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Supplemental Tables 1-2 & Figures 1-4 from Intrinsic Resistance to Cixutumumab Is Conferred by Distinct Isoforms of the Insulin Receptor

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posted on 2023-04-03, 16:28 authored by Amelie Forest, Michael Amatulli, Dale L. Ludwig, Christopher B. Damoci, Ying Wang, Colleen A. Burns, Gregory P. Donoho, Nina Zanella, Heinz H. Fiebig, Marie C. Prewett, David Surguladze, James T. DeLigio, Peter J. Houghton, Malcolm A. Smith, Ruslan Novosiadly

Supplemental Tables 1-2 & Figures 1-4. Supplemental Table 1. Total IR, IR-A, IR-B, IGF-IR, IGF-I and IGF-II mRNA expression in preclinical models of pediatric solid tumors. Supplemental Table 2. Patient-derived xenograft models of lung adenocarcinoma: clinical information and mutational status. Supplemental Figure 1. Effect of cixutumumab on circulating growth hormone (A), insulin (B) and IGF-I (C) levels in male C57BL/6 mice. Represented are mean values +/- SEM. p-values {less than or equal to} 0.05 indicate statistical significance (*) (Student's t-test). Supplemental Figure 2. IGF-I and IGF-II mRNA expression in patient-derived models of lung adenocarcinoma. Represented are mean values +/- SEM. Supplemental Figure 3. Overlay analysis of cell surface IR and IGF-IR expression by flow cytometry (A) and ligand-induced signal transduction by Western Blot (B) in A549 cell overexpressing empty vector (pLVX) or IR isoforms (IR-A and IR-B). Serum-starved cells (1h) were treated with rhIGF-I, rhIGF-II or recombinant human insulin (10 nM) for 5, 15 or 60 minutes. Proteins (10ug) extracted from cultured cells were size-fractionated by SDS-PAGE and immunoblotted with anti-phospho-IRbY1150/51/IGF-IRbY1135/36, anti-phospho-AktS473 and anti-phospho-p42/p44 MAPKT202/Y204 antibodies. Total level of proteins was demonstrated by immunoblotting with antibodies directed against total Akt and p42/p44 MAPK. Supplemental Figure 4. Anchorage-dependent cell viability assay. MCF-7-Mock, MCF-7-IR-A and MCF-7-IRB cells were treated with increasing concentrations of cixutumumab (0.015-1,000 nM). Maximum inhibition for MCF-7-Mock, MCF-7-IR-A and MCF-7-IRB were 60.4%, 0% and 26.9% respectively, p-values < 0.01(Student's t-test). Represented are mean values +/- SEM.

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ARTICLE ABSTRACT

Despite a recent shift away from anti–insulin-like growth factor I receptor (IGF-IR) therapy, this target has been identified as a key player in the resistance mechanisms to various conventional and targeted agents, emphasizing its value as a therapy, provided that it is used in the right patient population. Molecular markers predictive of antitumor activity of IGF-IR inhibitors remain largely unidentified. The aim of this study is to evaluate the impact of insulin receptor (IR) isoforms on the antitumor efficacy of cixutumumab, a humanized mAb against IGF-IR, and to correlate their expression with therapeutic outcome. The data demonstrate that expression of total IR rather than individual IR isoforms inversely correlates with single-agent cixutumumab efficacy in pediatric solid tumor models in vivo. Total IR, IR-A, and IR-B expression adversely affects the outcome of cixutumumab in combination with chemotherapy in patient-derived xenograft models of lung adenocarcinoma. IR-A overexpression in tumor cells confers complete resistance to cixutumumab in vitro and in vivo, whereas IR-B results in a partial resistance. Resistance in IR-B–overexpressing cells is fully reversed by anti–IGF-II antibodies, suggesting that IGF-II is a driver of cixutumumab resistance in this setting. The present study links IR isoforms, IGF-II, and cixutumumab efficacy mechanistically and identifies total IR as a biomarker predictive of intrinsic resistance to anti–IGF-IR antibody.Implications: This study identifies total IR as a biomarker predictive of primary resistance to IGF-IR antibodies and provides a rationale for new clinical trials enriched for patients whose tumors display low IR expression. Mol Cancer Res; 13(12); 1615–26. ©2015 AACR.

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