Toll-like receptor 9 agonist IMO cooperates with everolimus in renal cell carcinoma by interfering with tumour growth and angiogenesis.

BACKGROUND: Targeting the mammalian target of rapamycin by everolimus is a successful approach for renal cell carcinoma (RCC) therapy. The Toll-like receptor 9 agonist immune modulatory oligonucleotide (IMO) exhibits direct antitumour and antiangiogenic activity and cooperates with both epidermal growth factor receptor (EGFR) and vascular endothelial growth factor (VEGF) inhibitors. METHODS: We tested the combination of IMO and everolimus on models of human RCC with different Von-Hippel Lindau (VHL) gene status, both in vitro and in nude mice. We studied their direct antiangiogenic effects on human umbilical vein endothelial cells. RESULTS: Both IMO and everolimus inhibited in vitro growth and survival of RCC cell lines, and their combination produced a synergistic inhibitory effect. Moreover, everolimus plus IMO interfered with EGFR-dependent signaling and reduced VEGF secretion in both VHL wild-type and mutant cells. In RCC tumour xenografts, IMO plus everolimus caused a potent and long-lasting cooperative antitumour activity, with reduction of tumour growth, prolongation of mice survival and inhibition of signal transduction. Furthermore, IMO and everolimus impaired the main endothelial cell functions. CONCLUSION: A combined treatment with everolimus and IMO is effective in VHL wild-type and mutant models of RCC by interfering with tumour growth and angiogenesis, thus representing a potentially effective, rationale-based combination to be translated in the clinical setting.

Two novel human anti-ErbB2 immunoagents are active on trastuzumab-resistant tumours.

BACKGROUND: Overexpression of ErbB2 receptor in breast cancer is associated with disease progression and poor prognosis. Trastuzumab, the only humanised anti-ErbB2 antibody currently used in breast cancer, has proven to be effective; however, a relevant problem for clinical practice is that a high fraction of breast cancer patients shows primary or acquired resistance to trastuzumab treatment. METHODS: We tested on trastuzumab-resistant cells two novel human anti-tumour immunoconjugates engineered in our laboratory by fusion of a human anti-ErbB2 scFv, termed Erbicin, with either a human RNase or the Fc region of a human IgG1. Both Erbicin-derived immunoagents (EDIAs) are selectively cytotoxic for ErbB2-positive cancer cells in vitro and vivo, target an ErbB2 epitope different from that recognised by trastuzumab and do not show cardiotoxic effects. RESULTS: We report that EDIAs are active also on trastuzumab-resistant tumour cells both in vitro and in vivo, most likely because of the different epitope recognised, as EDIAs, unlike trastuzumab, were found to be able to inhibit the signalling pathway downstream of ErbB2. CONCLUSION: These results suggest that EDIAs are immunoagents that could not only fulfil the therapeutic need of patients ineligible to trastuzumab treatment due to cardiac dysfunction but also prove to be useful for breast cancer patients unresponsive to trastuzumab treatment.

Enzastaurin inhibits tumours sensitive and resistant to anti-EGFR drugs.

We investigated the antitumour effect and ability to overcome the resistance to anti-EGFR drugs of enzastaurin, an inhibitor of VEGFR-dependent PKCbeta signalling. Enzastaurin was evaluated alone and in combination with the EGFR inhibitor gefitinib, on growth and signalling protein expression in human cancer cells sensitive and resistant to anti-EGFR drugs, both in vitro and in nude mice. We demonstrated the marked inhibitory activity of enzastaurin against GEO colon and PC3 prostate cancer cells and their gefitinib-resistant counterparts GEO-GR and PC3-GR, accompanied by inhibition of pAkt and its effector pp70S6K, pGSK3beta and VEGF expression and secretion. Moreover, enzastaurin showed a cooperative effect with gefitinib in parental and in gefitinib-resistant cells. Remarkably, these results were confirmed in vivo, where enzastaurin showed antitumour activity and cooperativity with gefitinib in mice grafted with GEO and GEO-GR tumours, incrementing their median survival and inhibiting the aforesaid protein expression and secretion in tumour specimens. In conclusion, enzastaurin by interfering with signalling proteins implicated in EGFR drug resistance markedly cooperates with gefitinib in sensitive and gefitinib-resistant tumours, thus overcoming and reverting such resistance and providing a rational basis for its development in patients resistant to anti-EGFR drugs.

Inhibition of mTOR pathway by everolimus cooperates with EGFR inhibitors in human tumours sensitive and resistant to anti-EGFR drugs.

Inhibition of a single transduction pathway is often inefficient due to activation of alternative signalling. The mammalian target of rapamycin (mTOR) is a key intracellular kinase integrating proliferation, survival and angiogenic pathways and has been implicated in the resistance to EGFR inhibitors. Thus, mTOR blockade is pursued to interfere at multiple levels with tumour growth. We used everolimus (RAD001) to inhibit mTOR, alone or in combination with anti-EGFR drugs gefitinib or cetuximab, on human cancer cell lines sensitive and resistant to EGFR inhibitors, both in vitro and in vivo. We demonstrated that everolimus is active against EGFR-resistant cancer cell lines and partially restores the ability of EGFR inhibitors to inhibit growth and survival. Everolimus reduces the expression of EGFR-related signalling effectors and VEGF production, inhibiting proliferation and capillary tube formation of endothelial cells, both alone and in combination with gefitinib. Finally, combination of everolimus and gefitinib inhibits growth of GEO and GEO-GR (gefitinib resistant) colon cancer xenografts, activation of signalling proteins and VEGF secretion. Targeting mTOR pathway with everolimus overcomes resistance to EGFR inhibitors and produces a cooperative effect with EGFR inhibitors, providing a valid therapeutic strategy to be tested in a clinical setting.

The rationale for the combination of selective EGFR inhibitors with cytotoxic drugs and radiotherapy.

The epidermal growth factor receptor (EGFR) is frequently overexpressed in a wide range of human tumors; such overexpression often correlates with poor prognosis and worse clinical outcome. It has been demonstrated that the EGFR autocrine pathway plays a crucial role in human cancer since it contributes to a number of highly relevant processes in tumor development and progression, including cell proliferation, regulation of apoptotic cell death, angiogenesis and metastatic spread. For these reasons EGFR is one of the most studied and exploited targets for molecular cancer therapy. Two classes of anti-EGFR agents have entered clinical practice: monoclonal antibodies and small molecules targeting receptor tyrosine kinases. The possibility of combining conventional cytotoxic drugs with agents that specifically interfere with key pathways controlling cancer cell survival, proliferation, invasion and/or metastatic spread has generated wide interest. This could be a promising therapeutic approach for several reasons. First, the occurrence of cross-resistance is infrequent since the cellular targets and mechanisms of action of cytotoxic drugs and EGFR antagonists are different. Second, alterations in the expression and/or activity of genes that regulate mitogenic signals may either cause perturbation of cell growth or affect the sensitivity of cancer cells to conventional chemotherapy and radiotherapy. In fact, EGFR inhibitors have shown activity alone and/or in combination with conventional antitumor treatments.

The rationale for the combination of selective EGFR inhibitors with cytotoxic drugs and radiotherapy.

The epidermal growth factor receptor (EGFR) is frequently overexpressed in a wide range of human tumors; such overexpression often correlates with poor prognosis and worse clinical outcome. It has been demonstrated that the EGFR autocrine pathway plays a crucial role in human cancer since it contributes to a number of highly relevant processes in tumor development and progression, including cell proliferation, regulation of apoptotic cell death, angiogenesis and metastatic spread. For these reasons EGFR is one of the most studied and exploited targets for molecular cancer therapy. Two classes of anti-EGFR agents have entered clinical practice: monoclonal antibodies and small molecules targeting receptor tyrosine kinases. The possibility of combining conventional cytotoxic drugs with agents that specifically interfere with key pathways controlling cancer cell survival, proliferation, invasion and/or metastatic spread has generated wide interest. This could be a promising therapeutic approach for several reasons. First, the occurrence of cross-resistance is infrequent since the cellular targets and mechanisms of action of cytotoxic drugs and EGFR antagonists are different. Second, alterations in the expression and/or activity of genes that regulate mitogenic signals may either cause perturbation of cell growth or affect the sensitivity of cancer cells to conventional chemotherapy and radiotherapy. In fact, EGFR inhibitors have shown activity alone and/or in combination with conventional antitumor treatments.