activity of endostatin can be enhanced by the addition of an integrin targeting sequence. RGD sequence was added to either the amino or carboxyl terminus of endostatin containing a point mutation, P125A-endostatin. Earlier we have shown that the P125A mutation did not affect the biologic activity of endostatin but in fact had better antiangiogenic activity when compared to the native molecule. Further modification of P125A-endostatin with the RGD motif showed specific and increased binding to endothelial cells, and the increased binding coincided with improved antiangiogenic properties. Both amino and carboxyl terminal RGD-modification of P125A-endostatin resulted in greater inhibition of endothelial cell migration and proliferation. RGD modification increased tumor localization without affecting the circulatory half-life of P125A-endostatin, and RGD-modified P125A-endostatin was found to be more effective when compared to the P125A-endostatin in inhibiting ovarian and colon cancer growth in athymic mice. Complete inhibition of ovarian tumor growth was observed when P125A-endostatin-RGD was encapsulated into alginate beads. These studies demonstrate that addition of a vascular targeting sequence can enhance the biologic activity of an antiangiogenic molecule.

Immunopharmacological and antitumor effects of second-generation immunomodulatory oligonucleotides containing synthetic CpR motifs

Bacterial and synthetic DNAs containing CpG dinucleotides in specific sequence contexts (CpG DNA) activate the vertebrate immune system and produce potent Th1 immune responses. Recently, we reported immunomodulatory oligonucleotides (IMOs) containing 3'-3'-attached novel structures (immunomers) and synthetic immunostimulatory CpR (R=2'-deoxy-7-deazguanosine) dinucleotides show potent stimulatory activity with distinct cytokine secretion profiles. In the present study, we evaluated in vivo immunopharmacological and antitumor properties of second-generation immunomodulatory oligonucleotides (IMOs) either alone or in combination with chemotherapeutic agents. Repeated peritumoral administration of IMOs at 1 mg/kg to mice bearing established subcutaneous CT26 colon tumor or B16.F0 melanoma resulted in complete regression or strong inhibition of tumor growth. Direct peritoneal injection of IMOs at 2.5 mg/kg to mice bearing peritoneally implanted ascites CT26 or B16.F0 tumors completely eradicated or inhibited tumor growth. Treatment of mice bearing beta-gal expressing CT26.CL25 tumor with IMOs resulted in a significant tumor-specific CTL responses compared with treatment with a control non-CpG DNA or PBS. These responses correlated with IFN-gamma, but not IL-4 secreted in IMO, treated mice. A 5-fold increase in beta-gal specific IgG2a antibodies was found in mice, significantly increasing the IgG2a/IgG1 ratio. IMOs showed similar antitumor activity in both wt and IL-6 knockout (ko) C57BL/6 mice but failed to elicit activity in IL-12 ko C57BL/6 mice. Tumor-free mice from the IMO treatment group rejected the same tumor cell rechallenge, suggesting an adaptive immune response against these cells. Moreover, naïve mice quickly developed specific antitumor response without IMO treatment following adoptive transfer of splenocytes obtained from tumor free mice from the IMO treated group. Additionally, the co-administration of IMOs with chemotherapeutic agents, docetaxel and doxorubicin, resulted in synergistic antitumor effects in both B16.F0 melanoma and 4T1 breast carcinoma models. These results demonstrate potent antitumor activity of second-generation IMO compounds containing a synthetic CpR stimulatory motif in a broad spectrum of tumor models through induction of strong Th1 immune responses. IMO treatment resulted in the development of tumor-specific memory immune responses. No treatment-related toxicity was observed in mice at the doses and treatment schedules studied.

Characterization of new estrogen receptor destabilizing compounds: effects on estrogen-sensitive and tamoxifen-resistant breast cancer.

Antiestrogens of the selective estrogen receptor modulator (SERM) type, such as tamoxifen, have two major limitations: their mixed agonist and antagonist profile and the development of tumor resistance. We characterized two new pure antiestrogens-ZK-703 and ZK-253-that belong to the class of specific estrogen receptor destabilizers (SERDs), which includes fulvestrant, and compared their activity with that of fulvestrant and tamoxifen. Effects of antiestrogens on the growth of estrogen-dependent breast tumors in vivo were determined using several mouse xenograft models (including the tamoxifen-sensitive tumors MCF7, T47D, and MV3366 and the tamoxifen-resistant tumors ZR75-1 and MCF7/TAM) and chemically induced (nitrosomethyl urea [NMU] and dimethylbenzanthracene [DMBA]) rat breast cancer models (groups of 10 animals). We determined the initial response and effects on hormone receptor levels and the time to relapse after treatment (i.e., time to reach a predetermined tumor size threshold). Estrogen receptor (ER) levels were determined by immunoassay. ZK-703 (administered subcutaneously) and ZK-253 (administered orally) were more effective than tamoxifen or fulvestrant at inhibiting the growth of ER-positive breast cancer in all xenograft models. For example, MCF7 tumors relapsed (i.e., reached the size threshold) in 10 weeks in mice treated with tamoxifen but in 30 weeks in mice treated with ZK-703. ZK-703 and ZK-253 also prevented further tumor progression in tamoxifen-resistant breast cancer models to a similar extent (more than 30 weeks in mice with ZR75-1 and MCF7/TAM tumors). In the chemically induced rat breast cancer models, orally administered ZK-703 and ZK-253 caused a nearly complete (>80%) inhibition of tumor growth. ER levels were dramatically reduced in MCF7 tumors after 5 weeks of ZK-703 treatment compared with ER levels in vehicle-treated tumors; by contrast, ER levels in tamoxifen-treated tumors were higher than those in control tumors. ZK-703 and ZK-253 are potent, long-term inhibitors of growth in both tamoxifen-sensitive and tamoxifen-resistant breast cancer models.

Tumor specific novel taxoid-monoclonal antibody conjugates.

The basic principle of the targeted delivery approach is that the conjugation of a drug to a tumor-specific molecule renders the drug inactive until it reaches the target site. Monoclonal antibodies (mAbs), which have shown high binding specificity for tumor-specific antigens, could be used as targeting agents. Paclitaxel has brought significant impact on the current cancer chemotherapy, but seriously suffers from the lack of tumor specificity. A series of paclitaxel-monoclonal antibody conjugates via C-2' ester linkage were reported. Taking into account the fact that the cytotoxicity of paclitaxel is not good enough and thus not applicable to this target delivery prodrug approach, new taxoids bearing methyldisulfanyl(alkanoyl) groups were designed, synthesized, and their activities evaluated. A highly cytotoxic C-10 methyldisulfanyl-propanoyl taxoid, was conjugated to monoclonal antibodies recognizing the epidermal growth factor receptor (EGFR). These conjugates were shown to possess remarkable target-specific antitumor activity in vivo against EGFR-expressing A431 tumor xenografts in SCID mice, resulting in complete inhibition of tumor growth in all the treated mice without any noticeable toxicity to the animals.

Synthesis and cytotoxic and antitumor activity of 1,2-dihydroxy-1,2-dihydrobenzo[b]acronycine diacid hemiesters and carbamates.

A series of cis-1,2-dihydroxy-1,2-dihydrobenzo[b]acronycine diacid hemiesters and dicarbamates were prepared by acylation of cis-1,2-dihydroxy-6-methoxy-3,3,14-trimethyl-1,2,3,14-tetrahydro-7H-benzo[b]pyrano[3,2-h]acridin-7-one. The cytotoxicity of the dicarbamates depended on the steric hindrance of the esterifying groups at positions 1 and 2. Diacid hemiesters displayed significant in vitro cytotoxic activities and induced cell cycle perturbations similar to those obtained with cis-1,2-diacetoxy-1,2-dihydrobenzo[b]acronycine (S23906-1) currently under preclinical development. cis-1-Acetoxy-2-hemiglutaryloxy-1,2-dihydrobenzo[b]acronycine was the most promizing compound of the series, inducing complete inhibition of tumor growth when tested against C38 colon adenocarcinoma implanted in mice.

Responsiveness of experimental prostate carcinoma bone tumors to neutron or photon radiation combined with cytokine therapy

Purpose To improve the outcome of radiotherapy for prostate carcinoma bone tumors, we investigated bone tumor irradiation with photons or neutrons followed by interleukin 2 (IL-2) therapy in a tumor model. Methods and materials Implantation of PC-3 cells in nude mouse femur cavity induced a bone tumor that progressed to the formation of a palpable tumor, at the hip joint, by Day 20. Established bone tumors were irradiated with photons or neutrons, and a day later, mice were treated with IL-2 therapy for 3 weekly cycles. Results PC-3 bone tumors responded to radiation with photons or neutrons in a dose-dependent manner. Combination of photon or neutron radiation with IL-2 therapy increased tumor growth delay, compared to that with photons or neutrons alone. Radiation alone or combined with IL-2 significantly increased mouse survival compared to that with IL-2 or no treatment. After combined therapy, a complete inhibition of bone tumor growth was observed in 45% to 50% of the mice. Histologically, the combined therapy resulted in greater tumor destruction associated with fibrosis, new bone formation, and inflammatory infiltrates than that observed with each modality alone. Conclusions The efficacy of tumor irradiation with neutrons or photons was enhanced by IL-2 therapy for the treatment of prostate carcinoma bone tumors.

Antitumor capacity of a dominant-negative RET proto-oncogene mutant in a medullary thyroid carcinoma model.

Gain-of-function mutations in the RET proto-oncogene resulting in a constitutively active receptor tyrosine kinase have been identified as responsible for three subtypes of multiple endocrine neoplasia type 2 (MEN-2) and the development of sporadic medullary and papillary thyroid carcinoma. An important strategy in cancer gene therapy is the inhibition of oncogenic signal transduction by interfering with the molecular mechanisms of activation. In the present study, we tested the therapeutic capacity of an adenovirus expressing a dominant-negative (dn) RET mutant, RET(51).flag, under the control of a synthetic C cell-selective calcitonin promoter (TSE2.CP1) against human medullary thyroid cancer (MTC). Infection of human MTC-derived TT cells with Ad-TSE2.CP1-dn-RET(51).flag resulted in the accumulation of immature RET protein in the endoplasmic reticulum and a strong reduction of oncogenic RET receptor on the cell surface, indicating that RET(51).flag exhibits a dominant-negative effect over endogenous oncogenic protein. Analysis of potential downstream mechanisms associated with the inhibition of oncogenic RET signaling by overexpression of mutant RET(51).flag revealed a significant loss of cell viability in TT cells due to the induction of apoptosis. Finally, we examined the antitumor activity of the dominant-negative RET approach in vivo. Inoculation of Ad-TSE2.CP1- dn-RET(51).flag-expressing MTC cells into nude mice led to complete suppression of tumor growth. Moreover, a single intratumoral injection of Ad-TSE2.CP1-dn-RET(51).flag into established thyroid tumors resulted in prolonged survival of treated mice compared with the controls. Our data suggest that adenoviral delivery of dn-RET(51).flag may be a reliable strategy of effective molecular intervention for RET oncogene-related MTC.

Innate immunity mediated by the cytokine IL-1 homologue 4 (IL-1H4/IL-1F7) induces IL-12-dependent adaptive and profound antitumor immunity.

Recently, several novel members of the IL-1 family have been identified. The possible therapeutic utility and the underlying biologic role of these new members remain unclear. In the present study we analyzed the anti-tumor activity of human IL-1 homologue 4(IL-1H4; renamed IL-F7) by adenovirus-mediated gene transfer (AdIL-1H4) directly into murine tumors. In vitro expression analysis showed that IL-1H4 was a secretory protein. Treatment of an established MCA205 mouse fibrosarcoma by single intratumoral injection of AdIL-1H4 resulted in significant growth suppression. Furthermore, complete inhibition of tumor growth was observed following multiple injections of AdIL-1H4. The anti-tumor activity of IL-1H4 was abrogated in nude and SCID mice and in IL-12-, IFN-gamma-, or Fas ligand-deficient mice. In contrast, IL-1H4 was able to confer substantial anti-tumor effects in NKT-deficient mice. These results suggest that IL-1H4 could play an important role in the link between innate and adaptive immunity and may be useful for tumor immunotherapy.

Tumor-Specific Novel Taxoid−Monoclonal Antibody Conjugates

Taxoids bearing methyldisulfanyl(alkanoyl) groups for taxoid-antibody immunoconjugates were designed, synthesized and their activities evaluated. A highly cytotoxic C-10 methyldisulfanylpropanoyl taxoid was conjugated to monoclonal antibodies recognizing the epidermal growth factor receptor (EGFR) expressed in human squamous cancers. These conjugates were shown to possess remarkable target-specific antitumor activity in vivo against EGFR-expressing A431 tumor xenografts in severe combined immune deficiency mice, resulting in complete inhibition of tumor growth in all the treated mice.

Évaluation des lavages par la polyvidone iodée dans la prévention de l’ensemencement néoplasique péritonéal : étude expérimentale chez le rat

Goal: The aim of the study was to evaluate the in vitro cytototoxicity of diluted povidone iodine on colon cancer cells and its in vivo antitumoral effect in a model of peritoneal carcinomatosis in the rat. Methods: Cell cytotoxicity of a povidone iodine diluted solution was assessed, in vitro, on rat colon cancer cells (DHD/K12/PROb) and human colon cancer cells (HT29). The antitumoral effect of diluted povidone iodine washing was measured in BDIX rats after the intraperitoneal inoculation of 10 6 DHD/K12/PROb cells. Results were compared to an abdominal washing within a 9 g/l salinel solution. In one experiment, peritoneal scars and a colocolic anastomosis were performed after the injection of cancer cells. Results: A short 10 min incubation of human and rat colon cancer cells with diluted povidone iodine resulted in a complete cell killing. In animals, a peritoneal washing with 1 % diluted povidone iodine completely inhibited the tumor growth in parietal peritoneum. However, development of peritoneal tumor nodules was not inhibited in the omentum, in scarified peritoneum or in intestinal anastomosis. Conclusions: Despite its high in vitro efficacy, diluted povidone iodine has an incomplete effect in the prevention of peritoneal carcinomatosis, with only a partial inhibition in scarred peritoneum epiploïc area and intestinal anastomosis. In contrary, it procures a complete inhibition of tumor growth in normal peritoneum.

Hsp110 over-expression increases the immunogenicity of the murine CT26 colon tumor.

Several studies have suggested a positive correlation between heat shock protein (hsp) expression and tumor immunogenicity. Independently, many studies have shown that hsp purified from tumors can be used as a tumor-specific vaccine. In this study, we have explored the connection between hsp expression and anti-tumor immunity by transducing murine CT26 colon carcinoma cells with the cDNA of a major hsp, i.e. hsp110. We have shown that over-expression of hsp110 has no effect on CT26 tumor cell growth in vitro, and does not inhibit their anchorage-independent growth capacity. However, in situ, hsp110 over-expressing CT26 tumor (CT26-hsp110) grew at a significantly reduced rate as compared to the wild-type CT26 tumor in immunocompetent mice. Moreover, immunization of mice with inactivated CT26-hsp110 cells significantly inhibited the growth of wild-type CT26 tumor. This immunity was associated with an increased frequency of tumor-specific T cells after vaccination. An in vivo antibody depletion assay demonstrated that inactivated CT26-hsp110 cells elicited anti-tumor responses involving CD8(+) T cells and natural killer (NK) cells, but not CD4(+) T cells. Lastly, the effect of the addition of granulocyte-macrophage colony stimulating factor (GM-CSF) to these vaccine formulations was determined. Mice immunized with irradiated CT26-hsp110 cells combined with GM-CSF-producing bystander cells revealed a complete inhibition of CT26 tumor growth, indicating a synergy between inactivated CT26-hsp110 vaccine activity and GM-CSF. These observations demonstrate that manipulation of hsp110 expression in tumors, specifically when combined with GM-CSF, represents a potentially powerful approach to cancer vaccine formulation.

In vitro and preclinical targeted alpha therapy of human prostate cancer with Bi-213 labeled J591 antibody against the prostate specific membrane antigen

Limited options for the treatment of prostate cancer have spurred the search for new therapies. One innovative approach is the use of targeted alpha therapy (TAT) to inhibit cancer growth, using an alpha particle emitting radioisotope such as (213)Bi. Because of its short range and high linear energy transfer (LET), alpha-particles may be particularly effective in the treatment of cancer, especially in inhibiting the development of metastatic tumors from micro-metastases. Prostate-specific membrane antigen (PSMA) is expressed in prostate cancer cells and the neovasculature of a wide variety of malignant neoplasms including lung, colon, breast and others, but not in normal vascular endothelium. The expression is further increased in higher-grade cancers, metastatic disease and hormone-refractory prostate cancer (PCA). J591 is one of several monoclonal antibodies (mabs) to the extracellular domain of PSMA. Chelation of J591 mab with (213)Bi forms the alpha-radioimmunoconjugate (AIC). The objective of this preclinical study was to design an injectable AIC to treat human prostate tumors growing subcutaneously in mice. The anti-proliferative effects of AIC against prostate cancer were tested in vitro using the MTS assay and in vivo with the nude mice model. Apoptosis was documented using terminal deoxynucleotidyl transferase [TdT]-mediated deoxyuridinetriphosphate [dUTP] nick end-labeling (TUNEL) assay, while proliferative index was assessed using the Ki-67 marker. We show that a very high density of PSMA is expressed in an androgen-dependent human PCA cell line (LNCaP-LN3) and in tumor xenografts from nude mice. We also demonstrate that the AIC extensively inhibits the growth of LN3 cells in vitro in a concentration-dependent fashion, causing the cells to undergo apoptosis. Our in vivo studies showed that a local AIC injection of 50 microCi at 2 days post-cell inoculation gave complete inhibition of tumor growth, whereas results for a non-specific AIC were similar to those for untreated mice. Further, after 1 and 3 weeks post-tumor appearance, a single (100 microCi/100 microl) intra-lesional injection of AIC can inhibit the growth of LN3 tumor xenografts (volume<100 mm(3)) in nude mice. Tumors treated with AIC decreased in volume from a mean 46+/-14 mm(3) in the first week or 71+/-15 mm(3) in the third week to non-palpable, while in control mice treated with a non-specific AIC using the same dose, tumor volume increased from 42 to 590 mm(3). There were no observed side effects of the treatment. Because of its in vitro cytotoxicity and these anti-proliferative properties in vivo, the (213)Bi-J591 conjugate has considerable potential as a new therapeutic agent for the treatment of prostate cancer.

Lipofection of early passages of cell cultures derived from murine adenocarcinomas: in vitro and ex vivo testing of the thymidine kinase/ganciclovir system.

Early passages of cultured cells derived from four spontaneous Balb/c murine adenocarcinomas were used to explore the feasibility of a nonviral HSVtk-based suicide gene therapy system. After lipofection with pCMVtk, the transiently HSVtk expressing P07 (lung), M3, M05, and M38 (mammary gland) cells were, respectively, about 130-, 30-, 120-, and 170-fold more sensitive to ganciclovir (GCV) in vitro than their respective controls. Eighty percent of Balb/c mice subcutaneously inoculated with ex vivo pCMVtk-lipofected P07 cells, followed by intraperitoneal GCV injection for 7 days, displayed a complete inhibition of tumor growth for over 70 days. Control animals started to display tumors 13 days after inoculation. We present evidence showing that early passages of cultured tumor cells can efficiently express lipofected genes and that they are sensitive to the lipoplex-mediated HSVtk/GCV system.

Targeting cyclooxygenase 2 and her-2/neu pathways inhibits colorectal carcinoma growth

The cyclooxygenase 2 (COX-2) and ErbB/HER pathways are important modulators of cancer cell growth. We sought to determine the effects of treatment with a specific COX-2 inhibitor and/or a monoclonal antibody against the ErbB receptor subtype HER-2/neu on carcinoma cell growth. A cell-proliferation assay was used to determine the response of HCA-7 cells to the HER-3/HER-4 ligand heregulin beta-1 (HRGbeta-1). Both in vitro and in vivo assays were used to determine the effects of the selective COX-2 inhibitor, celecoxib, and/or an anti-HER-2/neu monoclonal antibody (either Herceptin [Genetech Inc., S. San Francisco, CA] or 2C4) on cell growth. HCA-7 cells express HER-2/neu messenger RNA and protein, and exposure of these cells to HRGbeta-1 results in a significant stimulation of cell growth. Celecoxib or Herceptin inhibits HCA-7 cell growth in vitro and in vivo. Combination therapy with celecoxib plus Herceptin or celecoxib plus 2C4 resulted in additive effects that resulted in almost complete inhibition of tumor growth. Combined treatment with COX-2 and HER-2/neu inhibitors more effectively reduces colorectal carcinoma growth than either agent alone. Therefore, targeting of both the COX-2 and ErbB signaling pathways may represent a novel approach for the treatment and/or prevention of colorectal cancer in humans.

Synergistic anti-tumor activity of a novel immunomodulator, BCH-1393, in combination with cyclophosphamide

N, N-dimethylaminopurine pentoxycarbonyl D-arginine (BCH-1393) is a novel low molecular weight synthetic immunomodulator that has been shown to significantly stimulate cytotoxic T-lymphocyte responses both in vitro and in vivo (Zacharie B, Gagnon L, Attardo G, Connolly TP, St-Denis Y, Penney CL. Synthesis and activity of 6-substituted purine linker amine immunostimulants. J. Med. Chem. 1997;40:2883–94). Prompted by this evidence, we extended evaluation of BCH-1393 for anticancer activity in syngeneic mouse experimental tumor models. Consistent with previous findings, in vitro assessment of BCH-1393 activity demonstrated a significant increase in the CTL responses in the range of 10 −9–10 −5 M. Treatment of mice with four consecutive daily intraperitoneal injections at 25 and 50 mg/kg resulted in a significant increase of the relative percentage of blood CD4+, CD8+, NK and monocyte subsets without any evidence of toxicity. In vivo anti-tumor activity of BCH-1393 was evaluated, either alone or in combination with subtherapeutic doses of cyclophosphamide (Cy), against weakly immunogenic mouse breast carcinoma DA-3 and strongly immunogenic colon adenocarcinoma MC38. Daily intraperitoneal injection of BCH-1393 at 50 mg/kg alone was well tolerated but produced a relatively weak anti-tumor effect in both tumor models. However, a significant inhibition of tumor outgrowth and suppression of established tumor growth was observed when BCH-1393 was administered in combination with subtherapeutic doses of Cy. Combination treatment of 50 mg/kg BCH-1393 with 100 mg/kg Cy (given as single intravenous bolus injection) starting 2 days prior to DA-3 tumor cell inoculation prevented tumor outgrowth in 70–80% of treated mice. In the remaining 20–30% of mice that had developed tumors, a nearly complete (90%) tumor growth inhibition was observed at days 22–24 post tumor implant. In the MC38 tumor model, combination treatment of established tumors with BCH-1393 and Cy (CTX) at 50 mg/kg resulted in a significant delay in tumor growth compared to CTX treatment alone. The observed concomitant anti-tumor activity of BCH-1393 with cyclophosphamide warrants further investigation of this immunomodulator as an adjunctive treatment of cancer.

Expression of the Death Gene Bik/Nbk Promotes Sensitivity to Drug-Induced Apoptosis in Corticosteroid-Resistant T-Cell Lymphoma and Prevents Tumor Growth in Severe Combined Immunodeficient Mice

Members of the Bcl-2 gene family have been implicated in the regulation of cell death induced by cytostatic drugs. In some malignancies such as B-cell lymphoma, there is evidence that high expression of Bcl-2 is an independent negative prognostic marker and the overexpression of Bcl-2 has been shown to confer resistance to cytotoxic drugs by preventing drug-induced apoptosis. This function of Bcl-2 can be antagonized by apoptosis-promoting members of the Bcl-2 family. We previously showed that overexpression of Bax restores the chemosensitivity of Bax-deficient breast cancer cell lines. Therefore, we investigated whether the death-promoting Bcl-2 homologue Bik/Nbk can enhance cytostatic drug-induced apoptosis. As a model, we used the T-cell leukemia H9 (CD3(+) and CD4(+)CD8(-)), which is resistant to corticosteroid-induced cell death and does not express endogenous Bik/Nbk. Sensitivity for drug-induced apoptosis was increased 10- to 39-fold in cells transfected with the full-length coding sequence of Bik/Nbk. In addition, apoptosis induced via CD95/Fas or heat shock was increased to a similar extent. These data show that Bik/Nbk, which, unlike Bax, carries only a BH3 but no BH1 or BH2 domain may be a target to enhance chemosensitivity. The complete suppression of tumor growth in a severe combined immunodeficient mouse xenotransplant model suggests that, in analogy to Bax, Bik/Nbk may function as a tumor suppressor gene.