Dhanvanthri S. Deevi

Synergistic Antitumor Effects of Combined Epidermal Growth Factor Receptor and Vascular Endothelial Growth Factor Receptor-2 Targeted Therapy

Purpose: Combination therapies that target the epidermal growth factor receptor (EGFR) and vascular endothelial growth factor receptor (VEGFR) pathways, are being actively tested for the treatment of cancer. In evaluating combination strategies, the ideal combination would be one in which the treatments interact in a way that is synergistic with regard to antitumor effects. Here, we have evaluated the interaction between anti-EGFR antibody Erbitux (cetuximab) and anti-VEGFR2 antibody, DC101, in preclinical models of pancreatic (BxPC-3) and colon (GEO) cancer. Experimental Design: Analysis of the interaction between cetuximab and DC101 in vivo used a novel method for establishing the upper 95% confidence limits for the combination index (CI) of isobologram analyses, where CI < 1 indicates synergy. Assessment of tumor cell proliferation, apoptosis, VEGF production, and hypoxia, as well as tumor vascularization, was performed to gain insights into the mechanistic basis for synergy between agents targeting different tumor compartments. Results: Monotherapy ED50 values for tumor growth inhibition ranged from 1.8 to 2.3 mg/kg and 10.5 to 16.6 mg/kg for cetuximab and DC101, respectively. From the dose response of the combination treatment, it was determined that cetuximab and DC101 are synergistic in the BxPC-3 (CI = 0.1, P < 0.01) and GEO (CI = 0.1, P < 0.01) models. Overlapping effects on the tumor cell and vascular compartments form a basis for the interaction, with VEGF production and hypoxia-inducible factor 1α potentially acting as molecular links between EGFR and VEGFR2 inhibition. Conclusions: Results show antitumor synergy for combined EGFR and VEGFR2 targeted therapy, supporting the significant therapeutic potential of this combination strategy.

Targeting the platelet-derived growth factor receptor α with a neutralizing human monoclonal antibody inhibits the growth of tumor xenografts: Implications as a potential therapeutic target

Platelet-derived growth factor receptor α (PDGFRα) is a type III receptor tyrosine kinase that is expressed on a variety of tumor types. A neutralizing monoclonal antibody to human PDGFRα, which did not cross-react with the β form of the receptor, was generated. The fully human antibody, termed 3G3, has a Kd of 40 pmol/L and blocks both PDGF-AA and PDGF-BB ligands from binding to PDGFRα. In addition to blocking ligand-induced cell mitogenesis and receptor autophosphorylation, 3G3 inhibited phosphorylation of the downstream signaling molecules Akt and mitogen-activated protein kinase. This inhibition was seen in both transfected and tumor cell lines expressing PDGFRα. The in vivo antitumor activity of 3G3 was tested in human glioblastoma (U118) and leiomyosarcoma (SKLMS-1) xenograft tumor models in athymic nude mice. Antibody 3G3 significantly inhibited the growth of U118 (P = 0.0004) and SKLMS-1 (P < 0.0001) tumors relative to control. These data suggest that 3G3 may be useful for the treatment of tumors that express PDGFRα.

Tumor necrosis factor-alpha and interleukin-1 antagonists alleviate inflammatory skin changes associated with epidermal growth factor receptor antibody therapy in mice.

Cancer patients receiving epidermal growth factor receptor (EGFR) antibody therapy often experience an acneiform rash of uncertain etiology in skin regions rich in pilosebaceous units. Currently, this condition is treated symptomatically with very limited, often anecdotal success. Here, we show that a monoclonal antibody targeting murine EGFR, ME1, caused a neutrophil-rich hair follicle inflammation in mice, similar to that reported in patients. This effect was preceded by the appearance of lipid-filled hair follicle distensions adjacent to enlarged sebaceous glands. The cytokine tumor necrosis factor-alpha (TNFalpha), localized immunohistochemically to this affected region of the pilosebaceous unit, was specifically up-regulated by ME1 in skin but not in other tissues examined. Moreover, skin inflammation was reduced by cotreatment with the TNFalpha signaling inhibitor, etanercept, indicating the involvement of TNFalpha in this inflammatory process. Interleukin-1, a cytokine that frequently acts in concert with TNFalpha, is also involved in this process given the efficacy of the interleukin-1 antagonist Kineret. Our results provide a mechanistic framework to develop evidence-based trials for EGFR antibody-induced skin rash in patients with cancer.

Abstract 5030: Necitumumab (IMC-11F8), a recombinant human anti EGFR antibody, increases the antitumor effects of cisplatin/paclitaxel in human NSCLC xenograft models

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FLnnEpidermal growth factor receptor (EGFR) targeted antibodies may have a role in the treatment of non-small cell lung cancer (NSCLC), based on preclinical and clinical data. Here we have demonstrated a significant benefit of adding necitumumab, a recombinant human IgG1 antibody targeting EGFR, to cisplatin+paclitaxel administered at its maximum tolerated dose, in subcutaneous xenograft tumor models established in nu/nu athymic mice with A549 or NCI-H1650 NSCLC cell lines.nnNecitumumab monotherapy or cisplatin+paclitaxel therapy inhibited xenograft tumor growth with a T/C% of 47 and 63, respectively in A549 tumor model. A similar anti tumor effect was seen in NCI-H1650 xenograft tumors treated with necitumumab monotherapy or cisplatin+paclitaxel therapy with a T/C% of 37 and 50, respectively. Furthermore, the combination of necitumumab with cisplatin+paclitaxel significantly inhibited tumor growth with a magnitude greater than any of the therapies alone in A549 xenograft tumors with a T/C% of 33 and in NCI-H1650 xenograft tumors with a T/C% of 18. Combination treatment resulted in partial regressions (3 out of 9 mice) in A549 and maximum tumor regressions (10 out of 12) in NCI-H1650. Chi-Squared test for all animals in the treatment groups showed p=0.02 in A549 and p=3.0E-08 in NCI-H1650 xenograft tumors.nnIn vitro analysis on A549 and NCI-H1650 cell lines treated with cisplatin+paclitaxel, with/without necitumumab (5 μg/ml) demonstrated that this combination benefit was associated with the increased apoptotic index in both models. Cell cycle arrest during combination therapy was also observed as an increase in the percentage of cells in G2/M phase in A549 and S or G2/M phase in NCI-H1650 cell lines. We are further utilizing SABiosciences PCR arrays to demonstrate combination benefit related gene function in apoptosis, cell cycle signaling pathways and drug specific biomarkers in both A549 and NCI-H1650 cell lines.nnThus our results demonstrate the potential utility of necitumumab for enhancing the efficacy of cytotoxic chemotherapy in both in A549 and NCI-H1650 cell lines and xenograft tumors models.nnCitation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5030. doi:10.1158/1538-7445.AM2011-5030

Abstract 1187: The role of PI3K as a biomarker

Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DCnnThe phosphatidylinositol 3-kinase (PI3K) signaling pathway has been implicated in tumorigenesis. Evidence over the past years suggests a pivotal role for the catalytic subunit, PIK3CA, in human cancers. Receptor tyrosine kinases (RTKs) on the cell surface are key regulators of PI3K activity, and RTK antagonists have demonstrated significant anticancer effects in animal models and in patients. However, among tumor harboring activating mutations of PIK3CA, the therapeutic value of RTK antagonists may be compromised. The mutation status of PIK3CA may, therefore, serve as a potential biomarker for both prognosis and response to RTK inhibitors.nnSomatic mutations of the pik3ca gene occur primarily in three hot spots, E542K, E545K (exon 9) and H1047R (exon 20). These mutations are detected in a broad spectrum of cancer indications including breast, colorectal and lung. This study was designed to explore the nature of PIK3CA mutations among numerous human cancer cell lines, utilizing mostly cells with the capacity to form tumors in mice. Primers corresponding to exon 9 and 20 of human pik3ca gene were designed. Genomic DNA was harvested from over 200 lines, derived from different human cancer indications. The pik3ca locus was sequenced and its WT or mutation status was determined.nnOf these cells, 8% presented mutations equally distributed between exon 9 and 20. The frequency of PIK3CA mutations was particularly high among colon cancer lines, with fewer incidences among lung, breast and ovarian cancer lines. Interestingly, 50% of the PIK3CA mutated lines presented a concurrent mutation of either KRAS or BRAF. The capacity of PIK3CA mutation-bearing cell lines to respond to RTK blockers was then evaluated, and the mechanism contributing to such phenomenon was further discussed.nnThese results stress PI3K diagnosis value and its potential role as a valid biomarker for patient susceptibility.nnCitation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1187.