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Background: Ipat is a potent oral AKT inhibitor that has been studied in multiple clinical trials, primarily in breast and prostate cancers. Combining fulv and AKT inhibition demonstrated efficacy in pts with HR+ aBC regardless of PI3K/AKT pathway alterations [Jones, Lancet Oncol 2020]. IPATunity150 (NCT04060862) was designed as a phase III trial with an open-label phase Ib portion adding ipat to palbo plus fulv in biomarker-unselected HR+ HER2-negative aBC. The biological rationale was to prevent or delay resistance to CDK4/6 inhibition plus endocrine therapy (ET). AKT1 alterations and PTEN loss have been implicated in resistance to CDK4/6 inhibitors [Wander, Cancer Discov 2020; Costa, Cancer Discov 2020]. We report results from the open-label phase Ib portion. Patients and Methods: The primary objective was to assess safety and pharmacokinetics (PK) of ipat in combination with palbo and fulv; several efficacy parameters were also analyzed. Pts with measurable disease who had not previously received a CDK4/6 inhibitor and had experienced relapse during adjuvant ET were treated with ipat at a dose of 300 mg/d, d1-21 q28d, plus standard-of-care doses of palbo (125 mg/d, d1-21 q28d) plus fulv (500 mg q28d with a loading dose in cycle 1). The selected ipat dose was lower than the 400 mg typically used in other studies because of the anticipated drug-drug interaction (DDI) when combining ipat (a sensitive CYP3A4 substrate and mild-to-moderate CYP3A inhibitor) with palbo (a weak time-dependent CYP3A4 inhibitor and CYP3A substrate). Results: Of the 20 pts treated, 20% were Asian, 65% had primary endocrine resistance (relapse ≤2 years after starting adjuvant ET), 80% had received prior (neo)adjuvant chemotherapy, and 60% had liver and/or lung metastases. At the data cutoff (19 Mar 2021; median follow-up 6.1 months), median treatment duration was 5.1, 5.9, and 5.3 months for ipat, palbo, and fulv, respectively. Treatment was ongoing in 13 pts. Grade 3/4 adverse events (AEs) occurred in 80% of pts (no grade 5). The most common AEs were diarrhea (80% any grade, 10% grade 3, no grade 4) and neutropenia (75% any grade, 45% grade 3, 20% grade 4). Other notable grade ≥3 AEs were grade 3 liver function test elevations in 10%. There were no cases of febrile neutropenia and only 1 case of pneumonitis (grade 1). AEs led to at least one dose reduction of ipat in 6 pts (30%; diarrhea n=3 [with vomiting in 1 pt], neutropenia n=3 [with fatigue in 1 pt]) and of palbo in 9 pts (45%; all for neutropenia). One pt (5%) discontinued ipat and palbo permanently due to ongoing neutropenia after protocol-defined dose reductions. As expected, a DDI led to increased ipat exposure (AUC0-24,ss ~60% and Cmax ~40%) when ipat and palbo were combined. Based on population PK analysis, palbo AUC0-24,ss was ~30% higher than reported from the PALOMA 1 and 2 trials, which was expected and consistent with previously reported physiologically based PK modeling of palbo exposure when administered with moderate CYP3A inhibitors [Yu, J Clin Pharmacol 2017]. All 20 pts had at least one post-baseline tumor assessment. Best overall response rate was 45% (95% CI: 23-68%), including confirmed responses in 7 pts (35%; 5% complete response, 30% partial response) at the clinical cutoff date. Median duration of response was 9.6 months (95% CI: 7.1-not estimable). An additional 10 pts (50%) had stable disease. Progression-free survival results were immature (events in 7 pts). There was no obvious association between efficacy and mutations in PIK3CA/AKT1 as tested in ctDNA. Conclusion: The triplet combination of ipat, fulv, and palbo had an acceptable safety profile generally consistent with that of the individual study drugs; ipat exposure was increased through a predicted DDI. Updated results will be presented. Citation Format: Mafalda Oliveira, Aditya Bardia, Sung-Bae Kim, Naoki Niikura, Cristina Hernando, Gustavo Werutsky, Yoland Antill, Pedro Liedke, Catherine Oakman, Eriko Tokunaga, Seth Wander, Vanessa Krause, Toshinari Yamashita, Frauke Schimmoller, Jacob Rotmensch, Heidi Savage, Rucha Sane, Nicholas Turner. Ipatasertib (ipat) in combination with palbociclib (palbo) and fulvestrant (fulv) in patients (pts) with hormone receptor-positive (HR+) HER2-negative advanced breast cancer (aBC) [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-16-11.

Supplementary Fig. from Proteomic analysis of breast cancer molecular subtypes and biomarkers of response to targeted kinase inhibitors using reverse-phase protein microarrays

Commentary on this Article from Molecular predictors of response to a humanized anti–insulin-like growth factor-I receptor monoclonal antibody in breast and colorectal cancer

The insulin-like growth factor-I receptor (IGF-IR) pathway is required for the maintenance of the transformed phenotype in neoplastic cells and hence has been the subject of intensive drug discovery efforts. A key aspect of successful clinical development of targeted therapies directed against IGF-IR will be identification of responsive patient populations. Toward that end, we have endeavored to identify predictive biomarkers of response to an anti-IGF-IR-targeting monoclonal antibody in preclinical models of breast and colorectal cancer. We find that levels of the IGF-IR itself may have predictive value in these tumor types and identify other gene expression predictors of in vitro response. Studies in breast cancer models suggest that IGF-IR expression is both correlated and functionally linked with estrogen receptor signaling and provide a basis for both patient stratification and rational combination therapy with antiestrogen-targeting agents. In addition, we find that levels of other components of the signaling pathway such as the adaptor proteins IRS1 and IRS2, as well as the ligand IGF-II, have predictive value and report on the development of a pathway-focused panel of diagnostic biomarkers that could be used to test these hypotheses during clinical development of IGF-IR-targeting therapies. [Mol Cancer Ther 2009;8(8):2110–21]

Supplementary Table S3 from Predictive Biomarkers of Sensitivity to the Phosphatidylinositol 3′ Kinase Inhibitor GDC-0941 in Breast Cancer Preclinical Models

Although breast cancer molecular subtypes have been extensively defined by means of gene expression profiling over the past decade, little is known, at the proteomic level, as to how signaling pathways are differentially activated and serve to control proliferation in different breast cancer subtypes. We used reverse-phase protein arrays to examine phosphorylation status of 100 proteins in a panel of 30 breast cancer cell lines and showed distinct pathway activation differences between different subtypes that are not obvious from previous gene expression studies. We also show that basal levels of phosphorylation of key signaling nodes may have diagnostic utility in predicting response to selective inhibitors of phosphatidylinositol 3-kinase and mitogen-activated protein kinase/extracellular signal-regulated kinase kinase. Finally, we show that reverse-phase protein arrays allow the parallel analysis of multiple pharmacodynamic biomarkers of response to targeted kinase inhibitors and that inhibitors of epidermal growth factor receptor and mitogen-activated protein kinase/extracellular signal-regulated kinase kinase result in compensatory up-regulation of the phosphatidylinositol 3-kinase/Akt signaling pathway. [Mol Cancer Ther 2008;7(12):3695–706]

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Purpose: We describe the preclinical pharmacology and antitumor activity of GDC-0068, a novel highly selective ATP-competitive pan-Akt inhibitor currently in clinical trials for the treatment of human cancers.Experimental Design: The effect of GDC-0068 on Akt signaling was characterized using specific biomarkers of the Akt pathway, and response to GDC-0068 was evaluated in human cancer cell lines and xenograft models with various genetic backgrounds, either as a single agent or in combination with chemotherapeutic agents.Results: GDC-0068 blocked Akt signaling both in cultured human cancer cell lines and in tumor xenograft models as evidenced by dose-dependent decrease in phosphorylation of downstream targets. Inhibition of Akt activity by GDC-0068 resulted in blockade of cell-cycle progression and reduced viability of cancer cell lines. Markers of Akt activation, including high-basal phospho-Akt levels, PTEN loss, and PIK3CA kinase domain mutations, correlate with sensitivity to GDC-0068. Isogenic PTEN knockout also sensitized MCF10A cells to GDC-0068. In multiple tumor xenograft models, oral administration of GDC-0068 resulted in antitumor activity ranging from tumor growth delay to regression. Consistent with the role of Akt in a survival pathway, GDC-0068 also enhanced antitumor activity of classic chemotherapeutic agents.Conclusions: GDC-0068 is a highly selective, orally bioavailable Akt kinase inhibitor that shows pharmacodynamic inhibition of Akt signaling and robust antitumor activity in human cancer cells in vitro and in vivo. Our preclinical data provide a strong mechanistic rationale to evaluate GDC-0068 in cancers with activated Akt signaling. Clin Cancer Res; 19(7); 1760–72. ©2012 AACR.

Supplementary Table from Proteomic analysis of breast cancer molecular subtypes and biomarkers of response to targeted kinase inhibitors using reverse-phase protein microarrays

Supplementary Figures 1-7 from Molecular predictors of response to a humanized anti–insulin-like growth factor-I receptor monoclonal antibody in breast and colorectal cancer

Purpose: The class I phosphatidylinositol 3′ kinase (PI3K) plays a major role in proliferation and survival in a wide variety of human cancers. A key factor in successful development of drugs targeting this pathway is likely to be the identification of responsive patient populations with predictive diagnostic biomarkers. This study sought to identify candidate biomarkers of response to the selective PI3K inhibitor GDC-0941.Experimental Design: We used a large panel of breast cancer cell lines and in vivo xenograft models to identify candidate predictive biomarkers for a selective inhibitor of class I PI3K that is currently in clinical development. The approach involved pharmacogenomic profiling as well as analysis of gene expression data sets from cells profiled at baseline or after GDC-0941 treatment.Results: We found that models harboring mutations in PIK3CA, amplification of human epidermal growth factor receptor 2, or dual alterations in two pathway components were exquisitely sensitive to the antitumor effects of GDC-0941. We found that several models that do not harbor these alterations also showed sensitivity, suggesting a need for additional diagnostic markers. Gene expression studies identified a collection of genes whose expression was associated with in vitro sensitivity to GDC-0941, and expression of a subset of these genes was found to be intimately linked to signaling through the pathway.Conclusion: Pathway focused biomarkers and the gene expression signature described in this study may have utility in the identification of patients likely to benefit from therapy with a selective PI3K inhibitor. Clin Cancer Res; 16(14); 3670–83. ©2010 AACR.

PDF file - 954K, Supplemental Figures: Figure S1. Chemical structure of the Akt inhibitor GDC-0068. Figure S2. Effect of GDC-0068 on cell cycle and apoptotic response on PC-3 and MCF7-neo/HER2 cell lines. Figure S3. Western blot analysis of indicated proteins in isogenic MCF10A cells with and without PTEN knockout in the presence of 20 or 0.2 ng/ml EGF. Figure S4. Representative immunohistochemistry staining images of cleaved caspase-3 in TOV-21G.x1 tumors. Figure S5. Combination effects between GDC-0068 and chemotherapeutic agents in vitro.

Purpose: The pathways underlying basal-like breast cancer are poorly understood, and as yet, there is no approved targeted therapy for this disease. We investigated the role of mitogen-activated protein kinase kinase (MEK) and phosphatidylinositol 3-kinase (PI3K) inhibitors as targeted therapies for basal-like breast cancer.Experimental Design: We used pharmacogenomic analysis of a large panel of breast cancer cell lines with detailed accompanying molecular information to identify molecular predictors of response to a potent and selective inhibitor of MEK and also to define molecular mechanisms underlying combined MEK and PI3K targeting in basal-like breast cancer. Hypotheses were confirmed by testing in multiple tumor xenograft models.Results: We found that basal-like breast cancer models have an activated RAS-like transcriptional program and show greater sensitivity to a selective inhibitor of MEK compared with models representative of other breast cancer subtypes. We also showed that loss of PTEN is a negative predictor of response to MEK inhibition, that treatment with a selective MEK inhibitor caused up-regulation of PI3K pathway signaling, and that dual blockade of both PI3K and MEK/extracellular signal–regulated kinase signaling synergized to potently impair the growth of basal-like breast cancer models in vitro and in vivo.Conclusions: Our studies suggest that single-agent MEK inhibition is a promising therapeutic modality for basal-like breast cancers with intact PTEN, and also provide a basis for rational combination of MEK and PI3K inhibitors in basal-like cancers with both intact and deleted PTEN.

Supplementary Data from In vivo Antitumor Activity of MEK and Phosphatidylinositol 3-Kinase Inhibitors in Basal-Like Breast Cancer Models

XLSX file - 74K, Table S1. Enzymatic potency, selectivity and cellular potency of GDC-0068 Table S2. GDC-0068 IC50 profile on cell viability and genetic background of cancer cell lines Table S3. Maximum percent body weight changes of mice treated with GDC-0068 single agent and in combination with chemotherapeutic agents

CCR Translation for this Article from Predictive Biomarkers of Sensitivity to the Phosphatidylinositol 3′ Kinase Inhibitor GDC-0941 in Breast Cancer Preclinical Models

Supplementary Figures from Predictive Biomarkers of Sensitivity to the Phosphatidylinositol 3′ Kinase Inhibitor GDC-0941 in Breast Cancer Preclinical Models

Purpose:Somatic mutations in phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA), which encodes the p110α catalytic subunit of PI3K, are found in multiple human cancers. While recurrent mutations in PIK3CA helical, regulatory, and kinase domains lead to constitutive PI3K pathway activation, other mutations remain uncharacterized. To further evaluate their clinical actionability, we designed a basket study for patients with PIK3CA-mutant cancers with the isoform-specific PI3K inhibitor taselisib.Patients and Methods:Patients were enrolled on the basis of local PIK3CA mutation testing into one of 11 histology-specific cohorts and treated with taselisib at 6 or 4 mg daily until progression. Tumor DNA from baseline and progression (when available) was sequenced using a next-generation sequencing panel. Exploratory analyses correlating genomic alterations with treatment outcomes were performed.Results:A total of 166 patients with PIK3CA-mutant cancers were enrolled. The confirmed response rate was 9%. Activity varied by tumor type and mutant allele, with confirmed responses observed in head and neck squamous (15.4%), cervical (10%), and other cancers, plus in tumors containing helical domain mutations. Genomic analyses identified mutations potentially associated with resistance to PI3K inhibition upfront (TP53 and PTEN) and postprogression through reactivation of the PI3K pathway (PTEN, STK11, and PIK3R1). Higher rates of dose modification occurred at higher doses of taselisib, indicating a narrow therapeutic index.Conclusions:Taselisib had limited activity in the tumor types tested and is no longer in development. This genome-driven study improves understanding of the activity, limitations, and resistance mechanisms of using PI3K inhibitors as monotherapy to target PIK3CA-mutant tumors.

Many, if not most philosophers, deny that a sentence like ‘Sherlock Holmes smokes’ is true. However, this attitude confl icts with speakers’ assignment of the value true to this sentence. Furthermore, making these assignments seem in no way distinct from the process that leads speakers to assign true to other sentences, sentences like ‘Bertrand Russell smokes.’ I will explore the idea that when speakers assign the value true to the first sentence, they are not making any kind of confused mistake — that we ought to take these assignments at face value. I show how the alternative view is inadequate for explaining various examples of fi ctional discourse. In addition, evidence that these truth value assignments to sentences are tracking semantic content, rather than pragmatic effects, is offered.

The identification of early breast cancer patients who may benefit from adjuvant chemotherapy has evolved to include assessment of clinicopathologic features such as tumor size and nodal status, as well as several gene-expression profiles for ER-positive, HER2-negative cancers. However, these tools do not reliably identify patients at the greatest risk of recurrence. The mutation and copy-number landscape of triple-negative breast cancer (TNBC) subtypes defined by gene expression is also largely unknown, and elucidation of this landscape may shed light on novel therapeutic opportunities. The USO01062 phase III clinical trial of standard chemotherapy (with or without capecitabine) enrolled a cohort of putatively high-risk patients based on clinical features, yet only observed a 5-year disease-free survival event rate of 11.6%. In order to uncover genomic aberrations associated with recurrence, a targeted next-generation sequencing panel was used to compare tumor specimens from patients who had a recurrence event with a matched set who did not. The somatic mutation and copy-number alteration landscapes of high-risk early breast cancer patients were characterized and alterations associated with relapse were identified. Tumor mutational burden was evaluated but was not prognostic in this study, nor did it correlate with PDL1 or CD8 gene expression. However, TNBC subtypes had substantial genomic heterogeneity with a distinct pattern of genomic alterations and putative underlying driver mutations. Implications: The present study uncovers a compendium of genomic alterations with utility to more precisely identify high-risk patients for adjuvant trials of novel therapeutic agents.

Background The PI3K/AKT signaling pathway plays a significant role in both HR+ BC and TNBC. IPATunity130 is a double-blind, placebo-controlled, randomized phase 3 trial of ipatasertib in combination with paclitaxel in patients with PI3K/AKT1/PTEN-altered HR+ or TNBC. A next-generation sequencing (NGS)-based assay from Foundation Medicine Inc. (FMI) was used to select patients prospectively for enrollment in this trial.Patients and methods An investigational clinical trial assay (CTA) of a composite 3-gene biomarker signature [Kim, Lancet Oncol 2017] based on the FoundationOne® CDx assay was used to identify patients with PI3K/AKT pathway-activated tumors as eligible for enrollment in IPATunity130 (NCT03337724). Qualifying alterations for the 3-gene signature (referred to as ‘biomarker-positive’) comprised activating mutations in PIK3CA and/or AKT1, and/or loss of function (LOF) alterations in PTEN represented by homozygous or heterozygous deletions, dominant-negative mutations, or inactivating mutations under loss of heterozygosity. Study sites were required to submit formalin-fixed paraffin-embedded archival or fresh biopsy tissue derived from primary or metastatic tumors for patient screening. IPATunity130 includes three independent cohorts: Cohort A (biomarker-positive TNBC); Cohort B (biomarker-positive HR+/HER2– BC); and Cohort C (biomarker-negative TNBC). Results In total, 1736 patients were screened, from whom 1690 samples were tested by FMI. Of these, 1475 (87%) produced a valid NGS result. The remaining 215 (13%) failed quality control for reasons including insufficient tissue, DNA yield, lab error, and computational failure. Alteration status for PIK3CA and/or AKT1 and/or PTEN was positive in 703 (49%) of 1440 CTA-evaluable samples. In the HR+/HER2– cohort, the breakdown of CTA-positive samples was 301/356 (85%) PIK3CA/AKT1 mutations and 86/356 (24%) PTEN LOF alterations. In TNBC, 183/347 (53%) had PIK3CA/AKT1 mutations and 193/347 (56%) had PTEN LOF alterations. CTA results according to baseline characteristics are shown overall and by subtype below. The most common mutations detected outside the 3-gene biomarker signature in the screened population were in the TP53, BRCA1, RB1, BRCA2, and NF1 genes in the TNBC cohort and the TP53, GATA3, CDH1, MAP3K1, and ESR1 genes in the HR+/HER2– cohort. In the HR+/HER2– cohort, 30 (14%) of 213 metastatic tumors had ESR1 mutations compared with 10/414 (2%) primary tumors. Tumor BRCA1 mutations were more common in patients aged ≤50 years whereas MAP3K1 mutations were more common in those aged >50 years.Conclusions IPATunity130 is the first reported pivotal trial to utilize the 3-gene biomarker CTA in HR+/HER2– BC and TNBC to screen for patients with PI3K/AKT pathway-activated tumors. The 3-gene biomarker CTA detected alterations in 49% of screened patients, with a higher prevalence of PIK3CA/AKT1/PTEN alterations in HR+/HER2– BC than TNBC. Additional analyses are planned to assess correlations between clinical outcomes and tumor characteristics. SubgroupPIK3CA/AKT1/PTEN alteration, n/N (%)OverallHR+/HER2–TNBCAll patient samples703/1440 (49)356/647 (55)347/793 (44)Age, years≤50219/475 (46)99/195 (51)120/280 (43)>50484/965 (50)257/452 (57)227/513 (44)Sample sourcePrimary456/941 (48)232/414 (56)224/527 (43)Metastatic222/448 (50)112/213 (53)110/235 (47)Geographic regionNorth America60/102 (59)33/49 (67)27/53 (51)Asia-Pacific172/330 (52)81/145 (56)91/185 (49)Europe301/633 (48)171/310 (55)130/323 (40)Rest of world170/375 (45)71/143 (50)99/232 (43) Citation Format: Heidi Savage, Wendy Lin, Mafalda Oliveira, Carlos Barrios, Joyce O’Shaughnessy, Nicholas Turner, Rebecca Dent, Steven J Isakoff, Shigehira Saji, Qinshu Lian, Denise Bradley, Sarah-Jayne Reilly, Heather Hinton, Matthew J Wongchenko, Aruna Mani, Sung-Bae Kim. Prospective testing for PIK3CA/AKT1/PTEN alterations in tumor tissue from 1440 patients with advanced hormone receptor-positive HER2-negative breast cancer (HR+/HER2- BC) or triple-negative breast cancer (TNBC) screened for the IPATunity130 randomized phase 3 trial [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS5-06.

Taselisib is a potent and selective tumor growth inhibitor through PI3K pathway suppression. Thirty-four patients with locally advanced or metastatic solid tumors were treated (phase I study, modified 3+3 dose escalation; 5 cohorts; 3–16 mg taselisib once-daily capsule). Taselisib pharmacokinetics were dose-proportional; mean half-life was 40 hours. Frequent dose-dependent, treatment-related adverse events included diarrhea, hyperglycemia, decreased appetite, nausea, rash, stomatitis, and vomiting. At 12 and 16 mg dose levels, dose-limiting toxicities (DLT) were observed, with an accumulation of higher-grade adverse events after the cycle 1 DLT assessment window. Pharmacodynamic findings showed pathway inhibition at ≥3 mg in patient tumor samples, consistent with preclinical PIK3CA-mutant tumor xenograft models. Confirmed response rate was 36% for PIK3CA-mutant tumor patients with measurable disease [5/14: 4 breast cancer (3 patients at 12 mg); 1 non–small cell lung cancer], where responses started at 3 mg, and 0% in patients with tumors without known PIK3CA hotspot mutations (0/15).Significance: Preliminary data consistent with preclinical data indicate increased antitumor activity of taselisib in patients with PIK3CA-mutant tumors (in comparison with patients with tumors without known activating PIK3CA hotspot mutations) starting at the lowest dose tested of 3 mg, thereby supporting higher potency for taselisib against PIK3CA-mutant tumors. Cancer Discov; 7(7); 704–15. ©2017 AACR.See related commentary by Rodon and Tabernero, p. 666.This article is highlighted in the In This Issue feature, p. 653

Background: The phosphatidylinositol 3-kinase (PI3K) pathway is frequently dysregulated in hormone receptor (HR)-positive breast cancer (BC), with activating mutations of PIK3CA detected in ~35–45% of patients (pts). Acquired mutations in the ESR1 gene, which encodes estrogen receptor α, may be associated with resistance to aromatase inhibitor (AI) therapy. Taselisib is a potent and selective PI3K inhibitor, with greater selectivity against mutant PI3Kα isoforms than wild-type (WT) via a unique mechanism. In phase I studies, taselisib plus fulvestrant had clinical activity and manageable tolerability in pts with HR-positive BC. We report exploratory analyses of PIK3CA and ESR1 from circulating tumor DNA (ctDNA). Methods: In this phase II, open-label, single-arm study (PMT4979g; NCT01296555), pts were postmenopausal with HER2-negative, HR-positive locally advanced or metastatic BC and progression or non-response to ≥1 prior endocrine therapy in the adjuvant or metastatic setting. Pts received taselisib (6 mg capsule orally, daily) plus fulvestrant (500 mg intramuscular on Days 1 and 15 of Cycle 1, then Day 1 of each 28-day cycle) until disease progression or unacceptable toxicity. PIK3CA-mutation testing on archival tumor tissue used the cobas® PIK3CA Mutation Test. The Sysmex Inostics' BEAMing Digital PCR platform was used for ctDNA analysis of ESR1 and PIK3CA mutations (pre-dose on Cycle 1, Day 1). Primary endpoints were objective response rate (ORR) and clinical benefit rate (CBR) in all pts and those with PIK3CA mutations. ORR was confirmed complete response (cCR) and confirmed partial response (cPR). CBR was cCR, cPR, or stable disease for ≥6 months. Secondary endpoints included safety, efficacy, pharmacokinetics, and exploratory biomarker analysis. Results: 60 pts were enrolled. Median age was 61.5 years (range 31–82). In the metastatic setting, pts had received prior chemotherapy (21.7%) and prior hormonal therapy (50.0%). 86.7% of pts had received prior treatment with an AI. 45 pts had PIK3CA mutation status from archival tumor tissue and ctDNA testing; concordance was 86.7% (39/45). ctDNA analysis, vs archival tumor tissue testing, identified 4 pts and 9 pts with PIK3CA mutations from pts with WT and unknown PIK3CA mutation status, respectively. Based on ctDNA analysis (N=60), 13 pts (21.7%) had mutations in both ESR1 and PIK3CA, 21 pts (35.0%) were 'mutation not detected' (MND) for both genes, 8 (13.3%) had ESR1 mutations and PIK3CA MND, and 18 (30.0%) had ESR1 MND and PIK3CA mutations. In pts with measurable disease at baseline, confirmed responses (all partial) were: PIK3CA mutation, 38.1% (8/21); PIK3CA MND, 8.7% (2/23); all pts, 22.7% (10/44). CBRs were: PIK3CA mutation, 42.9%; PIK3CA MND, 17.4%; all pts, 29.5%. ORR and CBR from ctDNA analyses were similar to archival tumor tissue data. Conclusions: ctDNA analysis identified PIK3CA mutations in pts with previously unknown or WT mutation status from archival tumor tissue; ORR and CBR were similar to those from archival tumor tissue suggesting that PIK3CA mutation testing from ctDNA may be used as a surrogate when tissue is unavailable. 21.7% of pts had mutations in both ESR1 and PIK3CA. Citation Format: Dickler MN, Saura C, Oliveira M, Richards DA, Krop IE, Cervantes A, Stout TJ, Jin H, Savage HM, Wilson TR, Baselga J. Phase II study of taselisib (GDC-0032) plus fulvestrant in HER2-negative, hormone receptor-positive advanced breast cancer: Analysis by PIK3CA and ESR1 mutation status from circulating tumor DNA [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P6-12-01.

Background There is an urgent requirement to identify biomarkers to tailor treatment in human epidermal growth factor receptor 2 (HER2)-amplified early breast cancer treated with trastuzumab/pertuzumab-based chemotherapy. Methods Among the 225 patients randomly assigned to trastuzumab/pertuzumab concurrently or sequentially with an anthracycline-containing regimen or concurrently with an anthracycline-free regimen in the Tryphaena trial, we determined the percentage of tumor-infiltrating lymphocytes (TILs) at baseline in 213 patients, of which 126 demonstrated a pathological complete response (pCR; ypT0/is ypN0), with 28 demonstrating event-free survival (EFS) events. We investigated associations between baseline TIL percentage and either pCR or EFS after adjusting for clinicopathological characteristics using logistic and Cox regression models, respectively. To understand TIL biology, we evaluated associations between baseline TILs and baseline tumor gene expression data (800 gene set by NanoString) in a subset of 173 patients. All statistical tests were two-sided. Results Among the patients with measurable TILs at baseline, the median level was 14.1% (interquartile range = 7.1%-32.4%). After adjusting for clinicopathological characteristics, baseline percentage TIL was not associated with pCR (adjusted odds ratio [aOR] for every 10-percentage unit increase in TILs = 1.12, 95% confidence interval [CI] = 0.95 to 1.31, P = .17). At a median follow-up of 4.7 years, for every increase in baseline TILs of 10%, there was a 25% reduction in the hazard for an EFS event (aOR = 0.75, 95% CI = 0.56 to 1.00, P = .05) after adjusting for baseline clinicopathological characteristics and pCR. Additionally, genes associated with epithelial-mesenchymal transition, angiogenesis, and T-cell inhibition such as SNAIL1, ZEB1, NOTCH3, and B7-H3 were statistically significantly inversely correlated with percentage TIL. Conclusions Baseline TIL percentage provides independent prognostic information in patients treated with trastuzumab/pertuzumab-based neoadjuvant chemotherapy. However, further validation is required.

Background: The OPPORTUNE Study randomized postmenopausal patients (pts) to receive 2-week preoperative treatment with anastrozole (ANA) plus pictilisib ("ANA+PIC" arm) or ANA alone. Patients had newly diagnosed, operable, ER+, HER2- invasive breast cancer of ≥1 cm size. The primary outcome at interim analysis (n=70) revealed that the addition of PIC significantly increased the anti-proliferative response to ANA as measured by reduction in Ki67 immunohistochemistry (IHC). Multivariate analyses suggested benefit of PIC for patients with luminal B disease (Schmid et al. SABCS 2014). Methods: RNA expression analysis of ∼800 breast cancer-related genes was performed on patients analyzed at the interim analysis, including 14 (ANA) and 20 (ANA+PIC) patients with matched pre- and post- treatment paired tumour samples using the nCounter platform (NanoString). Differential expression of individual genes by arm was assessed using paired and moderated t-tests and statistical significance assessed through false discovery rate (FDR). Ingenuity Pathway Analysis (IPA) of differentially expressed transcripts identified pathways of relevance. Protein expression was analyzed by reverse protein array ( RPPA) in pre- and post-treatment samples. Results: In an unsupervised analysis, down-regulation of genes associated with ER signaling was observed in patients who received single-agent ANA and ANA+PIC, which included genes that regulate the cell cycle, cell death, survival, growth and proliferation and known ER target genes (e.g., PGR, GREB1). In addition, transcripts related to growth factor signaling pathway appeared to be specifically modulated in the ANA+PIC arm, possibly via the upregulation of the expression of RTK ligands. There were no clear changes in PI3K-related phosphoproteins (e.g., AKT, S6, 4E-BP1) in the post-treatment samples by RPPA. However, known PI3K-regulated genes, IRS2 and PIK3IP1, were upregulated in the post-treatment samples and a composite PI3K gene expression signature score (O'Brien et al. 2010) was reduced in both study arms following treatment. This PI3K signature was associated with pre-treatment luminal B status (n=27) and, consistent with this finding, the baseline PI3K gene signature score in the ANA arm, but not the ANA+PIC arm, was inversely associated with the decrease in post treatment Ki67. The tumor immune microenvironment was analyzed though the use of composite gene sets. In our initial observations, analysis of pre- and post-treatment samples showed that 2-week treatment with ANA resulted in a modest increase in transcripts associated with multiple immune signatures, which was further enhanced by the addition of PIC. Conclusions: Gene expression analysis of pre- and post-treatment samples in the OPPORTUNE study demonstrates on-target inhibition of ER and PI3K signaling networks. The analysis of additional paired samples is in progress to further assess if 2-weeks of treatment with a regimen containing an AI in patients with early breast cancer impacts the tumor immune microenvironment. Citation Format: Schmid P, Pinder SE, Bundred N, Wheatley D, Macaskill J, Zammit C, Hu J, Price R, Shia A, Lim L, Parker P, Molinero L, Yu J, O'Brien C, Wilson T, Savage H, Derynck M, Lackner MR, Amler L, Purushotham A, Thompson A, Gendreau S. Transcript analysis of PI3K and immune-related genes and gene signatures in the pre- and post-treatment samples from the window of opportunity study of anastrozole and anastrozole with pictilisib (GDC-0941) in patients with HR-positive early breast cancer (OPPORTUNE study). [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P5-13-01.

Letrozole is a commonly used treatment option for metastatic hormone receptor-positive (HR+) breast cancer, but many patients ultimately relapse. Due to the importance of phosphoinositide-3 kinase (PI3K) in breast cancer, PI3K inhibitors such as taselisib are attractive for combination with endocrine therapies such as letrozole. Taselisib was evaluated as a single agent and in combination with letrozole in a breast cancer cell line engineered to express aromatase. The combination of taselisib and letrozole decreased cellular viability and increased apoptosis relative to either single agent. Signaling cross-talk between the PI3K and ER pathways was associated with efficacy for the combination. In a secreted factor screen, multiple soluble factors, including members of the epidermal and fibroblast growth factor families, rendered breast cancer cells non-responsive to letrozole. It was discovered that many of these factors signal through the PI3K pathway and cells remained sensitive to taselisib in the presence of the soluble factors. We also found that letrozole resistant lines have elevated PI3K pathway signaling due to an increased level of p110α, but are still sensitive to taselisib. These data provide rationale for clinical evaluation of PI3K inhibitors to overcome resistance to endocrine therapies in ER+ breast cancer.

Purpose: We describe the preclinical pharmacology and antitumor activity of GDC-0068, a novel highly selective ATP-competitive pan-Akt inhibitor currently in clinical trials for the treatment of human cancers. Experimental Design: The effect of GDC-0068 on Akt signaling was characterized using specific biomarkers of the Akt pathway, and response to GDC-0068 was evaluated in human cancer cell lines and xenograft models with various genetic backgrounds, either as a single agent or in combination with chemotherapeutic agents. Results: GDC-0068 blocked Akt signaling both in cultured human cancer cell lines and in tumor xenograft models as evidenced by dose-dependent decrease in phosphorylation of downstream targets. Inhibition of Akt activity by GDC-0068 resulted in blockade of cell-cycle progression and reduced viability of cancer cell lines. Markers of Akt activation, including high-basal phospho-Akt levels, PTEN loss, and PIK3CA kinase domain mutations, correlate with sensitivity to GDC-0068. Isogenic PTEN knockout also sensitized MCF10A cells to GDC-0068. In multiple tumor xenograft models, oral administration of GDC-0068 resulted in antitumor activity ranging from tumor growth delay to regression. Consistent with the role of Akt in a survival pathway, GDC-0068 also enhanced antitumor activity of classic chemotherapeutic agents. Conclusions: GDC-0068 is a highly selective, orally bioavailable Akt kinase inhibitor that shows pharmacodynamic inhibition of Akt signaling and robust antitumor activity in human cancer cells in vitro and in vivo. Our preclinical data provide a strong mechanistic rationale to evaluate GDC-0068 in cancers with activated Akt signaling. Clin Cancer Res; 19(7); 1760–72. ©2012 AACR.

Breast cancer is a heterogeneous disease and patients are managed clinically based on ER, PR, HER2 expression, and key risk factors. We sought to characterize the molecular landscape of high-risk breast cancer patients enrolled onto an adjuvant chemotherapy study to understand how disease subsets and tumor immune status impact survival. DNA and RNA were extracted from 861 breast cancer samples from patients enrolled onto the United States Oncology trial 01062. Samples were characterized using multiplex gene expression, copy number, and qPCR mutation assays. HR+ patients with a PIK3CA mutant tumor had a favorable disease-free survival (DFS; HR 0.66, P=0.05), however, the prognostic effect was specific to luminal A patients (Luminal A: HR 0.67, P=0.1; Luminal B: HR 1.01, P=0.98). Molecular subtyping of triple-negative breast cancers (TNBCs) suggested that the mesenchymal subtype had the worst DFS, whereas the immunomodulatory subtype had the best DFS. Profiling of immunologic genes revealed that TNBC tumors (n=280) displaying an activated T-cell signature had a longer DFS following adjuvant chemotherapy (HR 0.59, P=0.04), while a distinct set of immune genes was associated with DFS in HR+ cancers. Utilizing a discovery approach, we identified genes associated with a high risk of recurrence in HR+ patients, which were validated in an independent data set. Molecular classification based on PAM50 and TNBC subtyping stratified clinical high-risk patients into distinct prognostic subsets. Patients with high expression of immune-related genes showed superior DFS in both HR+ and TNBC. These results may inform patient management and drug development in early breast cancer.

Mutations in ESR1 have been associated with resistance to aromatase inhibitor (AI) therapy in patients with ER+ metastatic breast cancer. Little is known of the impact of these mutations in patients receiving selective oestrogen receptor degrader (SERD) therapy. In this study, hotspot mutations in ESR1 and PIK3CA from ctDNA were assayed in clinical trial samples from ER+ metastatic breast cancer patients randomized either to the SERD fulvestrant or fulvestrant plus a pan-PI3K inhibitor. ESR1 mutations are present in 37% of baseline samples and are enriched in patients with luminal A and PIK3CA-mutated tumours. ESR1 mutations are often polyclonal and longitudinal analysis shows distinct clones exhibiting divergent behaviour over time. ESR1 mutation allele frequency does not show a consistent pattern of increases during fulvestrant treatment, and progression-free survival is not different in patients with ESR1 mutations compared with wild-type patients. ESR1 mutations are not associated with clinical resistance to fulvestrant in this study., Fulvestrant degrades the oestrogen receptor. Here, the authors report on a clinical trial using fulvestrant and show that mutations in the oestrogen receptor alpha gene are prevalent in circulating tumour DNA and do not influence the clinical outcome of patients to fulvestrant.

The insulin-like growth factor-I receptor (IGF-IR) pathway is required for the maintenance of the transformed phenotype in neoplastic cells and hence has been the subject of intensive drug discovery efforts. A key aspect of successful clinical development of targeted therapies directed against IGF-IR will be identification of responsive patient populations. Toward that end, we have endeavored to identify predictive biomarkers of response to an anti-IGF-IR-targeting monoclonal antibody in preclinical models of breast and colorectal cancer. We find that levels of the IGF-IR itself may have predictive value in these tumor types and identify other gene expression predictors of in vitro response. Studies in breast cancer models suggest that IGF-IR expression is both correlated and functionally linked with estrogen receptor signaling and provide a basis for both patient stratification and rational combination therapy with antiestrogen-targeting agents. In addition, we find that levels of other components of the signaling pathway such as the adaptor proteins IRS1 and IRS2, as well as the ligand IGF-II, have predictive value and report on the development of a pathway-focused panel of diagnostic biomarkers that could be used to test these hypotheses during clinical development of IGF-IR-targeting therapies. [Mol Cancer Ther 2009;8(8):2110–21]

Purpose: The pathways underlying basal-like breast cancer are poorly understood, and as yet, there is no approved targeted therapy for this disease. We investigated the role of mitogen-activated protein kinase kinase (MEK) and phosphatidylinositol 3-kinase (PI3K) inhibitors as targeted therapies for basal-like breast cancer. Experimental Design: We used pharmacogenomic analysis of a large panel of breast cancer cell lines with detailed accompanying molecular information to identify molecular predictors of response to a potent and selective inhibitor of MEK and also to define molecular mechanisms underlying combined MEK and PI3K targeting in basal-like breast cancer. Hypotheses were confirmed by testing in multiple tumor xenograft models. Results: We found that basal-like breast cancer models have an activated RAS-like transcriptional program and show greater sensitivity to a selective inhibitor of MEK compared with models representative of other breast cancer subtypes. We also showed that loss of PTEN is a negative predictor of response to MEK inhibition, that treatment with a selective MEK inhibitor caused up-regulation of PI3K pathway signaling, and that dual blockade of both PI3K and MEK/extracellular signal–regulated kinase signaling synergized to potently impair the growth of basal-like breast cancer models in vitro and in vivo. Conclusions: Our studies suggest that single-agent MEK inhibition is a promising therapeutic modality for basal-like breast cancers with intact PTEN, and also provide a basis for rational combination of MEK and PI3K inhibitors in basal-like cancers with both intact and deleted PTEN.

The phosphoinositol-3-kinase (PI3K) signaling pathway is one of the most frequently activated pathways in cancer, and controls critical cellular processes such as proliferation, transcription and survival. Taselisib (GDC0032) is an orally bioavailable, potent, and selective inhibitor of Class I PI3K alpha, delta, and gamma isoforms, with 30fold less inhibition of the PI3K beta isoform relative to the PI3K alpha isoform. Previously published data demonstrated that taselisib has increased activity against PIK3CA mutant cancer cell lines (Ndubaku CO et al, J Med Chem, 2013). A panel of 50 breast cancer cell lines were profiled for sensitivity to taselisib using viability assays and subsequently correlated with PIK3CA mutations and expression of ER, PR, HER2 and PTEN. The PIK3CA mutant cell lines displayed an average of 30-fold greater sensitivity in IC50 compared to the PIK3CA wild-type cell lines. PTEN null cell lines were largely refractory to taselisib. Within the PIK3CA wild-type cell lines, the HER2 amplified cells showed an average of 18-fold greater sensitivity in IC50 over the HER2 wild-type cell lines. No differences in taselisib sensitivity were seen between the kinase, helical, or the C2 domain of the PIK3CA gene within the PIK3CA mutant cell lines. In HER2 amplified PIK3CA wild-type cell lines, taselisib did not fully suppress pAKT and pS6 signaling, however full suppression was observed in combination with the HER2 inhibitor, lapatinib. Increased apoptosis was observed with the combination of taselisib and lapatinib in the HER2 amplified, PIK3CA mutant cells, but not in HER2 amplified, PIK3CA wild-type cells. In PIK3CA mutant luminal cell lines, taselisib induced ESR1 transcriptional activity, but not in PIK3CA wild-type luminal lines, as assessed using PR, GREB1 and IGFBP4 gene expression. This induction was ER- and estradiol-dependent, and was suppressed using the estrogen degrader, fulvestrant. Lastly, as up to 40% of ER+, second line PIK3CA mutant metastatic tumors have been shown to harbor a mutation in the ESR1 gene (Spoerke JM et al., Nat Commun, 2016), we assessed the crosstalk between the PI3K and ER pathways in PIK3CA mutant MCF-7 cells that overexpress two ESR1 hotspot mutations. These ESR1 mutant cell lines displayed increased ER transcriptional activity, which was further activated by taselisib, an observation that was reversed with fulvestrant co-treatment. These data suggest that the combination of taselisib and an estrogen degrader can suppress crosstalk between PI3K and ER in both an ESR1 mutant and wild-type background, and supports the ongoing SANDPIPER study of taselisib in combination with fulvestrant in metastatic ER+, PIK3CA mutant cancers that have progressed following an aromatase inhibitor therapy (NCT02340221). Citation Format: Heidi M. Savage, Heather M. Moore, Carol L. O'Brien, Wei Zhou, Ciara Metcalfe, Mark R. Lackner, Timothy R. Wilson. Identifying preclinical predictive biomarkers to taselisib in breast cancer cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1780. doi:10.1158/1538-7445.AM2017-1780

First two authors contributed equally Last two authors contributed equally Recent studies have shown that ex vivo propagation of normal tissues or patient-derived tumor cells in presence of irradiated fibroblast feeder cells and ROCK inhibitor can rapidly establish conditionally reprogramed cells (CRCs). In case of normal tissues, the induction of CRCs was reversible when the ROCK inhibitor and the feeder cells were removed, resulting in CRCs differentiating to its tissue origin (Liu et al.2012). Previous publications suggested that the establishment of such cell models provides new strategies to understand acquired resistance during treatment (Crystal et al 2015) and to predict treatment response (Liu et al. 2014). However, gene expression modulations and genomic drifting during the establishment of CRC propagation have not been thoroughly studied. The primary goal of this study is to molecularly characterize alterations between the original tumor tissues and the derived models growing with or without ROCK inhibitor. Understanding in-depth molecular fluctuations in this patient-derived ex vivo system will facilitate its appropriate use for tumor biology experimentation. Herein, tumors from prostate cancer and breast cancer patients were surgical removed and cryopreserved at the clinical site then processed and cultured as previously described (Liu et al. 2012). Gene expression profiling and next-generation sequencing were carried out on the original tumor tissues and cellular models passaged during the CRC propagation in the presence or absence of ROCK inhibitor. Gene expression analysis of the prostate cancer cells and the breast cancer cells were carried out using a 93-gene prostate cancer-focused Fluidigm panel and a 800 gene NanoString breast cancer-focused panel, respectively. Cancer hotspot mutations were analyzed using the Ion Torrent Cancer Hotspot v2 NGS assay. Through aforementioned genomic and transcriptomic interrogations, we demonstrated the extent of indication-relevant gene expression modulation during establishment and propagation of these cells. We also characterize cancer hotspot mutations in the primary tumor cells and the stability of those mutations during ex vivo propagation. These results should begin to inform the appropriate use of the CRC model for tumor biology experimentation. Citation Format: Jessica Li, Bonnie Liu, Rin Nakamura, Heidi Savage, Shoji Ikeda, Timothy Wilson, Teiko Sumiyoshi, Garret Hampton, Lukas Amler, Mark Lackner, Shih-Min A. Huang, Walter C. Darbonne. Gene expression and genomic drift comparative analysis between patient-derived conditionally reprogrammed cells and original tumors. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4257.

The phosphoinositide-3 kinase (PI3K) pathway is one of the most frequently activated signaling pathways in human cancer and represents a promising therapeutic target. Taselisib (GDC-0032), a potent and selective inhibitor of the PI3K alpha, delta and gamma isoforms, but less potent in the beta isoform, displays increased sensitivity in models that are driven by mutant PIK3CA (Ndubaku CO et al, J Med Chem, 2013). Selective pressure caused by, small molecules such as PI3K inhibitors will ultimately lead to clinical resistance and disease progression. Recently, a published mechanism of acquired resistance to the PI3K inhibitor alpelisib (BYL-719, a PI3K alpha-selective inhibitor) reported that genomic loss of the tumor suppressor PTEN led to clinical progression in a metastatic breast cancer patient harboring an activating PIK3CA mutation (Juric D et al, Nature, 2014). As taselisib and alpelisib are next generation PI3K inhibitors, we sought to determine if there are any differences to drug sensitivity in PIK3CA mutant cells that have concomitant loss of PTEN. To investigate this hypothesis, we first measured viability in breast cancer cells, each with differing genetic backgrounds treated with two PI3K inhibitors, taselisib and alpelisib. We found that PIK3CA mutant and PIK3CA mutant/PTEN null cells were sensitive to treatment when compared to wild-type and PTEN null cells. However, PIK3CA mutant/PTEN null cells displayed higher IC50 values for both inhibitors when compared to PIK3CA mutant cells. To control for differences in genetic backgrounds, we next tested whether knockdown (KD) of PTEN expression in PIK3CA mutant breast cancer cells would promote resistance and reactivate the PI3K signaling cascade even in the presence of a PI3K inhibitor. Transient, siRNA-targeted KD of PTEN in the PIK3CA mutant breast cancer cell lines EFM19 and T47D conferred resistance to alpelisib after 6 days of drug treatment, an observation that was less pronounced in taselisib treated cells. Additionally, PTEN KD in DMSO-treated cells led to an increase in cell proliferation and enhanced downstream PI3K signaling as measured by phosphorylated AKT and S6 proteins when compared to non-targeting siRNA controls. The reactivated, pro-survival signaling with PTEN KD was maintained upon treatment with both inhibitors when compared to control cells, an observation that was more pronounced in alpelisib treated cells. These data suggest that PIK3CB can compensate for PIK3CA inhibition and that PTEN loss could lead to clinical resistance to alpelisib, but may not constitute a resistance mechanism to taselisib. As both molecules have entered phase 3 studies in estrogen receptor positive breast cancer, understanding how biomarker-defined genotypes respond to next generation PI3K inhibitors is critical for identifying patients most likely to gain therapeutic benefit. Citation Format: Heather M. Moore, Heidi M. Savage, Mark R. Lackner, Timothy R. Wilson. Deciphering the role of PTEN as a predictive biomarker to next generation isoform-selective PI3K inhibitors in PIK3CA mutant breast cancer cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 242.

The phosphoinositol-3-kinase (PI3K) signaling pathway is one of the most frequently activated pathways in oncogenesis, and controls critical cellular processes such as proliferation, transcription and survival. Taselisib (GDC 0032) is an orally bioavailable, potent, and selective inhibitor of Class I PI3K alpha, delta, and gamma isoforms, with 30 fold less inhibition of the PI3K beta isoform relative to the PI3K alpha isoform. The gene that encodes the p110 alpha isoform of PI3K, PIK3CA, is frequently mutated in breast, colorectal and endometrial cancers. Previously published data demonstrated that taselisib has increased activity against PIK3CA mutant cancer cell lines (Ndubaku CO et al, J Med Chem, 2013). To determine if there are additional predictive biomarkers outside of PIK3CA mutations for sensitivity to taselisib, we profiled over 550 cell lines, encompassing 13 of the main tissue types, to increasing concentrations of taselisib. As expected, a small percentage of all tumor types responded to taselisib and correlated strongly with PIK3CA mutations. Intriguingly, the majority of head and neck squamous cell cancer (HNSCC) cell lines showed an IC50 concentration of less than 0.5uM, suggesting that HNSCC cell lines may be particularly susceptible to PI3K inhibition. Interestingly, the majority of HNSCC cell lines displayed an activated PI3K pathway as evidenced by phosphorylation of key proteins, such as AKT, S6 and PRAS40 implicating that downstream phospho-protein analysis of the PI3K pathway may not predict for sensitivity to taselisib. Mutational profiling identified that only 3 out of the tested 31 HNSCC cell lines harbored a mutation within PIK3CA, suggesting that additional genotypes may explain the sensitivity to taselisib. Additional molecular analysis assessing both gene expression and copy number levels of genes relevant to the PI3K pathway found that many of the taselisib sensitive cell lines had an activated ERBB signaling node through low level amplification of ERBB receptors (e.g. EGFR, FGFR1) or via a NRG1:ERBB3 autocrine mechanism. As many ERBB receptors heterodimerize with ERBB3, which contains p85 binding sites and potentially activates the PI3K pathway, we next tested whether taselisib would enhance the potency of ERBB receptor inhibitors in biomarker-defined subset of HNSCC cell lines. Combination screens were performed in cell lines with amplified EGFR, ERBB2, and NRG1:ERBB3 autocrine signaling with taselisib + tarceva or lapatinib. We found that combination effects assessed using the BLISS excess method showed a synergistic interaction. These results suggest that taselisib may have therapeutic potential for the treatment of HNSCC. Citation Format: Heidi M. Savage, Carol O’Brien, Heather Moore, Mark R. Lackner, Robert Yauch, Jeffrey Settleman, Timothy R. Wilson. Taselisib enhances the potency of ERBB inhibitors in biomarker-defined subsets of head and neck squamous carcinoma cell lines. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 372.

Background Evaluation of cancer biomarkers from blood could significantly enable biomarker assessment by providing a relatively non-invasive source of representative tumor material. Circulating Tumor Cells (CTCs) isolated from blood of metastatic cancer patients hold significant promise in this regard. Methodology/Principal Findings Using spiked tumor-cells we evaluated CTC capture on different CTC technology platforms, including CellSearch® and two biochip platforms, and used the isolated CTCs to develop and optimize assays for molecular characterization of CTCs. We report similar performance for the various platforms tested in capturing CTCs, and find that capture efficiency is dependent on the level of EpCAM expression. We demonstrate that captured CTCs are amenable to biomarker analyses such as HER2 status, qRT-PCR for breast cancer subtype markers, KRAS mutation detection, and EGFR staining by immunofluorescence (IF). We quantify cell surface expression of EGFR in metastatic lung cancer patient samples. In addition, we determined HER2 status by IF and FISH in CTCs from metastatic breast cancer patients. In the majority of patients (89%) we found concordance with HER2 status from patient tumor tissue, though in a subset of patients (11%), HER2 status in CTCs differed from that observed in the primary tumor. Surprisingly, we found CTC counts to be higher in ER+ patients in comparison to HER2+ and triple negative patients, which could be explained by low EpCAM expression and a more mesenchymal phenotype of tumors belonging to the basal-like molecular subtype of breast cancer. Conclusions/Significance Our data suggests that molecular characterization from captured CTCs is possible and can potentially provide real-time information on biomarker status. In this regard, CTCs hold significant promise as a source of tumor material to facilitate clinical biomarker evaluation. However, limitations exist from a purely EpCAM based capture system and addition of antibodies to mesenchymal markers could further improve CTC capture efficiency to enable routine biomarker analysis from CTCs.

Purpose: The class I phosphatidylinositol 3′ kinase (PI3K) plays a major role in proliferation and survival in a wide variety of human cancers. A key factor in successful development of drugs targeting this pathway is likely to be the identification of responsive patient populations with predictive diagnostic biomarkers. This study sought to identify candidate biomarkers of response to the selective PI3K inhibitor GDC-0941. Experimental Design: We used a large panel of breast cancer cell lines and in vivo xenograft models to identify candidate predictive biomarkers for a selective inhibitor of class I PI3K that is currently in clinical development. The approach involved pharmacogenomic profiling as well as analysis of gene expression data sets from cells profiled at baseline or after GDC-0941 treatment. Results: We found that models harboring mutations in PIK3CA, amplification of human epidermal growth factor receptor 2, or dual alterations in two pathway components were exquisitely sensitive to the antitumor effects of GDC-0941. We found that several models that do not harbor these alterations also showed sensitivity, suggesting a need for additional diagnostic markers. Gene expression studies identified a collection of genes whose expression was associated with in vitro sensitivity to GDC-0941, and expression of a subset of these genes was found to be intimately linked to signaling through the pathway. Conclusion: Pathway focused biomarkers and the gene expression signature described in this study may have utility in the identification of patients likely to benefit from therapy with a selective PI3K inhibitor. Clin Cancer Res; 16(14); 3670–83. ©2010 AACR.

Although breast cancer molecular subtypes have been extensively defined by means of gene expression profiling over the past decade, little is known, at the proteomic level, as to how signaling pathways are differentially activated and serve to control proliferation in different breast cancer subtypes. We used reverse-phase protein arrays to examine phosphorylation status of 100 proteins in a panel of 30 breast cancer cell lines and showed distinct pathway activation differences between different subtypes that are not obvious from previous gene expression studies. We also show that basal levels of phosphorylation of key signaling nodes may have diagnostic utility in predicting response to selective inhibitors of phosphatidylinositol 3-kinase and mitogen-activated protein kinase/extracellular signal-regulated kinase kinase. Finally, we show that reverse-phase protein arrays allow the parallel analysis of multiple pharmacodynamic biomarkers of response to targeted kinase inhibitors and that inhibitors of epidermal growth factor receptor and mitogen-activated protein kinase/extracellular signal-regulated kinase kinase result in compensatory up-regulation of the phosphatidylinositol 3-kinase/Akt signaling pathway. [Mol Cancer Ther 2008;7(12):3695–706]

The PI3K signaling pathway is one of the most frequently activated pathways in cancer, and controls critical proliferative and survival processes. Taselisib (GDC-0032) is a novel, oral, selective inhibitor of PI3K, sparing inhibition of PI3Kβ. (Ndubaku CO et al. J Med Chem 2013). Preclinically, taselisib demonstrates enhanced potency in PIK3CA mutant cells, with greater signaling and growth/survival effects in mutant cells vs. wild-type cells. In the phase Ia study with single agent taselisib, partial responses (PRs) were observed in 6/34 enrolled patients. All 6 responses were observed in patients with PIK3CA mutant tumors (Juric D. et al. AACR 2013), indicating the need to determine PIK3CA mutation status from patients treated with taselisib. Circulating tumor DNA (ctDNA) provides an up-to-date liquid biopsy that can be used to detect somatic mutation in oncogenes especially when tumor tissue is not available. Analysis was undertaken on five cohorts of patients enrolled onto the taselisib Phase 1b study comprised of 106 patients. In total, 87 patients had tumor tissue available for PIK3CA mutation testing. Plasma was collected from 91 patients prior to treatment and 10 patients had matched plasma collected at the study discontinuation visit. Tissue samples were assessed using the Roche cobas PIK3CA mutation test that detects 17 PIK3CA hotspot mutations, of which 47 (54%) tissue samples were positive for PIK3CA mutations. Analysis of PIK3CA and AKT1 mutations from plasma was performed using an Inostics BEAMing digital PCR Oncobeam panel, which detects 8 PIK3CA hotspot mutations and 1 AKT1 mutation. ctDNA was detected in all 91 patients sampled, and ranged from 1.3ng to 1ug per milliliter of plasma. PIK3CA mutations were identified in the plasma of 51 patients (56%), and an AKT1 mutation was identified in one patient. For 80 patients that had both tissue and plasma available for PIK3CA mutation testing, a sensitivity of 81%, a specificity of 74% and an overall accuracy of 78% was observed. Allele frequency ranged from 0.02% to 75.58% with a median of 2.72%. In two cohorts that enrolled hormone receptor positive, HER2 negative breast cancer patients, a sensitivity of 84%, a specificity of 87% and an overall accuracy of 88% was observed between 42 matched tissue and plasma samples. In a patient who experienced a sustained clinical partial response to taselisib in combination with fulvestrant, a decrease in plasma PIK3CA allele frequency was observed from 3.05% in the pretreatment sample to 0.14% in the study discontinuation sample. In summary, analysis of PIK3CA mutations from plasma in patients enrolled onto trials testing the efficacy of taselisib may be a useful surrogate for patients when tissue is unavailable, an observation that is being assessed in ongoing trials. Citation Format: Timothy R. Wilson, Heidi Savage, Junko Aimi, Jessica Jin, Hema Parmar, Jerry Hsu, Ian Krop, Cristina Saura, Andres Cervantes, Jasgit Sachdev, Manish Patel, Juan Cejalvo, Mafalda Oliveira, Eric Winer, Daniel Von Hoff, Jose Baselga, Dejan Juric. Circulating tumor DNA (ctDNA) analysis of PIK3CA and AKT1 mutations in patients enrolled onto the Phase 1b study of the PI3K inhibitor taselisib (GDC-0032) in solid malignancies. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2399. doi:10.1158/1538-7445.AM2015-2399

The phosphoinositol-3-kinase (PI3K) signaling pathway is one of the most frequently activated pathways in oncogenesis, and controls critical cellular processes such as proliferation, transcription and survival. GDC 0032 is an orally bioavailable, potent, and selective inhibitor of Class I PI3K alpha, delta, and gamma isoforms, with 30 fold less inhibition of the PI3K beta isoform relative to the PI3K alpha isoform. The gene that encodes the p110 alpha isoform of PI3K, PIK3CA, is frequently mutated in breast, colorectal and endometrial cancers. Previously published data demonstrated that GDC-0032 has increased activity against PIK3CA mutant cancer cell lines (Olivero A. et al., DDT02-01, AACR 2013). To determine if there are additional predictive biomarkers outside of PIK3CA mutations for sensitivity to GDC-0032, we profiled over 550 cell lines, encompassing 13 of the main cancer types, to increasing concentrations of GDC-0032. As expected, a small percentage of all tumor types responded to GDC-0032 and, were assessed, correlated strongly with PIK3CA mutations. Intriguingly, the majority of head and neck squamous cell cancer (HNSCC) cell lines showed an IC50 concentration of less than 1uM, suggesting that HNSCC cell lines may be particularly susceptible to PI3K inhibition. Molecular profiling identified that only 10% of the HNSCC cell lines had a mutation within PIK3CA, and no consistent copy number alterations were observed in receptor tyrosine kinases or PTEN. To investigate this observation further, we developed a ”PIK3CA activation” algorithm, by utilizing the mutant-selective property of GDC-0032, and calculated a ratio of GDC-0032 to GDC-0941, a pan PI3K inhibitor with reduced in vitro selectivity for PIK3CA mutations. Following a proof of principle experiment in breast cancer cells, a GDC-0941/GDC-0032 ratio of ≥4 enriched for breast cancer cells with mutant PIK3CA (11 out of 13 cell lines). Application of the algorithm, to our panel of HNSCC cell lines identified 10 out of 34 cell lines that had a GDC-0941/GDC-0032 ratio of ≥4, with only 1 of these cell lines being PIK3CA mutant. Further molecular characterization identified high mRNA expression of neuregulin-1, the ligand for HER3, in GDC-0032 cell lines with a GDC-0941/GDC-0032 ratio of ≥4, compared to cell lines with a ratio of In conclusion, high throughput cell line screening identified a “PIK3CA activated” subset of HNSCC cell lines that correlated with increased neuregulin-1 expression and corresponding sensitivity to GDC-0032, outside of PIK3CA mutations. Citation Format: Heidi M. Savage, Carol O'Brien, Bob Yauch, Jeff Settleman, Mark R. Lackner, Timothy R. Wilson. Neuregulin-1 expression correlates with sensitivity to the PI3K inhibitor, GDC-0032, outside of PIK3CA mutations in head and neck cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2631. doi:10.1158/1538-7445.AM2014-2631

The phosphoinositol-3-kinase (PI3K) signaling pathway is one of the most frequently activated pathways in oncogenesis, and controls critical cellular processes such as proliferation, transcription and survival. GDC 0032 is an orally bioavailable, potent, and selective inhibitor of Class I PI3K alpha, delta, and gamma isoforms, with 30 fold less inhibition of the PI3K beta isoform relative to the PI3K alpha isoform. The gene that encodes the p110 alpha isoform of PI3K, PIK3CA, is frequently mutated in breast, colorectal and endometrial cancers. Preclinical data indicate that GDC-0032 has increased activity against PIK3CA mutant cancer cell lines. As presented by Juric et al., 34 patients were enrolled in this study and dose escalation has been completed (Juric D. et al. AACR 2013, Abstract LB-64). Metabolic partial responses via FDG-PET were observed in 6 out of 13 patients assessed, and clinical partial responses (PRs) were observed in 6 patients, with 5 of these patients having PIK3CA mutant cancers. Tumor tissue was obtained from 30 out of 34 patients enrolled onto the study. The most frequently recruited cancer type was breast cancer (41%) followed by colorectal (15%) and lung cancer (15%). Fourteen patients enrolled onto the study had PIK3CA mutant tumors (41%) as determined using a TaqMan-based PCR assay. Three patients had total loss of PTEN expression, as assessed by immunohistochemical staining, and a further 3 patients had reduced PTEN expression based on a H-score assessment. Out of the 6 patients that had reduced PTEN expression, 2 patients had coexisting mutations with PIK3CA. Where known, 4 out of 5 patients that had a PR showed intact PTEN expression, with the fifth patient having reduced PTEN expression. Samples were further analyzed using an in-house developed PCR-based multiplexed assay that detects activating mutations within an additional ten oncogenes. PIK3CA mutations were largely mutually exclusive with mutations in the Ras pathway, however 3 out of the 14 PIK3CA mutant patients had a coexisting mutation within KRas. A further 2 patients had KRas mutations, 1 patient had a NRas mutation and 2 patients had EGFR mutations. Preliminary analysis suggests lack of benefit in patients with KRas mutations treated with GDC-0032 single agent. Optional on-study biopsies were collected from 2 patients and demonstrated pharmacodynamic inhibition of the PI3K pathway as assessed by reverse phase protein array for approximately 45 endpoints, including 1 patient at the lowest dose. In conclusion, our preliminary data indicates that GDC-0032 demonstrates single agent activity in patients with PIK3CA mutations tumors with unaltered PTEN or MAP-kinase pathways. Citation Format: Timothy R. Wilson, Heidi Savage, Carol O'Brien, Sandra Sanabria, Ray S. Lin, Marie-Claire Wagle, Yibing Yan, Mark R. Lackner, Hema Parmar, Jerry Y. Hsu, Dejan Juric, Ian E. Krop, Ramesh K. Ramanathan, Daniel D. Von Hoff, Jose Baselga. Expanded biomarker results from a phase I dose escalation study of GDC-0032, a beta isoform-sparing PI3K inhibitor. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 915. doi:10.1158/1538-7445.AM2014-915

Current clinical practice stratifies patients for treatment and enrollment in clinical breast cancer studies based on ER, PR and HER2 status. Emerging data suggest that these three biomarkers do not fully capture the heterogeneity that exists within breast cancer. In addition, understanding the genetic landscape of breast cancer is critical for defining clinically actionable patient subsets that may derive benefit from targeted therapies. PIK3CA and TP53 mutations have previously been reported to be the most common mutations within breast cancer; however the prognostic significance of these alterations, as well as their overlap/exclusivity with other important biomarkers is poorly understood. USON 01062 (O’Shaughnessy J, et al. Proc SABCS, 2010, abstract S4-2) is a phase III study that evaluated the addition of capecitabine to standard adjuvant chemotherapy. The study did not meet its primary endpoint of 5-year disease-free survival (HR 0.84, p=0.125), but showed improvement in overall survival (HR 0.68, p=0.011). FFPE primary breast cancers were obtained from approximately 2000 of the 2610 patients (pts) enrolled, and here we report results from our comprehensive biomarker analyses on 817 of these pts. Samples were profiled using a multiplexed PCR-based platform to determine somatic mutations in 6 key oncogenes, as well as assayed for 800 breast cancer-related genes encompassing a variety of signaling pathways and published breast cancer signatures. IHC for the proliferation marker Ki67 and the tumor suppressor PTEN was also performed. Intrinsic subtyping analysis determined that 296 pts’ cancers were basal, 69 were HER2-enriched, 327 were luminal A, 124 were luminal B and 1 was normal-like. Approximately 80% of the triple negative breast cancers (TNBCs) were of the basal subtype, and 80% of the ER+ cancers were of the luminal A and B subtypes. The most heterogeneous group was the HER2-enriched group that was comprised of 60% HER2+ and 30% TN cancers. PIK3CA mutations were found at a frequency of 44% in luminal A, 24% in luminal B, 33% in HER2-enriched and 3% in the basal subtypes. Within the TNBC subset, 42% were basal like 1/2, 21% were immunomodulatory, 15% were mesenchymal stem-like, 13% were mesenchymal-like and 9% were luminal AR. In conclusion, luminal A breast cancers were the most common subtype treated with chemotherapy on this adjuvant trial that evaluated the effectiveness of the anti-proliferative agent, capecitabine. Intrinsic subtyping and molecular profiling of pts’ primary breast cancer reveals the substantial heterogeneity of early breast cancers, highlighting the challenges in identifying patients who may benefit from adjuvant capecitabine based on standard clinical-pathologic features. Analysis of intrinsic subtyping, Ki67 levels and genomic alterations of these early breast cancer pts will be presented. Citation Format: Carol Lynn O'Brien, Tim R. Wilson, Jill M. Spoerke, Yuanyuan Xiao, Heidi Savage, Rachel Tam, Erica Schleifman, Rajesh Patel, Ling Huw, Hartmut Koeppen, Ling Fu, Teiko Sumiyoshi, Joyce O'Shaughnessy, Mark Lackner. Molecular biomarker profiling of archival primary breast cancers from a Phase 3 adjuvant study of capecitabine in early stage breast cancer patients. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-179. doi:10.1158/1538-7445.AM2014-LB-179

582 Background: The phase III trial USON 01062 (O’Shaughnessy J, et al Proc SABCS, 2010, abstract S4-2) showed that the addition of capecitabine to docetaxel following doxorubicin/cyclophosphamide ...

Introduction. Understanding the genetic landscape of estrogen receptor positive breast caner is critical for defining clinically actionable alleles that may be targeted using next generation biologics. PIK3CA mutations have previously been reported to be the most common mutation within estrogen receptor positive breast cancer, however the overlap and mutual exclusivity with other key driving alleles is poorly understood, especially how these biomarkers change following treatment failure. We custom designed a mutation, copy number variation and RNA expression panels to profile biomarkers from low quality formalin fixed paraffin embedded extracted material. Data relating to key pathways, such as PI3K and immune modulatory pathways, and overlap with other biomarkers will be presented. Methods. Formalin fixed paraffin embedded material was available from 195 primary cases and 95 paired metastatic estrogen receptor positive breast cancer patients. Samples were assayed for the expression of PTEN by IHC, hotspot mutations in 11 oncogenic driving genes using Q-PCR based technology, copy number alterations in 42 genes using Q-PCR based technology and RNA expression using a custom designed 400 breast cancer specific NanoString gene panel. Results. PTEN loss, as defined by H-score of 0, was found in 4.5% of primary samples and in 4.3% of metastatic samples. PIK3CA mutations were found in 43% of primary samples and in 38% of metastatic samples and were largely found to be mutually exclusive with PTEN loss. AKT1 mutations were found in 5.4% of primary samples and in 4.1% of metastatic samples. Less frequent mutations in KRAS (7.0%) and BRAF (1.7%) were found in the primary sample, and some were co-existing with PIK3CA mutations. Copy number gains were found in CCND1 (23.5%), ZNF703 (19.5%), FGFR1 (16.8%) and PAK1 (11.4%) and were largely concordant with the paired metastatic sample. Analysis with clinical outcome is ongoing. Conclusions. PIK3CA was the most frequently altered gene detected and was mutually exclusive with other key driving mutations within the PI3K pathway (AKT1 and PTEN). A strong concordance was observed between genetic alterations found in the primary sample and the paired metastatic sample. Mutations and expression gains within clinically actionable targets were found less frequently, but may provide alternative treatment strategies for these patients following failure of endocrine therapy. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P6-05-12.

Introduction The PI3-Kinase pathway is one of the most commonly mutated pathways in cancer and plays a major role in cell proliferation and survival. Mutations in PIK3CA, the gene encoding the p110 subunit of PI3K, are among the most common alterations in breast cancer, occurring in approximately 45% of luminal A, 30% of luminal B, 30% of HER2 positive and 8% of triple negative breast cancers. Additional pathway activating alterations include loss of PTEN, AKT mutations and overexpression of PIK3CA and HER2. Development of a pharmacodynamic biomarker is challenging with the more isoform specific PI3K inhibitors as multiple upstream pathways can funnel into common downstream immunohistochemical evaluable endpoints. In addition, phosphorylated epitopes are often labile and do not always lend themselves to immunohistochemical evaluation in the clinical setting. GDC-0032, which is currently under clinical investigation, is a class I PI3K inhibitor with 30-fold less inhibition on PI3K beta relative to PI3K alpha, and the development of a predictive and on-study pharmacodynamic signature may prove informative as compared to traditional IHC endpoints. Methods We screened a panel of 53 breast cancer cell lines, incorporating all subtypes, to GDC-0032 using the cell proliferation assay cell titer glo. To determine if there was a relationship between pathway activation and sensitivity to GDC-0032, we correlated response to PIK3CA mutations, loss of PTEN and HER2 overexpression. Using RNA sequencing, we compared the baseline gene expression between the sensitive and refractory cell lines. Next, to identify an on-study pharmacodynamic gene expression signature, we treated both sensitive and refractory cell lines with GDC-0032 and ran an in-house custom designed 800 gene NanoString breast cancer gene set that incorporated published PI3K pathway signatures, intrinsic subtyping genes and immunological related genes. Finally, the GDC-0032 signature was applied to a set of 160 FFPE breast cancer samples and overlaid with relevant biomarkers. Results and Conclusions Sensitivity to GDC-0032 correlated strongly with PI3K pathway activation including PIK3CA mutations and HER2 overexpression in breast cancer cells. Comparing baseline whole genome RNA expression of GDC-0032 sensitive and refractory cell lines, we identified 293 genes that were differentially expressed. Applying a more stringent statistical cutoff (greater than 2 fold difference and t-test less than 0.01) refined the gene list to 51 genes, which defined the baseline GDC-0032 sensitivity signature. Applying the 800 gene breast cancer NanoString panel to a set of 160 FFPE breast cancer samples, the GDC-0032 sensitivity signature correlated with luminal status and was enriched in PIK3CA mutant tumors. In conclusion, our in-house designed GDC-0032 sensitivity signature correlated strongly with PIK3CA mutations in clinical specimens. However the lack of complete correlation may identify tumors that have an activated PI3K pathway outside of PIK3CA mutations and/or HER2 amplification that may derive clinical benefit from GDC-0032. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P6-05-09.