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Background: TAPUR is a phase II basket study evaluating antitumor activity of commercially available targeted agents in pts with advanced cancers with specific genomic alterations. Results in a cohort of pts with solid tumors with PTEN mut treated with T are reported. Methods: Eligible pts had no standard treatment (tx) options, measurable disease, ECOG performance status (PS) 0-2, and adequate organ function. Genomic testing was performed in CLIA-certified, CAP-accredited site selected labs. After antihistamine pre-tx, 25 mg T was infused over 30-60 minutes once weekly until disease progression. Low accruing histology-specific cohorts with PTEN mut were collapsed into 1 histology-pooled cohort for analysis. Primary end point was disease control (DC), defined as complete or partial response (PR) or stable disease (SD) of at least 16+ weeks (wks) duration (SD16+) per RECIST v1.1. The primary end point was summarized as a proportion and the hypothesized null DC rate of 15% was evaluated by a 1-sided exact binomial test with alpha 0.10. Other end points were progression-free survival (PFS), overall survival (OS), duration of response (DOR), duration of SD and safety. DOR is defined as time from pt’s first documented objective response (OR) until progressive disease (PD). Duration of SD is defined as time from tx start to PD. Results: 34 pts with solid tumors (10 tumor types; 20/34 pts had prostate, soft tissue sarcoma [STS] or lung) with PTEN mut were enrolled. 6 pts were unevaluable for efficacy, 1 was ineligible. Table shows demographics, outcomes and toxicity. 2 pts obtained PR (prostate and STS) and 5 had SD16+ (lung, head and neck, breast, uterine, and site unspecified) for DC rate of 26% (1-sided 90% CI: 15.1%, 100%) and OR rate of 7% (95% CI: 1%, 24%). The null DC rate was rejected. Median PFS was 10 wks and median OS was 32 wks. Conclusions: T met prespecified criteria to declare a signal of activity in pts with solid tumors with PTEN mut. Table: Baseline Characteristics (N=34); Efficacy Outcomes (n=27); Toxicity Outcomes (N=34) Median (Med) age, years (range) 65 (42, 84) Female, % 14 (41) ECOG PS, % 0 13 (38) 1 18 (53) 2 3 (9) Prior systemic regimens, % 1 3 (9) 2 5 (15) ≥3 26 (77) DC rate, % (OR and SD16+) (1-sided 90% CI) 26 (15.1, 100) OR rate, % (95% CI) 7 (1, 24) Med PFS, wks (95% CI) 10 (7, 17) Med OS, wks (95% CI) 32 (13, 42) DOR, wks (n=2) 11 and 55 Med duration SD, wks (n=5) 27 (24, 36) Number of pts1 with tx-related grade 3-4 AE or SAE AE2 12 (35) SAE3 6 (18) 1Pts may have experienced one or more events 2alkaline phosphatase increase, anemia, chronic kidney disease, diarrhea, fatigue, hyperglycemia, hypokalemia, hypophosphatemia, lung infection, oral pain and all SAEs 3acute kidney injury, dyspnea, generalized muscle weakness, hypophosphatemia, lung infection, oral mucositis, pericardial effusion Citation Format: Kristopher Wentzel, Michael Rothe, Pam K. Mangat, Elizabeth Garrett-Mayer, Li Ding, Evan Pisick, Mridula Krishnan, Deepti Behl, Olatunji B. Alese, Carmen J. Calfa, Kathrine A. Cooper, Herbert L. Duvivier, Raghava Reddy Induru, Janet C. Ruzich, Song Zhao, Gina N. Grantham, Abigail Gregory, Susan Halabi, Richard L. Schilsky. Temsirolimus (T) in patients (pts) with solid tumors with PTEN mutation (mut): results from the Targeted Agent and Profiling Utilization Registry (TAPUR) Study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr CT231.

Background: TAPUR is a phase II basket study evaluating antitumor activity of commercially available targeted agents in pts with advanced cancers with specific genomic alterations. Results in a cohort of pts with solid tumors and MET amp or mut treated with C are reported. Methods: Eligible pts had no standard treatment (tx) options, had measurable disease, ECOG performance status (PS) 0-2, and adequate organ function. Pts with non-small cell lung cancer were excluded. Genomic testing was performed in CLIA-certified, CAP-accredited site selected labs. Pts received C at 250 mg orally BID until disease progression. Low accruing histology-specific cohorts with MET amp or mut were collapsed into 1 histology-pooled cohort for analysis. Primary end point was disease control (DC) per investigator, defined as complete or partial response (PR) or stable disease (SD) of at least 16+ weeks (wks) duration (SD16+) per RECIST v1.1. The primary end point was summarized as a proportion and the hypothesized null DC rate of 15% was evaluated by a 1-sided exact binomial test with α= 0.10. Other end points were progression-free survival (PFS), overall survival (OS), duration of response and SD, and safety. Results: 31 pts with solid tumors (12 tumor types) and MET mut only (n=10), amp only (n=19), or mut and amp (n=1) were enrolled; 1 pt with overexpression was ineligible. 3 additional pts were unevaluable for efficacy. Table shows demographics and outcomes. 2 PR (both esophageal adenocarcinoma with MET amp) and 4 SD16+ (2 renal cell carcinoma, 1 with mut, 1 with amp; colorectal with amp; small intestine with amp) were observed for DC rate of 21% (1-sided 90% CI: 12%, 100%) and objective response rate of 7% (95% CI: 1%, 24%). The null DC rate was not rejected. 5 pts had ≥1 grade 3 tx-related adverse or serious adverse event. Conclusions: C did not meet prespecified criteria to declare a signal of activity in pts with solid tumors with MET amp or mut. Table: Baseline Characteristics (N=31); Efficacy Outcomes (n=28); Toxicity Outcomes (N=31) Median (Med) age, years (range) 61 (30, 82) Female, % 16 (52) ECOG PS, % 0 15 (48) 1 12 (39) 2 4 (13) Prior systemic regimens, % 1 3 (10) 2 5 (16) ≥3 23 (74) DC rate, % (OR and SD16+) (1-sided 90% CI) 21 (12, 100) Objective response rate, % (95% CI) 7 (1, 24) Med PFS, wks (95% CI) 8 (8, 13) Med OS, wks (95% CI) 37 (26, 68) Duration of response, wks (n=2) 14 and 20 Med duration SD, wks (n=4) 27 (26, 28) Number of pts1 with tx-related grade 3 adverse or any grade serious adverse event AE2 5 (16) SAE3 3 (10) 1Pts may have experienced one or more events 2ALT increase, diarrhea and all SAEs 3Acute kidney injury, ALT increase, AST increase, blood bilirubin increase, creatinine increase, dehydration, fatigue, GGT increase, hyperkalemia, nausea, sinus bradycardia Citation Format: Kathryn F. Mileham, Michael Rothe, Pam K. Mangat, Elizabeth Garrett-Mayer, Herbert L. Duvivier, Carmen J. Calfa, Carrie L. Dul, Alissa S. Marr, Eugene R. Ahn, Deepti Behl, Michael J. Hall, Inderjit Mehmi, Anu Gaba, Rom Leidner, Mark M. Zalupski, Gina N. Grantham, Abigail Gregory, Susan Halabi, Richard L. Schilsky. Crizotinib (C) in patients (pts) with solid tumors with MET amplification (amp) or mutation (mut): Results from the Targeted Agent and Profiling Utilization Registry (TAPUR) Study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr CT226.

Background: TAPUR is a phase II basket study evaluating anti-tumor activity of commercially available targeted agents in pts with advanced cancers with specific genomic alterations. Results in a cohort of pts with solid tumors with ATM mutation (mut) or deletion (del) treated with O are reported. Methods: Eligible pts had solid tumors, no standard treatment (tx) options, measurable disease, ECOG Performance Status (PS) 0-2, and adequate organ function. Genomic testing was performed in CLIA-certified, CAP-accredited site selected labs. Pts received O tablets (300mg) or capsules (400mg) orally twice daily until disease progression. Low accruing histology-specific cohorts with the same genomic alteration were collapsed into one histology-pooled cohort for this analysis. Primary endpoint was disease control (DC) (complete (CR) or partial (PR) response or stable disease at 16+ wks (SD16+)) (RECIST v1.1). For histology-specific cohorts a Simon 2-stage design with a null DC rate of 15% vs. 35% (power = 0.85; α = 0.10) requires 28 pts with futility stopping after 10 pts. For histology-pooled cohorts with sample size > 28, if the lower limit of a one-sided 90% CI is >15%, the null hypothesis of a DC rate of 15% is rejected. 2-sided 95% CIs were used for other efficacy endpoint estimates. Secondary endpoints were progression-free survival (PFS), overall survival (OS) and safety. Results: 39 pts with solid tumors (17 histologies) with ATM mut (n=36) or del (n=3) were enrolled from 6/2016 to 1/2019. 3 pts were unevaluable for efficacy. Table 1 shows demographics and outcomes. 1 CR (prostate), 2 PR (unknown primary) and 6 SD16+ were observed in pts with ATM mut for a DC rate of 25% (90% CI: 16%, 100%) and an OR rate of 8% (95% CI: 2%, 23%). The null DC rate was rejected. 9 pts had ≥1 Grade 3 tx-related adverse or serious adverse event related to O. Conclusions: Monotherapy O showed evidence of anti-tumor activity in pts with various solid tumors with ATM mut. Table 1. Demographics and Baseline Characteristics (N=39); Efficacy Outcomes (N=36); Toxicity Outcomes (N=39) Median (Med) age, years (range) 65 (35, 77) Female, % 46 ECOG PS, % 0 33 1 59 2 8 Prior systemic regimens, % 0 3 1 8 2 15 ≥3 74 DC rate, % (OR or SD 16+) (1-sided 90% CI) 25 (16, 100) OR rate, % (95% CI) 8 (2, 23) Med PFS, wks (95% CI) 8.4 (8.0, 15.9) Med OS, wks (95% CI) 40.4 (30.3, 50.7) Med duration OR (range), wks 18.9 (4.3, 24.4) Med duration SD16+ (range), wks 27.3 (19.4, 31.0) Number of pts1 with treatment-related adverse or serious adverse events (all Grade 3) AE2 9 SAE3 4 1Patients may have experienced one or more events2anemia, anorexia, dehydration, fatigue, hypokalemia, nausea 3colitis, dizziness, lung infection, proteinuria, urinary tract infection/obstruction Citation Format: Kathryn F. Mileham, Michael Rothe, Pam K. Mangat, Elizabeth Garrett-Mayer, Eddy S. Yang, Olatunji B. Alese, Angela Jain, Herbert L. Duvivier, Phillip Palmbos, Eugene R. Ahn, Jeanny B. Aragon-Ching, Kathleen W. Beekman, Deepti Behl, Funda Meric-Bernstam, Rodolfo Gutierrez, Amy Sanford, Ramya Thota, Michael Zakem, Song Zhao, Raegan O'Lone, Gina N. Grantham, Susan Halabi, Richard L. Schilsky. Olaparib (O) in patients (pts) with solid tumors with ATM mutation or deletion: Results from the Targeted Agent and Profiling Utilization Registry (TAPUR) Study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr CT110.

Background: TAPUR is a phase II basket study evaluating anti-tumor activity of commercially available targeted agents in pts with advanced cancers with genomic alterations. S is an oral multi-kinase inhibitor that inhibits Fibroblast Growth Factor Receptor family members 1-4 (FGFR1-4) in biochemical and cellular assays and is FDA approved in several tumor types. Results in a cohort of mBC pts with FGFR1 mutations (mut) or amplifications (amp) treated with S are reported. Methods: Eligible pts had mBC, no standard treatment (tx) options, measurable disease, ECOG Performance Status (PS) 0-2, and adequate organ function. Genomic testing was performed in CLIA-certified, CAP-accredited site selected labs. Pts received S 50 mg orally daily for four weeks followed by two weeks off, until tumor progression. Simon 2-stage design tested the null disease control (DC) - defined as partial (PR), complete response (CR) or stable disease at 16+ weeks (SD 16+) - rate of 15% vs. 35% (power = 0.85; α = 0.10). If ≥2 of 10 pts in stage 1 have DC, 18 more pts are enrolled. If ≥7 of 28 pts have DC, the null DC rate is rejected. Secondary endpoints are progression-free survival (PFS), overall survival (OS) and safety. Results: Thirty pts with mBC with FGFR1 mut (1 pt), amp (28 pts), or both (1 pt) were enrolled from Oct 2016 to June 2019. 3 were not evaluable and excluded from efficacy analyses. Demographics and outcomes are summarized in Table 1. Two partial responses (PR) and 5 SD16+ (FGFR1 amp only) were observed for DC and OR rates of 29% (95% CI: 13%, 42%) and 7% (95% CI: 1%, 24%), respectively, and the null DC rate of 15% was rejected (p=0.09). S related grade 3-5 TAEs (Table 1) were consistent with the product label for S except encephalopathy. Conclusions: Monotherapy S showed modest anti-tumor activity and clinically significant TAEs in heavily pre-treated pts with mBC with FGFR1 amplification. Table 1.Demographics, Efficacy (N=27) and Toxicity Outcomes (N=30)Median age, yrs (range)61 (28, 81)Female, %97ECOG PS, %047137217Prior systemic regimens, %1-210≥390Hormone Receptor (HR) & HER2 Status, %HR (+) HER2 (-)77HR (-) HER2(-)13HR (+) HER2 (+)7Not reported3DC rate, % (OR or SD16+) (95% CI)29 (13, 42)OR rate, % (95% CI)7 (1, 24)Median PFS, wks (95% CI)8.7 (8.1, 15.7)Median OS, wks (95% CI)33.9 (23.0, 49.0)Number of Pts with Treatment-related AEs/SAEs (TAEs, maximum grade reported)Grade 211Grade 329Grade 4321Skin infection (SAE)2Cytopenia, encephalopathy (SAE), febrile neutropenia (SAE), increased alkaline phosphatase, Palmar-plantar erythrodysesthesia syndrome, vomiting3Cytopenia, hypertension Citation Format: Carmen Calfa, Michael Rothe, Pam K. Mangat, Elizabeth Garrett-Mayer, Eugene Ahn, Keerthi Gogineni, Nitin Rohatgi, Monika L. Burness, Anu Gaba, Omid Hamid, Tareq Albaghdadi, Alison Conlin, Philip Gold, Jordi Rodon, Ramya Thota, Richard L. Schilsky. Sunitinib (S) in patients (pts) with metastatic breast cancer (mBC) with FGFR1 mutations or amplifications: Results from the Targeted Agent and Profiling Utilization Registry (TAPUR) Study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr CT173.

GARP encoded by the Lrrc32 gene is the cell surface docking receptor for latent TGFβ, which is expressed naturally by platelets and regulatory T cells (Treg). Although Lrrc32 is amplified frequently in breast cancer, the expression and relevant functions of GARP in cancer have not been explored. Here, we report that GARP exerts oncogenic effects, promoting immune tolerance by enriching and activating latent TGFβ in the tumor microenvironment. We found that human breast, lung, and colon cancers expressed GARP aberrantly. In genetic studies in normal mammary gland epithelial and carcinoma cells, GARP expression increased TGFβ bioactivity and promoted malignant transformation in immunodeficient mice. In breast carcinoma–bearing mice that were immunocompetent, GARP overexpression promoted Foxp3+ Treg activity, which in turn contributed to enhancing cancer progression and metastasis. Notably, administration of a GARP-specific mAb limited metastasis in an orthotopic model of human breast cancer. Overall, these results define the oncogenic effects of the GARP–TGFβ axis in the tumor microenvironment and suggest mechanisms that might be exploited for diagnostic and therapeutic purposes. Cancer Res; 76(24); 7106–17. ©2016 AACR.

Repetitive stimulation of T-cell receptor (TCR) with cognate antigen results in robust proliferation and expansion of the T cells, and also imprints them with replicative senescence signatures. Our previous studies have shown that life-span and antitumor function of T cells can be enhanced by inhibiting reactive oxygen species (ROS) or intervening with ROS-dependent JNK activation that leads to its activation-induced cell death. Because tumor suppressor protein p53 is also a redox active transcription factor that regulates cellular ROS generation that triggers downstream factor–mediating apoptosis, we determined if p53 levels could influence persistence and function of tumor-reactive T cells. Using h3T TCR transgenic mice, with human tyrosinase epitope–reactive T cells developed on p53 knockout (KO) background, we determined its role in regulating antitumor T-cell function. Our data show that as compared with h3T cells, h3T-p53 KO T cells exhibited enhanced glycolytic commitment that correlated with increased proliferation, IFNγ secretion, cytolytic capacity, expression of stemness gene signature, and decreased TGF-β signaling. This increased effector function correlated to the improved control of subcutaneously established murine melanoma after adoptive transfer of p53-KO T cells. Pharmacological inhibition of human TCR-transduced T cells using a combination of p53 inhibitors also potentiated the T-cell effector function and improved persistence. Thus, our data highlight the key role of p53 in regulating the tumor-reactive T-cell response and that targeting this pathway could have potential translational significance in adoptive T-cell therapy. Cancer Res; 76(18); 5229–40. ©2016 AACR.

Effector CD8+ T cells conditioned with IL12 during activation mediate enhanced antitumor efficacy after adoptive transfer into lymphodepleted hosts; this is due in part to improved IL7 responsiveness. Therefore, we hypothesized that increasing the intensity or type of lymphodepletion would deplete more IL7-consuming host cells and improve the persistence and antitumor activity of IL12-conditioned CD8+ T cells. Using cyclophosphamide, fludarabine, and total body irradiation (TBI, 6 Gy) either individually or in combination, we found that combined lymphodepletion best enhanced T-cell engraftment in mice. This improvement was strongly related to the extent of leukopenia, as posttransfer levels of donor T cells inversely correlated to host cell counts after lymphodepletion. Despite the improvement in engraftment seen with combination lymphodepletion, dual-agent lymphodepletion did not augment the antitumor efficacy of donor T cells compared with TBI alone. Similarly, IL7 supplementation after TBI and transfer of tumor-reactive T cells failed to improve persistence or antitumor immunity. However, IL15 or IL2 supplementation greatly augmented the persistence and antitumor efficacy of donor tumor-reactive T cells. Our results indicate that the amount of host IL7 induced after single agent lymphodepletion is sufficient to potentiate the expansion and antitumor activity of donor T cells, and that the efficacy of future regimens may be improved by providing posttransfer support with IL2 or IL15. Significance: The relationship between lymphodepletion and cytokine support plays a critical role in determining donor T-cell engraftment and antitumor efficacy. Cancer Res; 78(11); 3067–74. ©2018 AACR.

Background: TAPUR is a phase II multi-basket study evaluating the anti-tumor activity of commercially available targeted agents in pts with advanced cancers with genomic alterations known to be drug targets. S is an oral multi-kinase inhibitor that inhibits Fms-like tyrosine kinase-3 (FLT-3) in biochemical and cellular assays. Results from mCRC pts each with FLT-3 amplification treated with S are reported. Methods: Simon’s optimal two stage design was used to test the null hypothesis of 15% response rate versus the alternative of 35%. Power and alpha were set at 85% and 10%, respectively. Response was assessed per RECIST v1.1. This design requires 10 pts in stage 1 and if Results: Ten pts were enrolled from November 2016 to April 2018. Pts received S at a dose of 50 mg orally once daily for four weeks followed by two weeks off. All pts had mCRC with FLT-3 amplification. All pts are included in the data analysis for demographics, safety, PFS and OS (Table 1). No ORs were observed and despite observation of 2 pts with SD at 16 wks, the cohort was closed after further examination revealed both pts with SD died due to disease progression shortly after the 16 wk evaluation. Grades 3-4 AEs at least possibly related to drug are required to be reported. A single grade 3 diarrhea was reported as possibly related to S. Conclusions: Monotherapy with S does not have sufficient clinical activity in mCRC pts with FLT-3 amplification for continued evaluation in this pt population. Other treatments should be considered for these pts, including treatments offered in clinical trials. Table 1:Baseline demographics, clinical characteristics and outcomes by cohortSunitinib targeting FLT-3Tumor TypemCRC (N=10)Median age, yrs (range)56 (41, 71)Male, %80ECOG Performance Status, %01230700Median PFS, wks (90% CI)10.1 (7.4, 15.9)Median OS, wks (90% CI)29.5 (15.7, 39.9)Drug-related AEs, grades 3-4 (% of pts)10 Citation Format: Ricardo H. Alvarez, Elizabeth Garrett-Mayer, Susan Halabi, Pam K. Mangat, Tareq Al Baghdadi, Eugene R. Ahn, Seungjean Chai, Andrew L. Rygiel, Kaitlyn R. Antonelli, Samiha Islam, Suanna S. Bruinooge, Richard L. Schilsky. Sunitinib (S) in patients (Pts) with metastatic colorectal cancer (mCRC) with FLT-3 alterations: Results from the Targeted Agent and Profiling Utilization Registry (TAPUR) Study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr CT146.

Background: The TAPUR Study is a phase II multi-basket study that evaluates the anti-tumor activity of commercially available targeted agents in pts with advanced cancers with genomic alterations known to be drug targets. Results in two cohorts of BC and NSCLC pts each without reported KRAS, NRAS, BRAF mutationstreated with C are reported. Methods: Simon’s optimal two stage design was used to test the null hypothesis of 15% response rate versus the alternative of 35%. Power and alpha were set at 85% and 10%, respectively. Response was assessed per RECIST v1.1. This design requires 10 pts in stage 1 and if Results: Ten pts were enrolled in the BC cohort from June 2016 to May 2018. Ten pts were enrolled in the NSCLC cohort from January 2017 to May 2018. Pts received C at an initial intravenous infusion of 400 mg/m2 over 120 minutes and subsequent weekly infusions of 250 mg/m2 over 60 minutes. All pts are included in the data analysis for demographics, safety, PFS and OS (Table 1). No ORs or SD at 16 wks were observed in either cohort and both were therefore closed. Grades 3-4 adverse events (AEs) at least possibly related to drug are required to be reported. A single grade 3 AE of hypomagnesemia was reported in the NSCLC cohort as possibly related to C. Conclusions: Results from these cohorts suggest monotherapy with C does not have clinical activity in BC or NSCLC pts without reported KRAS, NRAS, BRAF mutations. Other treatments should be considered for these pts, including treatments offered in clinical trials. Table 1:Baseline demographics, clinical characteristics and outcomes by cohortCetuximab targeting tumors without reported KRAS, NRAS, BRAF mutationsTumor TypeBC (N=10)NSCLC (N=10)Median age, yrs (range)59 (45, 65)60 (55, 82)Male, %060ECOG Performance Status, %01260301010900Median PFS, wks (90% CI)6.7 (4.0, 7.9)8.0 (7.6, 8.4)Median OS, wks (90% CI)11.0 (4.9, 30.1)19.6 (8.6, 25.6)Drug-related AEs, grades 3-4 (% of pts)010 Citation Format: Julie Fisher, Elizabeth Garrett-Mayer, Susan Halabi, Pam K. Mangat, Ricardo H. Alvarez, Timothy L. Cannon, Pamela Crilley, Theodore Pollock, Tareq Al Baghdadi, Jared A. Cotta, Andrew L. Rygiel, Kaitlyn R. Antonelli, Samiha Islam, Suanna S. Bruinooge, Richard L. Schilsky. Cetuximab (C) in patients (Pts) with breast cancer (BC) and non-small cell lung cancer (NSCLC) without reported KRAS, NRAS, BRAF mutations: Results from the Targeted Agent and Profiling Utilization Registry (TAPUR) Study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr CT135.

Purpose: Post-mastectomy radiation therapy (PMRT) has been shown to provide improved locoregional control and overall survival in intermediate and high-risk breast cancer patients1,2. To investigate the frequency of utilization and potential barriers to receipt of adjuvant PMRT in South Carolina, an analysis of the S.C. Cancer Registry (SCCR) database was undertaken. Methods: Epidemiological factors such as age, race, and geographic distance from a radiation therapy facility were analyzed as potential predictors of whether patients received the indicated radiotherapy. Post-mastectomy breast cancer patients (n=1080) recorded in the SCCR from 2006–2008 were stratified as low (Tis/T1, N0) (43%), intermediate (T1/T2, N1) (27%), or high (T3/T4, N2/N3) (30%) risk. Logistic regression models were used to predict receipt of adjuvant radiotherapy based on factors such as race, age, and geographical distance from radiation treatment facility. Results: Adjuvant PMRT was received in 4% (19/469) of low risk patients, 23% (67/288) of intermediate risk patients, and 44% (141/323) of high risk patients. In both intermediate and high risk groups, older patients were less likely to receive PMRT. The odds of receiving PMRT decreased by 3% (OR= 1.03; 95% CI: 1.01−1.05; p=0.01) and 2% (OR=1.02; 95% CI: 1.002−1.035; p=0.03) per year in the intermediate and high risk groups, respectively. Race and distance from treatment facility were not predictive of receipt of PMRT in high risk patients. Conclusion: Utilization of adjuvant PMRT in intermediate and high risk breast cancer patients in South Carolina is low, despite the evidence that it confers a survival benefit in both premenopausal and postmenopausal women. Of the patient-related factors analyzed, only age predicted lower odds of receiving radiotherapy in both the intermediate and high risk group. In high risk patients of advanced age (>70), breast cancer continues to be a major cause of mortality which can be improved by radiotherapy3. Future educational efforts should focus on addressing patient and physician bias against radiotherapy based on patient age. Further study is needed to identify socio-economic-, physician- and health system- related factors that maybe impacting the underutilization of PMRT in South Carolina. References: 1. Overgaard, M, PS Hansen, et al. 1997. Postoperative radiotherapy in high-risk premenopausal women with breast cancer who receive adjuvant chemotherapy. NEJM, 337:14, pg 949–955. 2. Overgaard M, Jensen M-J, Overgaard J, et al. 1999. Postoperative radiotherapy in high-risk postmenopausal breast-cancer patients given adjuvant tamoxifen: Danish Breast Cancer Cooperative Group DBCG 82c randomised trial. Lancet. 353: 1641–1648. 3. Smith, BD, BG Haffty, A Hurria, DH Galusha, and CP Gross. 2006. Postmastectomy radiation and survival in older women with breast cancer. JCO, 24:30, pg 4901–4907. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P1-11-05.

Background: NOV-002 (a formulation of disodium glutathione disulfide) modulates signaling pathways involved in tumor cell proliferation and metastasis and enhances anti-tumor immune responsiveness in tumor models. The addition of NOV-002 to a range of cytotoxic chemotherapeutic regimens has been shown to increase their anti-tumor efficacy in several early phase oncology trials and in animal models. Pathological complete response (pCR) has been demonstrated to be associated with favorable overall survival in primary breast cancer, and neoadjuvant treatment of early breast cancer aims at achieving high rates of pCR. In patients with HER-2 (-) breast cancer pCR rates with anthracycline and taxane combinations have been reported to be approximately 10-20% depending on hormone receptor status. We conducted a clinical trial in HER-2 negative patients (pts) combining daily N0V-002 with AC→T. Methods: Women with newly diagnosed stages II-III HER-2 (-) breast cancer received AC x 4 [60/600 mg/m2] followed by T [100 mg/m2] x 4 every 3 weeks in conjunction with daily N0V-002 [60mg IV day 1 and subcutaneously days 2-21 of each cycle]. The primary endpoint is pCR, defined as: (i) ypN0, and (ii) ypT0 or presence of invasive tumor Sample size (n=46 total patients) was calculated using a Simon 2-stage optimal design assuming a doubling of the historical pCR rate with the addition of NOV-002 to AC→T from a p0 of 0.16 to a p1 of 0.32. If a total of 12 or more patients experience a pCR by the end of the trial, then the treatment regimen will be declared active. The calculation assumes an alpha of 0.05 and 80% power. Results: A total of 39 pts have been enrolled to date across three study sites, with 31 patients having completed chemotherapy and undergone surgery. One patient dropped out during cycle 1 and was not assessable for response; 5 are currently receiving chemotherapy; and 2 patients have completed all chemotherapy, but have not yet undergone surgery. A total of 292 chemotherapy cycles have been administered, with 92% of all patients being able to complete all 8 cycles of planned chemotherapy. Of the 31 evaluable patients, 12 achieved a pCR (39%), meeting the primary endpoint of the trial. In patients with residual invasive primary breast tumor Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P1-11-05.

These studies were undertaken to determine the potential role of thromboxane receptors (TP) in bladder cancer. The data reported herein show that expression of the TP-β receptor protein is increased in tissue obtained from patients with bladder cancer and associated with a significantly poorer prognosis (P < 0.005). Bladder cancer cell lines express the TP-β isoform, unlike immortalized nontransformed urothelial cells (SV-HUC) that express only the TP-α isoform. TP-β receptor expression, but not TP-α, promoted cell proliferation, migration, and invasion in vitro, and also resulted in malignant transformation of SV-HUC cells in vivo. Agonist-mediated phosphorylation of extracellular signal-regulated kinase and FAK was dependent on the expression of TP-β. Furthermore, TP-β mediated multiple biological effects by signaling through either G-protein α subunit 12 or β-arrestin 2. Treatment of mice with the TP receptor antagonist GR32191, alone or in combination with cisplatin, significantly delayed tumor onset and prolonged survival of mice transplanted with TCC-SUP bladder cancer cells compared with vehicle or cisplatin alone. These results support the model that the TP-β receptor isoform plays a unique role in bladder cancer progression and its expression may have predictive value and provide a novel therapeutic target. [Cancer Res 2008;68(11):4097–104]

If detected early, breast cancer is eminently curable. To detect breast cancer in samples with little cellularity, a high level of sensitivity is needed. Tumor-specific promoter hypermethylation has provided such a valuable tool for detection of cancer cells in biological samples. To accurately assess promoter hypermethylation for many genes simultaneously in small samples, we developed a novel method, quantitative multiplex-methylation-specific PCR (QM-MSP). QM-MSP is highly sensitive (1 in 104–105 copies of DNA) and linear over 5 orders of magnitude. For RASSF1A, TWIST, Cyclin D2, and HIN1, we observed significant differences in both the degree (P < 0.003) and incidence (P < 0.02) of hypermethylation between normal and malignant breast tissues. Evaluation of the cumulative hypermethylation of the four genes within each sample revealed a high level of sensitivity (84%) and specificity (89%) of detection of methylation. We demonstrate the application of this technique for detecting hypermethylated RASSF1A, TWIST, Cyclin D2, HIN1, and RARB in 50–1000 epithelial cells collected from breast ducts during endoscopy or by lavage. Such an approach could be used in a variety of small samples derived from different tissues, with these or different biomarkers to enhance detection of malignancy.

Sphingosine 1-phosphate (S1P), a pro-proliferative sphingolipid generated by Sphingosine Kinases is upregulated in many cancers. Telomerase (hTERT), is a ribonucleoprotein that extends the ends of chromosomes (telomeres), to promote lung cancer growth. We discovered that SK2 generated S1P binds and stabilizes telomerase in the nuclear periphery by allosterically mimicking protein phosphorylation. Mechanistically, S1P binding protected telomerase from MKRN1 mediated degradation thereby preventing telomere dysfunction and senescence. The objective of this study is to delineate the molecular mechanism of SK2-S1P mediated telomere dysfunction in aging and lung cancer. SK2 knockout mice tissues showed increased telomeric DNA damage and senescence associated-beta galactosidase activity. Moreover, SK2 knockout mice displayed aging phenotypes by reduced subcutaneous fat, atrophy in spleens and hypoplasia in mice testes. Also, SK2 knockout fibroblasts showed increased senescence and robust p16 expression and interestingly, p16 ablation rescued SK2 knockout fibroblasts from senescence. Importantly, we found SK2 and telomerase protein levels to be upregulated in lung cancer tumor tissues (n=48). Mechanistically, pharmacological inhibition of SK2 by ABC294640 (in Phase II clinical trials) leads to telomere dysfunction in A549 cells and interestingly wild type and a phosphomimicking S921D-hTERT mutant of human telomerase overexpression prevented telomere dysfunction whereas S1P defective mutant D684A-hTERT did not. In vivo, inhibition of SK2 by ABC294640 or shRNA knockdown displayed reduced tumor volume, decreased telomerase protein levels and increased TUNEL positive cells. Interestingly, qPCR based gene array revealed robust increase in TCF21 tumor suppressor in stable shSK2 tumors. Furthermore, shRNA against TCF21 following SCID mice xenograft studies showed protection against ABC294640 induced tumor suppression. Interestingly, SK2 inhibition showed ATM/ATR kinase mediated p-CHK1 increase and subsequent activation of caspase-3 in both in vivo and in vitro experiments and stable TCF21 knockdown prevents apoptotic cell death. Importantly, p16 overexpression in p16 deficient A549 lung cancer cells prevented caspase-3 activation and leading to senescence induction following inhibition of SK2-S1P by ABC294640 or shRNA against SK2. Overall, our data suggest that SK2-S1P regulates telomere dysfunction through p16 mediated aging in normal somatic tissues and TCF21 mediated tumor suppression in lung cancer and interestingly SK2-S1P and p16 acts as a molecular switch between senescence and apoptosis in normal and cancer cells respectively. Citation Format: Shanmugam Panneer Selvam, Marion Cooley, Kristi Helke, Elizabeth Garrett-Mayer, Charles D. Smith, Besim Ogretmen. Sphingosine kinase 2/sphingosine 1-phosphate signaling regulates p16 mediated accelerated aging in normal somatic tissues and tcf21 mediated tumor suppression in lung cancer [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 5474. doi:10.1158/1538-7445.AM2017-5474

The epithelial–mesenchymal transition (EMT) and its reversal, mesenchymal–epithelial transition (MET), are fundamental processes involved in tumor cell invasion and metastasis. SEMA3F is a secreted semaphorin and tumor suppressor downregulated by TGF-β1 and ZEB1-induced EMT. Here, we report that neuropilin (NRP)-2, the high-affinity receptor for SEMA3F and a coreceptor for certain growth factors, is upregulated during TGF-β1–driven EMT in lung cancer cells. Mechanistically, NRP2 upregulation was TβRI dependent and SMAD independent, occurring mainly at a posttranscriptional level involving increased association of mRNA with polyribosomes. Extracellular signal—regulated kinase (ERK) and AKT inhibition blocked NRP2 upregulation, whereas RNA interference-mediated attenuation of ZEB1 reduced steady-state NRP2 levels. In addition, NRP2 attenuation inhibited TGF-β1–driven morphologic transformation, migration/invasion, ERK activation, growth suppression, and changes in gene expression. In a mouse xenograft model of lung cancer, NRP2 attenuation also inhibited locally invasive features of the tumor and reversed TGF-β1–mediated growth inhibition. In support of these results, human lung cancer specimens with the highest NRP2 expression were predominantly E-cadherin negative. Furthermore, the presence of NRP2 staining strengthened the association of E-cadherin loss with high-grade tumors. Together, our results demonstrate that NRP2 contributes significantly to TGF-β1–induced EMT in lung cancer. Cancer Res; 73(23); 7111–21. ©2013 AACR.

Background: Cancer patients who self-report tobacco use within the past 30 days are at high risk for relapse and benefit from cessation support. However, there is little data on whether assessment of tobacco use within the past 30 days increases referral rates to a cessation program across cancer disease sites. Methods: Structured tobacco assessments were administered to 426 new adult patients in a Radiation Oncology clinic at a single institution. Multiple logistic regression was used to assess if clinical aspects of cancer diagnosis (age, gender, race, disease site [head/neck or thoracic {HNT} vs. other], stage [advanced vs. early], and purpose [curative vs. palliative]) affected cessation service referral rates identified using assessment of tobacco use within the past 30 days. Results: Complete medical and tobacco use data were compiled for 387 adult cancer patients (91%). Of these, 147 (38%) were women, 259 (67%) were Caucasian, 115 (30%) were black, 177 (46%) were diagnosed with HNT cancer, and 289 (74%) were being considered for curative cancer treatment. Ever smoking was reported in 273 patients (71%), and 83 (21%) reported current smoking everyday or some days. In 180 former smokers who reported time since last cigarette, 21 (12%) reported smoking within with past 30 days and 9 (7%) of these reported smoking within the past day or week. In 21 patients who were identified for cessation support through evaluation of smoking within the past 30 days, 6 (29%) were women, 7 (33%) were black, 15 (71%) were being considered for curative treatment, and 15 (71%) were diagnosed with a HNT cancer. Multiple logistic regression identified that younger age (10 yr OR 2.41, 95% CI 1.47 - 3.98) and HNT cancer (OR 8.13, 95% CI 2.2-30.3) but not gender, race, stage, and purpose were significantly associated with referral based on assessment of tobacco use within the past 30 days. Identification of smoking within the past 30 days increased cessation referrals by 25% from 83 to 114 patients. Conclusions: Assessment of tobacco use within the past 30 days increased referrals to a smoking cessation program from a Radiation Oncology clinic using a standardized tobacco assessment. In clinical trials and practice, including this question has particular benefit for younger patients and patients with head/neck or thoracic tumors. Citation Format: Samuel Lewis Cooper, Katherine Hoover, Elizabeth Garrett-Mayer, Kenneth Michael Cummings, Matthew J. Carpenter, Kelly Crowley, David T. Marshall, Graham W. Warren. Assessment of tobacco use within 30 days increases referral for cessation support. [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 3736. doi:10.1158/1538-7445.AM2015-3736

A subset of triple negative breast cancer (TNBC) is characterized by overexpression of the epidermal growth factor receptor (EGFR) and loss of PTEN, and patients with these factors have a poor prognosis. In addition, approximately half of all basal breast cancers show autocrine activation of EGFR by amphiregulin (AREG), resulting in altered receptor trafficking and downstream signaling. However, despite the strong rationale for targeting EGFR in the treatment of TNBC, EGFR inhibitors have performed poorly in clinical trials. To further analyze the signaling processes active in this subset of TNBC, we performed reverse-phase protein array (RPPA) analysis on SUM149 cells, a highly tumorigenic and metastatic PTEN-null cell line that expresses high levels of AREG and EGFR, and MCF10A cells, a non-transformed mammary epithelial cell line cultured in the presence of either EGF or AREG. This analysis identified a number of signaling components mediated specifically by amphiregulin, including marked changes in integrin expression and increased levels of fibronectin, which is known to be a mediator of invasive capacity via interaction with integrin β1. RPPA analysis also revealed elevated AKT activity in SUM-149 cells, which was not surprising given the lack of PTEN expression. Interestingly, this AKT activity is not abrogated by treatment with the EGFR inhibitor gefitinib, nor by treatment with PI3K inhibitors. These findings and the results of preliminary experiments in SUM149 cells re-expressing PTEN suggest that AKT activity in these cells is, at least partly, independent of EGFR and PI3K activity. In addition, expression profiling demonstrated that AREG-activated EGFR regulates gene expression differently than EGF-activated EGFR, and functional analysis via genome-scale shRNA screening identified a set of genes, including PLK1 and BIRC5, that are regulated by EGFR in MCF10A cells but uncoupled from EGFR signaling in SUM149 cells, although they are essential for the survival of SUM-149 cells. Thus, our results demonstrate that in cells with constitutive EGFR activation and PTEN loss, critical survival genes are uncoupled from regulation by EGFR, a mechanism that is likely to mediate resistance to EGFR inhibitors. Citation Format: Christiana S. Kappler, Stephen T. Guest, Ericka L. Smith, Jonathan C. Irish, Elizabeth Garrett-Mayer, Zachary Kratche, Stephen Ethier. Oncogenic signaling in amphiregulin- and EGFR-expressing PTEN-null human breast cancer. [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 1949. doi:10.1158/1538-7445.AM2015-1949

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA Objectives: Primary aim of the ongoing study ([NCT01898091][1]) is to determine whether a mouthrinse containing extract of neem leaf (Azadirachta indica A. Juss.), a tropical evergreen tree with anti-inflammatory and anti-microbial medicinal properties, will lessen the severity of oral mucositis (OM) in patients undergoing radiation therapy (RT) to the head and neck. Secondary aims are to assess the effects of the neem mouthrinse on the microbial environment of the oral cavity and on quality of life. Methods: Design is a double-blind, controlled, randomized, parallel-group Phase II clinical trial at a single NCI designated center. Block-randomization was used for patient assignment, stratified by tobacco use. Eligibles were adults with histologically confirmed head and neck cancer (HNC) whose treatment includes RT for 7 weeks. HNC includes malignancy of the oral cavity, oro-pharynx and larynx (ICD-9 codes 140 - 149, 161; ICD-O morphology code of 2 or 3). Exclusion criteria include prior HNC radiotherapy, baseline mouth and throat soreness (MTS) score of >3, ECOG performance status >2, allergy or inability to use mouthrinse, and language barrier. Evaluable participants receive > 40 Gy RT and participate to week 6 RT. Study duration is 12 weeks with visits at baseline, weekly during RT, 3 telephone visits post RT, and one-month follow-up visit. Data were collected using the Oral Mucositis Daily Questionnaire, Functional Assessment of Cancer Therapy-Head and Neck, and Symptom Distress Scale questionnaires; CariScreen for oral microbial load, and ELISA and flow cytometry for salivary analyte measures. Results: Of 53 patients enrolled, 3 screen failed and 8 withdrew leaving 42 with evaluable data. Neem (n = 23) and placebo (n = 19) groups were not statistically different for demographic and clinical variables. Major hypothesis assessed as the maximum change in MTS score from baseline during 6 weeks of RT resulted in a larger change, 1.91 with SD 1.34 for neem group vs. 1.71 with SD 1.29 for placebo group based on a Wilcoxon rank sum test with one-sided alpha = 0.05. Preliminary results suggest no difference in the maximum change in severity from baseline (p = 0.85). Neem group had higher adherence to mouthrinse protocol measured as ≥4 days mouthrinse use per week for six weeks (OR 2.56, p = 0.19). Additional outcomes of ongoing comparisons across groups include time to maximum OM severity, time to onset of OM, percent of patients with MTS scores

During the course of tumorigenesis, cancer cells acquire a large number of genetic and epigenetic alterations. A subset of these alterations results in the generation of driver oncogenes that play critical roles in initiating and maintaining the transformed phenotype. We hypothesize that most solid tumors are simultaneously driven by multiple driver oncogenes that together form an oncogene network and that durable therapies for solid tumors will need to target multiple oncogenes. Identifying multiple driver oncogenes from among the large number of mutated/amplified genes present in most solid tumors is a nontrivial task that will require novel approaches. In this work, we have combined genome-wide, RNAi-based dropout screens with copy number, expression and exome sequencing analysis to identify the full complement of driver oncogenes that function in a panel of breast cancer cell lines. The panel of cell lines used for this study is the SUM panel of breast cancer cells which contains 10 cell lines and includes representatives of HER2 positive, ER positive, and triple negative breast cancer subtypes. For each line we have identified multiple activated driver oncogenes that when combined define a complete oncogene signature for the cell line. All of the oncogene signatures contain both well-characterized oncogenes that are known to play a role in breast cancer as well as novel driver genes that do not have a well-characterized role in breast cancer. Subsequent work has confirmed the role of several of the novel driver oncogenes including the epigenetic modifier KAT6A and the apoptotic regulator BCL2L1. Additionally, we show that the cell line oncogene signatures can be used to successfully predict sensitivity of individual cell lines to small molecule inhibitors. Predictions of sensitivity to inhibitors were confirmed through the integration of oncogene signatures with published data from large-scale drug screens across numerous cancer cell lines. Finally, we demonstrate the utility of oncogene signatures through their use in the design of novel therapeutic strategies that effectively reduce cancer cell viability. Citation Format: Stephen Guest, Zachary Kratche, Jonathan Irish, Aliccia Bollig-Fischer, Elizabeth Garrett-Mayer, Stephen Ethier. Oncogene signatures identify novel therapeutic targets and combinatorial strategies in breast cancer. [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 1116. doi:10.1158/1538-7445.AM2015-1116

The ability of cancers to evade immune surveillance and resist immunotherapy raises a fundamental question of how tumor cells survive in the presence of a competent immune system. Studies to address this question have primarily focused on mechanisms by which tumor cells avoid recognition by or induce tolerance in the immune system. However, little is known about whether cancer cells also acquire an intrinsic ability to resist killing by immune effectors. We find that cancer cells enhance their ability to withstand an attack by cytotoxic immune effector cells via acquisition of specific genetic alterations that interfere with the shared mitochondrial death signaling pathway entrained by granzyme B, IFN-γ, and Apo2 ligand/tumor necrosis factor–related apoptosis inducing ligand (Apo2L/TRAIL), three key mediators of immunologic cell–mediated cytotoxicity. We show that the coexistence of specific mitochondrial signaling defects (either deletion of Bax, overexpression of Bcl-xL, or deletion of Smac) with expression of X-linked inhibitor of apoptosis protein decreases the sensitivity of cancer cells to IFN-γ/Apo2L/TRAIL– or granzyme B–induced apoptosis, lymphocyte-mediated cytotoxicity in vitro, and adoptive cellular immunotherapy in vivo. Conversely, negating X-linked inhibitor of apoptosis protein expression or function in tumor cells with defective mitochondrial signaling enables direct activation of caspase-3/-7 by granzyme B or Apo2L/TRAIL, and restores their susceptibility to immunologic cytotoxicity. These findings identify an important mechanism by which cancers evade elimination by immune effector cells and suggest that cancer immunotherapy might be improved by concurrent strategies to alleviate or circumvent the intrinsic mitochondrial death signaling defects that help cancer cells resist immunologic cytotoxicity. (Cancer Res 2006; 66(3): 1730-9)

In cancer patients and in those at high risk, systemic exposure to agents for therapy or prevention is accompanied by undesirable side effects. We hypothesized that it is possible to prevent and treat breast cancer by introducing anticancer agents into the mammary ductal network. Here, we show the efficacy of intraductally administered anticancer agents 4-hydroxytamoxifen and pegylated liposomal doxorubicin (PLD) in the prevention and treatment of breast cancer using the rat N-methyl-N′-nitrosourea–induced and spontaneous HER-2/neu transgenic mouse (neu-N) models of breast cancer. Intraductal administration of PLD to neu-N mice caused regression of established tumors and prevented tumor development more effectively than i.v. injection (P < 0.0001). Intraductal administration resulted in lower circulating levels of PLD compared with i.v. administration, with no evidence of systemic toxicity or long-term histopathologic changes in the mammary gland. Compared with systemic administration, intraductal injection provides direct access to breast lesions with higher local and lower systemic drug exposure. These studies suggest that this approach has potential for application to prevention and neoadjuvant therapy of early breast cancer. (Cancer Res 2006; 66(2): 638-45)

The majority of colorectal cancers have lost/inactivated the p53 tumor suppressor gene. Using isogenic human colon cancer cells that differ only in their p53 status, we demonstrate that loss of p53 renders tumor cells relatively resistant to the topoisomerase I inhibitor, irinotecan. Whereas irinotecan-induced up-regulation of the proapoptotic proteins PUMA and Noxa requires p53, we find that irinotecan inhibits Janus kinase 2 (JAK2)-signal transducer and activator of transcription 3 and 5 (STAT3/5) signaling in both p53-proficient and p53-deficient tumor cells. We show that irinotecan inhibits JAK2-STAT3/5-dependent expression of survival proteins (Bcl-xL and XIAP) and cooperates with Apo2 ligand/tumor necrosis factor-related apoptosis-inducing ligand (Apo2L/TRAIL) to facilitate p53-independent apoptosis of colon cancer cells. Whereas xenografts of p53-deficient colon cancer cells are relatively resistant to irinotecan compared with their p53-proficient counterparts, combined treatment with irinotecan and Apo2L/TRAIL eliminates hepatic metastases of both p53-proficient and p53-deficient cancer cells in vivo and significantly improves the survival of animals relative to treatment with either agent alone. Although the synergy between chemotherapy and Apo2L/TRAIL has been ascribed to p53, our data demonstrate that irinotecan enhances Apo2L/TRAIL-induced apoptosis of tumor cells via a distinct p53-independent mechanism involving inhibition of JAK2-STAT3/5 signaling. These findings identify a novel p53-independent channel of cross-talk between topoisomerase I inhibitors and Apo2L/TRAIL and suggest that the addition of Apo2L/TRAIL can improve the therapeutic index of irinotecan against both p53-proficient and p53-deficient colorectal cancers, including those that have metastasized to the liver.

The tumor suppressor FOXO3 is a transcription factor whose activity and stability are regulated by phosphorylation on S294 via extracellular regulated kinases (ERK1/2). Previous data by our group has demonstrated overexpressing the thromboxane A2 isoform-β receptor (TPβ) increased ERK activation and induced malignant transformation of immortalized bladder cells. These data describe a novel mechanism of TPβ mediated regulation of FOXO3 via phosphorylation by ERK. Immunohistochemical studies revealed increased expression of nuclear pFOXO3-S294 in late stage bladder tumors, whereas total FOXO3 nuclear expression was reduced in high grade tumors. Both cytoplasmic and nuclear pFOXO3-S294 staining scores positively correlated with TPβ staining scores. Overexpressing TPβ in non-transformed urothelial cell lines, UROsta and SV-HUC, induced phosphorylation of FOXO3 at -S294. Knockdown of FOXO3 expression by shRNA increased cell migration and invasion. Conversely, overexpression of mutated FOXO3 that is resistant to ERK phosphorylation reduced the overall and TP agonist (U46619) mediated increase in malignant UMUC3 cell migration and invasion. Stimulation of UMUC3 cells with U46619 increased pFOXO3-S294 expression which could be attenuated with TP antagonist (PTXA2) pre-treatment. A dual luciferase assay demonstrated that U46619 stimulation decreased overall FOXO3 transcriptional activity. Additionally, U46619 stimulation initially caused nuclear accumulation of pFOXO3-S294, but prolonged stimulation increased FOXO3 cytoplasmic localization. In summary, the data describe a novel mechanism of TPβ signaling in bladder cancer that results in the ERK mediated regulation of FOXO3 activity, which could contribute to tumor progression. Citation Format: Philip M. Sobolesky, Perry V. Halushka, Elizabeth Garrett-Mayer, Omar Moussa. Thromboxane-A2 receptor β-isoform regulates the activity of the tumor suppressor FOXO3 via phosphorylation by ERK in urothelial 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 3392. doi:10.1158/1538-7445.AM2014-3392

Large-scale efforts such as The Cancer Genome Atlas (TCGA) have identified the landscape of genetic, epigenetic and proteomic alterations in breast cancer through analysis of ∼1000 primary breast tumors. Importantly, all of the samples analyzed by TCGA also come with long-term clinical follow-up data that allows for examination of how identified alterations affect patient outcomes. Although powerful, the sheer number of alterations identified by TCGA and similar efforts makes it difficult to separate alterations that drive cancer from those that are merely passengers. We have been combining data from TCGA with data from genome-wide, RNAi-based, functional genetic screens that we have been performing on a panel of breast cancer cell lines that were developed in our lab. This combination of functional genetic data with data from the TCGA has resulted in the identification of genes that are responsible for driving breast cancer cell growth and survival that also affect patient outcomes. Using this approach, we have identified the TRiC chaperonin protein complex as playing a critical role in driving breast cancer and affecting patient outcomes. In the SUM-52 breast cancer cell line, a member of the TRiC complex known as TCP-1 was a hit in the genome wide, RNAi-based screen and examination of TCP1 in the TCGA database revealed that it is altered in ∼9% (85 of 929 cases) of breast cancers and patients with TCP1 alterations have significantly reduced long-term survival (pval Citation Format: Stephen Guest, Jon Irish, Zachary Kratche, Elizabeth Garrett-Mayer, Robert Wilson, Stephen Ethier. The TRiC chaperonin in breast cancer cell growth, survival and response to taxane therapy. [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 440. doi:10.1158/1538-7445.AM2014-440

The selection of clinical endpoints when designing a clinical trial is critically important to ensure appropriate evaluation of the agent or regimen of interest. Endpoints in early stage clinical trials differ from those in later stage trials, and the nature of the endpoints (e.g., continuou,s categorical) has an impact on both the parameters that can be estimated and the power to detect significant differences. This talk will focus on the selection of continuous (e.g. Ki67, breast density, change in tumor size), categorical (e.g. RECIST response, pathologic complete response), and time to event endpoints (e.g. progression-free survival, overall survival) when designing clinical trials. Discussion will also include interpretation of resulting measures of clinical efficacy (e.g. hazard ratios), approaches for stating trial objectives and implications of endpoint selection on statistical power. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr ES03-3.

Abstract #5145 Purpose: To describe 4-year local and ipsilateral breast recurrence-free intervals, overall freedom from disease failure, and disease-specific and overall survivals in a cohort of MammoSite Brachytherapy (MBT) patients with mature follow-up, treated at a single institution over a 6-year period. Methods and Materials: An analysis of MBT cases was performed using prospectively collected quality assurance database, departmental chart review, and electronic medical records. Patient-, tumor-, treatment-, and outcome-specific data were extracted and recorded into a research database. Eligible patients were required to have >6 months post-MBT follow-up. Local tumor failure was defined as pathologically-proven recurrence within 2 cm of the resected tumor bed, with all other ipsilateral breast recurrences (excluding axilla or internal mammary nodal failures) defined as ipsilateral breast failures. Any local, ipsilateral breast, nodal, or distant failure was censored for overall freedom from failure. Results: From 2004 through 2008, 115 MBT patients have been treated, of which 104 were eligible for the present analysis. Median patient age was 62 years (range 25–86). Primary tumors included ductal carcinoma in situ (DCIS) for 20 patients, T1a (12), T1b (31), T1c (34), and T2 (7). Lymph nodes were assessed in 94 patients (including all patients with invasive carcinomas), and revealed pN0 disease in 85 patients, pN1(mic) (3), and pN1a (6). Histologies of invasive carcinomas were ductal in 68 patients, lobular (3), mixed ductal/lobular (3), mucinous/colloid (7), medullary (2), and tubular (1). At a median follow up of 45.7 months (range 7.3-71.4), 6 patients have experienced disease recurrence, including 4 within the ipsilateral breast. Three patients with ipsilateral breast recurrence underwent salvage mastectomy, two of whom remain without evidence of disease recurrence at 56 and 14 months, respectively. Seven patients have died, 3 of or with recurrent breast cancer. Estimated 4-year outcomes for the entire cohort were: local tumor control 100%, ipsilateral breast control 97.0% (95% CI, 93.6%-100%), overall freedom from failure 93.8% (88.5%-99.5%), disease specific survival 97.8% (94.8%-100%) and overall survival 92.7% (87.2%-98.6%). Conclusions: The present study demonstrates low rates of local and ipsilateral breast disease failure in a well-defined cohort of MBT patients with mature follow-up. Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 5145.

Abstract #2141 Background: Chemotherapy induced anemia is a common problem in breast cancer. Anemic events [Hgb Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2141.