PDF file - 20K, Gene notations and ABI assay identifiers.

PDF file - 51K, Figitumumab potency in 93 cell lines annotated by normalized mRNA expression of selected IGF pathway genes.

Supplementary Fig. S2 from Antitumor activity and pharmacokinetic properties of PF-00299804, a second-generation irreversible pan-erbB receptor tyrosine kinase inhibitor

PDF file - 33K, Figitumumab sensitivity gene target correlates rank ordered by statistical significance and annotated with false discovery rates and chromosomal positions of gene locations.

PDF file - 23K, Summary of figitumumab potency in selected cell lines in vitro and tumor growth inhibition for cell lines growth as subcutaneous tumor xenografts.

Supplementary Table 1 from PF-03814735, an Orally Bioavailable Small Molecule Aurora Kinase Inhibitor for Cancer Therapy

PDF file - 312K, DLDA gene expression classifier.

PDF file - 266K, Single agent and combination activity of figitumumab and correlation of combination synergy with the genetic status of major cancer drivers or RNA expression of cancer drivers and IGF pathway alterations.

PDF file - 64K, Supplementary summary of performance of DLDA gene expression signature classifiers across cell lines including cell line classification and mean number of genes in classifier and including classifier compositions rank ordered by t value.

Abstract Figitumumab (CP-751,871), a potent and fully human monoclonal anti–insulin-like growth factor 1 receptor (IGF1R) antibody, has been investigated in clinical trials of several solid tumors. To identify biomarkers of sensitivity and resistance to figitumumab, its in vitro antiproliferative activity was analyzed in a panel of 93 cancer cell lines by combining in vitro screens with extensive molecular profiling of genomic aberrations. Overall response was bimodal and the majority of cell lines were resistant to figitumumab. Nine of 15 sensitive cell lines were derived from colon cancers. Correlations between genomic characteristics of cancer cell lines with figitumumab antiproliferative activity revealed that components of the IGF pathway, including IRS2 (insulin receptor substrate 2) and IGFBP5 (IGF-binding protein 5), played a pivotal role in determining the sensitivity of tumors to single-agent figitumumab. Tissue-specific differences among the top predictive genes highlight the need for tumor-specific patient selection strategies. For the first time, we report that alteration or expression of the MYB oncogene is associated with sensitivity to IGF1R inhibitors. MYB is dysregulated in hematologic and epithelial tumors, and IGF1R inhibition may represent a novel therapeutic opportunity. Although growth inhibitory activity with single-agent figitumumab was relatively rare, nine combinations comprising figitumumab plus chemotherapeutic agents or other targeted agents exhibited properties of synergy. Inhibitors of the ERBB family were frequently synergistic and potential biomarkers of drug synergy were identified. Several biomarkers of antiproliferative activity of figitumumab both alone and in combination with other therapies may inform the design of clinical trials evaluating IGF1R inhibitors. Mol Cancer Ther; 12(12); 2929–39. ©2013 AACR.

Abstract The PI3K pathway plays a pivotal role in many cellular functions that include regulation of cell proliferation, survival, growth, angiogenesis, and motility. Dysregulation of the PI3K/AKT pathway is a common occurrence in cancer and could be a result of mutations in PIK3CA and PTEN, as well as loss of heterozygosity of PTEN. In endometrial cancer patients, the most frequent aberration is the activation of the PI3K/mAKT pathway with; (1) 80% loss of PTEN function and 30% PIK3CA mutations in type I endometrial cancer and (2) 20% PIK3CA mutation and 46% PIK3CA amplification in type II endometrial cancer. In this study, we investigated the functional consequences of the dual PI3K/mTOR inhibitors PF-04691502 and PF-05212384 in endometrial cancer cell lines for antiproliferative effects, pathway signaling inhibition and tumor growth inhibition. Both compounds had antiproliferative activity in vitro that translated to tumor growth inhibition in vivo. Moreover, in the MFE-280 xenograft model that harbors a PIK3CA mutation (P1047R), the dual PI3K/mTOR inhibitors showed tumor regression. These studies support the use of the dual PI3K/mTOR inhibitors for endometrial cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2817. doi:1538-7445.AM2012-2817

Abstract PF-03814735 is a novel, reversible inhibitor of Aurora kinases A and B that finished a phase I clinical trial for the treatment of advanced solid tumors. To find predictive biomarkers of drug sensitivity, we screened a diverse panel of 87 cancer cell lines for growth inhibition upon PF-03814735 treatment. Small cell lung cancer (SCLC) and, to a lesser extent, colon cancer lines were very sensitive to PF-03814735. The status of the Myc gene family and retinoblastoma pathway members significantly correlated with the efficacy of PF-03814735. Whereas RB1 inactivation, intact CDKN2A/p16, and normal CCND1/Cyclin D1 status are hallmarks of SCLC, activation or amplification of any of the three Myc genes (MYC, MYCL1, and MYCN) clearly differentiated cell line sensitivity within the SCLC panel. By contrast, we found that expression of Aurora A and B were weak predictors of response. We observed a decrease in histone H3 phosphorylation and polyploidization of sensitive lines, consistent with the phenotype of Aurora B inhibition. In vivo experiments with two SCLC xenograft models confirmed the sensitivity of Myc gene-driven models to PF-03814735 and a possible schedule dependence of MYC/c-Myc–driven tumors. Altogether our results suggest that SCLC and other malignancies driven by the Myc family genes may be suitable indications for treatment by Aurora B kinase inhibitors. Mol Cancer Ther; 11(3); 710–9. ©2012 AACR.

Abstract The Aurora family of highly related serine/threonine kinases plays a key role in the regulation of mitosis. Aurora1 and Aurora2 play important but distinct roles in the G2 and M phases of the cell cycle and are essential for proper chromosome segregation and cell division. Overexpression and amplification of Aurora2 have been reported in different tumor types, including breast, colon, pancreatic, ovarian, and gastric cancer. PF-03814735 is a novel, potent, orally bioavailable, reversible inhibitor of both Aurora1 and Aurora2 kinases that is currently in phase I clinical trials for the treatment of advanced solid tumors. In intact cells, the inhibitory activity of PF-03814735 on the Aurora1 and Aurora2 kinases reduces levels of phospho-Aurora1, phosphohistone H3, and phospho-Aurora2. PF-03814735 produces a block in cytokinesis, resulting in inhibition of cell proliferation and the formation of polyploid multinucleated cells. Although PF-03814735 produces significant inhibition of several other protein kinases, the predominant biochemical effects in cellular assays are consistent with inhibition of Aurora kinases. Once-daily oral administration of PF-03814735 to mice bearing human xenograft tumors produces a reduction in phosphohistone H3 in tumors at doses that are tolerable and that result in significant inhibition of tumor growth. The combination of PF-03814735 and docetaxel in xenograft mouse tumor models shows additive tumor growth inhibition. These results support the clinical evaluation of PF-03814735 in cancer patients. Mol Cancer Ther; 9(4); 883–94. ©2010 AACR.

Abstract Targeting the insulin like growth factor receptor (IGF1R) signaling pathway is a promising approach against a variety of cancers and several clinical trials are now underway designed to evaluate the efficacy of IGF1R inhibitors singly or in combination with chemotherapeutic agents. Identification of the optimal patient population likely to respond is critical to the success of this class of targeted agents. To identify potentially predictive biomarkers, we sought to correlate response to figitumumab (CP-751,871), a fully human monoclonal antibody against IGF1R with molecular characteristics across a panel of cell lines. Growth inhibition to CP-751,871 was assessed in a panel of non-small cell lung carcinoma (NSCLC) (n=27) and colon cancer cell lines (n=21) cultured in anchorage and anchorage-independent conditions. Mutation status, whole-genome mRNA expression and DNA copy number were correlated to drug sensitivity. In addition, RNA expression of key genes in relevant signaling pathways was also interrogated via real time PCR. In the present studies, the frequency of response was greater in colon cancer cell lines than in NSCLC. Tissue or cell origin-specific expression levels of insulin-like growth factor binding proteins (IGFBPs) were associated with sensitivity. Low IGFBP3 expression [p=0.0035] correlated with response in NSCLC cell lines whereas low IGFBP6 [p=0.013] and IGF2BP3 [p=0.038] expression correlated with response in colon cancer lines. Increased IGF1R mRNA [p=0.045] levels, microsatellite stability, and PIK3CA wild-type status were also correlated with sensitivity in colon cancer lines. In addition, high expression of erbB2 [p=0.04] and erbB3 [p=0.04] were also associated with sensitivity of colon cancer lines. Collectively, findings from these studies are of potential value to identify patient populations with increased probability of benefit from IGF1R targeted agents. Our results further indicate that molecular predictors associated with the IGF1R pathway are potentially distinct between lung and colon cancer lines and additional studies to extend to other tumor types are warranted. Citation 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 LB-220.

Abstract The PI3K pathway plays a role in key cellular functions that include regulation of cell growth, proliferation, survival, angiogenesis, and motility. PI3K/AKT pathway aberrations are common in cancer comprising PTEN loss of function through mutations, deletion, and methylation events and gain of function at the PIK3CA locus including mutations and amplification events resulting in deregulation of this pathway. Thus, pharmacological intervention of this pathway should impact cellular functions central to survival of cancer cells. PIK3CA mutations/amplification and PTEN loss of heterozygosity have been reported to occur in approximately 35-40% and 20% of breast cancer, respectively and suggest this patient population could benefit from treatment using a small molecule inhibitor that targets the PI3K pathway. PF-04691502 which is a potent inhibitor of all PI3K isoforms and mTOR (TORC1 and TORC2) presently in Phase 1 trials was evaluated for its growth inhibitory or cytoreductive activity over a panel of 30 breast cancer cell lines. PF-04691502 had robust <300nM (IC50%) antiproliferative activity across the panel of breast cell lines. We observed a trend whereby cell lines that were hormone receptor (ER and/or PR) and HER2 negative (triple negative) were least sensitive and cell lines that were HER2(+) and/or hormone receptor positive (ER and/or PR) were more sensitive to PF-04691502. These studies were extended in vivo with PF-04691502 as a single agent and in combination in a HER2, ER and PR positive xenograft model with robust tumor growth inhibition in combination with docetaxel or with the pan-HER inhibitor, PF-00299804. In a xenograft that was triple negative, administration of single agent PF-4691502 at the MTD resulted in 60% tumor growth inhibition. In conclusion, our findings suggest that the ER/PR and/or HER2 positive breast cancer segment may particularly benefit from PI3K/mTOR inhibitor based treatment regimens. Citation 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 LB-302.

Abstract The identification of patient populations predicted to derive clinical benefit from systemic treatment regimens is of critical importance for development of targeted therapies. Collective clinical data available for the Aurora kinase inhibitor class of targeted therapies has indicated that broad single agent clinical activity is not readily apparent. We screened a panel of established lung (mostly non-small cell) and colon cell lines for growth inhibitory sensitivity to PF-03814735, an Aurora family kinase inhibitor, revealing a potential correlation with Myc family amplification or expression. The Aurora kinases A and B have been shown to be functionally linked with Myc family oncoproteins in a number of tumor types. Myc family gene amplification events have been reported in about 30% of small cell lung cancer (SCLC) primary tumors and around 50% of cell lines established from SCLC. We next screened a panel of around 20 SCLC lines for sensitivity to PF-03814735 and its relationship to Myc family gene copy number and gene expression levels. Sensitivity to PF-03814735 in vitro was strongly correlated with amplification events in at least one of the Myc family genes (c-Myc, L-Myc, N-Myc) as well as mRNA expression levels of those genes. Myc family expression demonstrated a significant correlation with gene copy number. Follow up studies to evaluate antitumor efficacy in two SCLC xenograft models, H82 (c-Myc) and H69 (N-Myc), indicated significant tumor growth inhibition by PF-03814735. However, the SCLC models were not appreciably more sensitive to PF-03814735 than several non-Myc amplified tumors studied previously (HCT-116, COLO-205, MDA-MB-231) indicating that further study of optimal dose schedule and/or the molecular basis of sensitivity is warranted. Citation 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 2615.

Abstract Anti angiogenic agents that target the VEGF pathway (bevacizumab, sorafenib and sunitinib (SU)) have demonstrated improved clinical benefit as single agents or in combination with chemotherapy in a variety of solid tumor types. Although this class of agents has provided an improved treatment option to certain tumor types, it is evident that some patients do not respond to therapy or will relapse despite an initial response to treatment due to an acquired resistance. To better understand this mode of acquired resistance to this class of agents, a model of Renal Cell Carcinoma (RCC) which has shown regression, eventually developed evasive resistance during long term administration of the single agent sunitinib (a small-molecule inhibitor of the VEGF-1, 2, 3, PDGF-α,β, KIT, CSF-1R and FLT-3 receptor tyrosine kinases). Despite the observation that resistant tumors under continuous treatment with sunitinib (60 mg/kg PO, QD) demonstrated a gross phenotypic reduction of micro-vascular density (MVD), they continued to show a progressive increase in size. Because the observed low MVD phenotype was suggestive of an ability of resistant tumor cells to survive in an increasingly hypoxic environment with a reduced functional vasculature, emphasis was placed on identifying tumor cell derived resistance factors. Gene expression and/or proteomic profiling studies accompanied by confirmatory immunoblotting indicated a subset of differentially expressed genes and/or proteins in resistant vs. sensitive treated tumors. Although profiling studies did not clearly elucidate definitive resistance mechanisms, the dysregulation of EGFR, FGFR1, & TGFB1 pathways and downstream signaling through PI3K and MEK were potentially implicated. Comparative genomic hybridization studies did not identify any resistance mechanisms at the cytogenetic level. Sunitinib combination studies designed to target pathways potentially involved in resistance (SU + a PI3K/mTOR inhibitor; SU + a MEK inhibitor) demonstrated the ability to restore sensitivity in the sunitinib resistance model. The ability to identify and characterize resistance mechanisms and circumvent resistance through combination approaches may have utility in developing future combination regimens for anti angiogenic therapies. Citation 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 387.

Abstract Signaling through the erbB receptor family of tyrosine kinases contributes to the proliferation, differentiation, migration, and survival of a variety of cell types. Abnormalities in members of this receptor family have been shown to play a role in oncogenesis, thus making them attractive targets for anticancer treatments. PF-00299804 is a second-generation irreversible pan-erbB receptor tyrosine kinase inhibitor currently in phase I clinical trials. PF-00299804 is believed to irreversibly inhibit erbB tyrosine kinase activity through binding at the ATP site and covalent modification of nucleophilic cysteine residues in the catalytic domains of erbB family members. Oral administration of PF-00299804 causes significant antitumor activity, including marked tumor regressions in a variety of human tumor xenograft models that express and/or overexpress erbB family members or contain the double mutation (L858R/T790M) in erbB1 (EGFR) associated with resistance to gefitinib and erlotinib. Furthermore, PF-00299804 shows exceptional distribution to human tumor xenografts and excellent pharmacokinetic properties across species. [Mol Cancer Ther 2008;7(7):1880–9]