Inhibition Of Human Tumor Growth Research Articles

Physical and biological properties of homophilic therapeutic antibodies.

Homophilic antibodies have been discovered in mice and primates and can also be engineered. Compared to conventional antibodies, homophilic antibodies form lattices on targets leading to enhanced binding via polyvalent attachment. Previously, we have observed a paradoxical dose/potency effect with an engineered homophilic antibody against a human lung cancer tumor. Here, we have investigated some biophysical properties of homophilic antibodies and also studied the inhibition of human tumor growth in a xenograft model using homophilic Herceptin. Dimerization and viscosity of two homophilic antibodies are greater at physiological temperature than at 4°C. Similarly, binding to solid-phase antigen is greater at 37°C than at room temperature or 4°C. Dimer formation is higher at therapeutic concentration, supporting the notion that preformed dimers in solution are the effective molecular species responsible for polyvalent target binding and enhanced therapeutic potency.

AT7867 Is a Potent and Oral Inhibitor of AKT and p70 S6 Kinase That Induces Pharmacodynamic Changes and Inhibits Human Tumor Xenograft Growth

The serine/threonine kinase AKT plays a pivotal role in signal transduction events involved in malignant transformation and chemoresistance and is an attractive target for the development of cancer therapeutics. Fragment-based lead discovery, combined with structure-based drug design, has recently identified AT7867 as a novel and potent inhibitor of both AKT and the downstream kinase p70 S6 kinase (p70S6K) and also of protein kinase A. This ATP-competitive small molecule potently inhibits both AKT and p70S6K activity at the cellular level, as measured by inhibition of GSK3beta and S6 ribosomal protein phosphorylation, and also causes growth inhibition in a range of human cancer cell lines as a single agent. Induction of apoptosis was detected by multiple methods in tumor cells following AT7867 treatment. Administration of AT7867 (90 mg/kg p.o. or 20 mg/kg i.p.) to athymic mice implanted with the PTEN-deficient U87MG human glioblastoma xenograft model caused inhibition of phosphorylation of downstream substrates of both AKT and p70S6K and induction of apoptosis, confirming the observations made in vitro. These doses of AT7867 also resulted in inhibition of human tumor growth in PTEN-deficient xenograft models. These data suggest that the novel strategy of AKT and p70S6K blockade may have therapeutic value and supports further evaluation of AT7867 as a single-agent anticancer strategy.

Inhibition of human tumor growth in mice by an oncolytic herpes simplex virus designed to target solely HER-2-positive cells

Oncolytic virotherapy exploits the ability of viruses to infect, replicate into, and kill tumor cells. Among the viruses that entered clinical trials are HSVs. HSVs can be engineered to become tumor-specific by deletion of selected genes or retargeting to tumor-specific receptors. A clinically relevant surface molecule is HER-2, hyperexpressed in one fourth of mammary and ovary carcinomas, and associated with high metastatic ability. As a previously undescribed strategy to generate HSV recombinants retargeted to HER-2 and detargeted from natural receptors, we replaced the Ig-folded core in the receptor-binding virion glycoprotein gD with anti-HER-2 single-chain antibody. The recombinant entered cells solely via HER-2 and lysed HER-2-positive cancer cells. Because of the high specificity, its safety profile in i.p. injected mice was very high, with a LD(50) >5 x 10(8) pfu, a figure at least 10,000-fold higher than that of corresponding WT-gD carrying virus (LD(50) approximately 5 x 10(4) pfu). When administered intratumorally to nude mice bearing HER-2-hyperexpressing human tumors, it strongly inhibited progressive tumor growth. The results provide a generally applicable strategy to engineer HSV recombinants retargeted to a wide range of receptors for which a single-chain antibody is available, and show the potential for retargeted HSV to exert target-specific inhibition of human tumor growth. Therapy with HER-2-retargeted oncolytic HSV could be effective in combined or sequential protocols with monoclonal antibodies and small inhibitors, particularly in patients resistant to HER-2-targeted therapy because of alterations in HER-2 signaling pathway, or against brain metastases inaccessible to anti-HER-2 antibodies.

Targeting Vacuolar H+-ATPases as a New Strategy against Cancer

Growing evidence suggests a key role of tumor acidic microenvironment in cancer development, progression, and metastasis. As a consequence, the need for compounds that specifically target the mechanism(s) responsible for the low pH of tumors is increasing. Among the key regulators of the tumor acidic microenvironment, vacuolar H(+)-ATPases (V-ATPases) play an important role. These proteins cover a number of functions in a variety of normal as well as tumor cells, in which they pump ions across the membranes. We discuss here some recent results showing that a molecular inhibition of V-ATPases by small interfering RNA in vivo as well as a pharmacologic inhibition through proton pump inhibitors led to tumor cytotoxicity and marked inhibition of human tumor growth in xenograft models. These results propose V-ATPases as a key target for new strategies in cancer treatment.

53 INVITED HPMA copolymer-TNP-470 (caplostatin) and Avastin show synergistic inhibition of human tumor growth in mice

Progression of cancer is dependent on acquisition of vascular networks within the tumor. Tumor angiogenesis is dependent on up-regulation of angiogenesis stimulators to overcome the endogenous anti-angiogenic barrier. Such disruption of angiogenesis balance to favor neovascularization is a key step for progression of tumor growth and metastasis. In this regard, the vascular basement membrane and the extracellular matrix have been found to be rich sources of angiogenesis stimulators and inhibitors that become bioavailable on proteolysis of the matrix by tumor microenvironment–related enzymes. In this review the subgroup of endogenous angiogenesis stimulators and inhibitors is discussed, and their mechanism of action during tumor angiogenesis is evaluated. The role in regulating tumor growth and the possibility of using them as prognostic markers for human gastrointestinal cancers is discussed. Furthermore, we specifically address the role of vascular endothelial growth factor in human gastrointestinal cancers and discuss the development and use of bevacizumab (Avastin; anti–vascular endothelial growth factor antibody [Genentech, CA]) in the treatment of colorectal and other gastrointestinal cancers.

The phosphoinositide 3-OH kinase/AKT2 pathway as a critical target for farnesyltransferase inhibitor-induced apoptosis.

Farnesyltransferase inhibitors (FTIs) represent a novel class of anticancer drugs that exhibit a remarkable ability to inhibit malignant transformation without toxicity to normal cells. However, the mechanism by which FTIs inhibit tumor growth is not well understood. Here, we demonstrate that FTI-277 inhibits phosphatidylinositol 3-OH kinase (PI 3-kinase)/AKT2-mediated growth factor- and adhesion-dependent survival pathways and induces apoptosis in human cancer cells that overexpress AKT2. Furthermore, overexpression of AKT2, but not oncogenic H-Ras, sensitizes NIH 3T3 cells to FTI-277, and a high serum level prevents FTI-277-induced apoptosis in H-Ras- but not AKT2-transformed NIH 3T3 cells. A constitutively active form of AKT2 rescues human cancer cells from FTI-277-induced apoptosis. FTI-277 inhibits insulin-like growth factor 1-induced PI 3-kinase and AKT2 activation and subsequent phosphorylation of the proapoptotic protein BAD. Integrin-dependent activation of AKT2 is also blocked by FTI-277. Thus, a mechanism for FTI inhibition of human tumor growth is by inducing apoptosis through inhibition of PI 3-kinase/AKT2-mediated cell survival and adhesion pathway.

Inhibition of the prenylation of K-Ras, but not H- or N-Ras, is highly resistant to CAAX peptidomimetics and requires both a farnesyltransferase and a geranylgeranyltransferase I inhibitor in human tumor cell lines.

The farnesyltransferase (FTase) inhibitor FTI-277 is highly effective at blocking oncogenic H-Ras but not K-Ras4B processing and signaling. While inhibition of processing and signaling of oncogenic K-Ras4B is more sensitive to the geranylgeranyltransferase I (GGTase I) inhibitor GGTI-286 than it is to FTI-277 in K-Ras4B-transformed NIH3T3 cells, the sensitivity of K-Ras as well as H- and N-Ras to the CAAX peptidomimetics in human tumor cell lines is not known. Here, we report that a panel of five human carcinoma cell lines from pancreatic, pulmonary, and bladder origins all express H-, N-, and K-Ras, and their respective prenylation sensitivities to the FTase and GGTase I inhibitors is variable. In all of the cell lines investigated, the prenylation of N-Ras was highly sensitive to FTI-277, and in two of the cell lines, N-Ras showed slight sensitivity to GGTI-298, an analog of GGTI-286. Although the prenylation of H-Ras was also sensitive to FTI-277, complete inhibition of H-Ras processing even at high concentrations of FTI-277 and/or GGTI-298 was never achieved. The prenylation of K-Ras, on the other hand, was highly resistant to FTI-277 and GGTI-298. Most significantly, treatment of human tumor cell lines with both inhibitors was required for inhibition of K-Ras prenylation. In one cell line, the human lung adenocarcinoma A-549, prenylation of K-Ras was highly resistant even when co-treated with both inhibitors. Furthermore, soft agar experiments demonstrated that in all the human tumor cell lines tested inhibition of K-Ras prenylation was not necessary for inhibition of anchorage-independent growth. In addition, although GGTI-298 had very little effect on soft agar growth, the combination of FTI-277 and GGTI-298 resulted in significant growth inhibition. Therefore, the results demonstrate that while FTI-277 inhibits N-Ras and H-Ras processing in the human tumor cell lines evaluated, inhibition of K-Ras processing requires both an FTase inhibitor as well as a GGTase I inhibitor, and that inhibition of human tumor growth in soft agar does not require inhibition of oncogenic K-Ras processing.

Expression behavior of 85-kDa membrane protein in adriamycin-resistant tumor cells and the inhibition of human tumor growth in athymic mice by MRK-20 monoclonal antibody against the protein.

Treatment of tumors with monoclonal antibodies against tumor antigen is one of the selective modalities for cancer therapy. We examined the therapeutic effect of MRK‐20 (IgG1), a murine monoclonal antibody against resistance‐associated 85‐kDa membrane protein. The 85‐kDa protein is expressed on the surface of multidrug‐resistant cells induced by adriamycin. This protein is also expressed in some multiple‐drug‐resistant cells, including atypical multidrug‐resistant cells. The protein, once lost during long‐term culture without adriamycin, was rapidly induced by treatment with adriamycin but not with vinblastine or etoposide, suggesting a close relationship of the protein with adriamycin resistance but not with multidrug resistance. The antibody MRK‐20 suppressed the growth of subcutaneously implanted tumors expressing the 85‐kDa protein. Adriamycin‐resistant human ovarian tumor 2780AD and innately resistant human erythroleukemia HEL cells in athymic mice were completely cured by treatment with MRK‐20 antibody when the antibody was administered i.v. 2 days after s.c. tumor implantation. On the other hand, MRK‐20 did not show any effect on the growth of the 85‐kDa protein‐negative A2780 human ovarian tumor. These results indicate that the effect of MRK‐20 is highly specific to cells expressing 85‐kDa protein.

Inhibition of human tumor growth in nude mice by a conjugate of doxorubicin with monoclonal antibodies to epidermal growth factor receptor.

Monoclonal antibodies that recognize the extracellular domain of the epidermal growth factor receptor (mAb108) were conjugated with doxorubicin through a dextran bridge. Several antibody-drug conjugates, containing different amounts of doxorubicin, retained binding capacity to human epidermoid carcinoma (KB) cells overexpressing epidermal growth factor receptors. Slight decrease in drug cytotoxicity was seen in in vitro tests, as determined either by inhibition of thymidine incorporation into cells or by reduction in number and size of KB-cell colonies. Yet, when tested in vivo against KB tumor xenografted into nude mice, the anti-epidermal growth factor-receptor drug conjugates with high drug-substitution levels were significantly more effective than free doxorubicin, antibody alone, mixture of dextran-doxorubicin and antibody, or drug conjugated with irrelevant antibody. When the labile covalent bonds linking antibody to dextran bridge were stabilized by reduction, the therapeutic efficacy of the conjugate was markedly decreased. These results show that antibodies against the extracellular domain of the epidermal growth factor can deliver doxorubicin specifically and efficiently to tumor sites that express high receptor levels exerting a specific antitumor effect.

Inhibition of human tumor growth by IgG2A monoclonal antibodies correlates with antibody density on tumor cells.

Eight monoclonal antibodies (MAb) of IgG2a isotype that were produced against human melanomas were tested for tumor growth-inhibiting properties in nude mice injected with human melanoma cells of various origins. Four of the eight MAb inhibited growth of these tumors, and all four of these antibodies reacted in antibody-dependent macrophage-mediated cytotoxicity (ADMC) assays in vitro. The MAb that were inactive in vivo also did not react in these assays in vitro. The number of antibody-binding sites per cell on the tumor cell surface was significantly higher for tumoricidal MAb as compared to unreactive MAb. On the other hand, the percentage of tumor cells binding the MAb and the binding affinity to these cells were the same for the two groups of MAb. Also, tumoricidal and nontumoricidal MAb bound with similar affinity and antibody density to Fc receptors on macrophages. The importance of the number of antibody sites on the tumor cell surface for tumor destruction by MAb was confirmed by the demonstration of tumoricidal effects of mixtures of MAb that were by themselves not tumoricidal. MAb binding to different molecules on melanoma cells were complementary in ADMC, whereas MAb directed to the same molecule but to different epitopes were not.