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Purpose ASCO and the College of American Pathologists (ASCO-CAP) recently recommended further changes to the evaluation of human epidermal growth factor receptor 2 gene (HER2) amplification by fluorescent in situ hybridization (FISH). We retrospectively assessed the impact of these new guidelines by using annotated Breast Cancer International Research Group (BCIRG) -005, BCIRG-006, and BCIRG-007 clinical trials data for which we have detailed outcomes. Patients and Methods The HER2 FISH status of BCIRG-005/006/007 patients with breast cancers was re-evaluated according to current ASCO-CAP guidelines, which designates five different groups according to HER2 FISH ratio and average HER2 gene copy number per tumor cell: group 1 (in situ hybridization [ISH]–positive): HER2-to-chromosome 17 centromere ratio ≥ 2.0, average HER2 copies ≥ 4.0; group 2 (ISH-positive): ratio ≥ 2.0, copies < 4.0; group 3 (ISH-positive): ratio < 2.0, copies ≥ 6.0; group 4 (ISH-equivocal): ratio < 2.0, copies ≥ 4.0 and < 6.0; and group 5 (ISH-negative): ratio < 2.0, copies < 4.0. We assessed correlations with HER2 protein, clinical outcomes by disease-free survival (DFS) and overall survival (OS) and benefit from trastuzumab therapy (hazard ratio [HR]). Results Among 10,468 patients with breast cancers who were successfully screened for trial entry, 40.8% were in ASCO-CAP ISH group 1, 0.7% in group 2; 0.5% in group 3, 4.1% in group 4, and 53.9% in group 5. Distributions were similar in screened compared with accrued subpopulations. Among accrued patients, FISH group 1 breast cancers were strongly correlated with immunohistochemistry 3+ status (P < .0001), whereas groups 2, 3, 4, and 5 were not; however, groups 2, 4 and, 5 were strongly correlated with immunohistochemistry 0/1+ status (all P < .0001), whereas group 3 was not. Among patients accrued to BCIRG-005, group 4 was not associated with significantly worse DFS or OS compared with group 5. Among patients accrued to BCIRG-006, only group 1 showed a significant benefit from trastuzumab therapy (DFS HR, 0.71; 95% CI, 0.60 to 0.83; P < .0001; OS HR, 0.69; 95% CI, 0.55 to 0.85; P = .0006), whereas group 2 did not. Conclusion Our findings support the original categorizations of HER2 by FISH status in BCIRG/Translational Research in Oncology trials.

Background The optimal regimen for adjuvant breast cancer chemotherapy is undefined. We compared sequential to concurrent combination of doxorubicin and cyclophosphamide with docetaxel chemotherapy in women with node-positive non-metastatic breast cancer. We report the final, 10-year analysis of disease-free survival (DFS), overall survival (OS), and long-term safety. Patients and methods A total of 3298 women with HER2 nonamplified breast cancer were randomized to doxorubicin and cyclophosphamide every 3 weeks for four cycles followed by docetaxel (AC → T) every 3 weeks for four cycles or docetaxel, doxorubicin, and cyclophosphamide (TAC) every 3 weeks for six cycles. The patients received standard radiotherapy and endocrine therapy and were followed up for 10 years with annual clinical evaluation and mammography. Results The 10-year DFS rates were 66.5% in the AC → T arm and 66.3% in the TAC arm (P = 0.749). OS was 79.9% in the AC → T arm and 78.9% in the TAC arm (P = 0.506). TAC was associated with higher rates of febrile neutropenia, although G-CSF primary prophylaxis greatly reduced this risk. AC → T was associated with a higher rate of myalgia, hand-foot syndrome, fluid retention, and sensory neuropathy. Conclusion This 10-year analysis of the BCIRG-005 trial confirmed that the efficacy of TAC was not superior to AC → T in women with node-positive early breast cancer. The toxicity profiles differ between arms and were consistent with previous reports. The TAC regimen with G-CSF support provides shorter adjuvant treatment duration with less toxicity. Trial Registration ClinicalTrials.gov Identifier NCT00312208.

Background: Adjuvant trastuzumab (TRZ) is the standard of care for HER2-overexpressing (HER2+) early stage breast cancer (EBC) patients (PTS), with five-year survival rates exceeding 90%. However, significant cardiac toxicities are observed, with a fivefold increase in clinical heart failure (HF). Left ventricular (LV) remodeling (increased heart size and mass) is an early indicator of cardiac injury, progressing to further LV dysfunction, reduced exercise tolerance and overt HF. Therefore, effective prevention of such negative sequelae is of enormous clinical interest. As the pivotal TRZ trials assessed cardiac function with MUGA or echocardiography, insensitive modalities to evaluate LV remodeling, the long-term sequelae of TRZ remain unknown. Objective: to determine the long term effects of trastuzumab on cardiopulmonary and left ventricular function in women with HER2 overexpressing breast cancer. Additionally as aerobic training is an effective intervention in HF PTS who adhere to prescribed exercise, a sub-analysis compared those who adhered during a 4 month exercise intervention (AEX) vs those who did not adhere (NEX). Methods: 16 PTS (mean age 58 ± 7) who participated in an exercise intervention study during the first 4 months of TRZ therapy were recruited, with an average of 4 years elapsing since TRZ completion. Cardiopulmonary exercise (VO2peak) testing and resting cardiac MRI (CMR) were performed and compared with baseline and 4 month assessments. Adherence to exercise intevention was defined as attendance ≥80% prescribed sessions. Results: All 16 PTS reported independent living with no limitations to ADLs. At 4 years, mean VO2peak for all PTS was 22.4 ml/kg/min (20.0 at baseline and 22.0 at 4 months). In AEX PTS, higher VO2peak persisted 4 years after cessation of therapy, 4.1 mL/kg/min higher than NEX PTS (24.9 and 20.8 mL/kg/min, respectively ). Mean LVEF for all PTS was 60 ± 6%, not significantly different from baseline or 4 months (61 ± 5 and 55 ± 4%, respectively) . Statistically significant interactions of exercise adherence to other CMR metrics were not observed. Conclusions: Clinically significant impairment of cardiopulmonary function (equal to 14 years of aging) are present before therapy and persist in PTS four years following exposure to TRZ-based chemotherapy. This observation is consistent with our other work, and occurs on a background of normal LVEF, implying additional negative effects to other components of the oxygen cascade. As mortality risk has been shown to decrease by 17% for every 3.5 mL/kg/min difference in aerobic capacity in healthy females, these findings indicate adherence to exercise interventions during TRZ-based therapy has potentially important long-term implications. Citation Format: Pituskin E, Paterson I, Ghosh S, Mackey JR, Haykowsky MJ. Long term effects of trastuzumab on cardiopulmonary and left ventricular function in women with HER2 overexpressing breast cancer. [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 P4-10-16.

Background: Genetic predisposition to docetaxel (Doc) toxicity contributes to unacceptable toxicity and reduced dose intensity, and may influence disease outcomes. We previously reported variants associated with Doc toxicity in candidate gene single nucleotide polymorphism (SNP) associations in a breast cancer treatment setting (Damaraju et al. Eur J Cancer (suppl); Vol 8 (7), page 175, 2010) and the identified variants were confirmed in an independent validation study (Damaraju et al, J Clin Oncol Vol 33, Issue 15 suppl, 2015: 540). Others have reported candidate SNP (Breast Cancer Res Treat, 2011 SWOG 0221 study) and GWAS (Clin Cancer Res 2012 CALGB 40101 study) identified variants associated with paclitaxel mediated peripheral neuropathy. However, the overlap on the variants identified thus far between Doc and paclitaxel are limited, prompting a genome wide search to find variants contributing to Doc specific toxicity. Methods: TRIO-012 is a double blinded, multinational trial that randomized 1,144 patients with advanced breast cancer to receive first-line Doc in combination with ramucirumab (RAM) or placebo (Mackey et al. J Clin Oncol Jan 10, 2015:141-148). Study subjects (n=719) in the Doc+RAM or Doc+placebo arm with available germline DNA are being genotyped; all subjects provided ethics-committee approved prospective consent for this genetic study. Genotyping are being performed with Affymetrix SNP 6.0 arrays. Genotype data will be filtered for deviations from Hardy Weinberg Equilibrium and minor allele frequency of >0.05. Doc-induced adverse events (AEs) are based on CTCAE (Common Terminology Criteria for Adverse Events v4.1) toxicity grades. Toxicities >2 scored for fatigue (n=96), myalgia (n=22), peripheral neuropathy (n=17) will be analysed as individual phenotypes in comparison with the no toxicity group (toxicity grades 0-1) and in a combined analysis of all Doc induced toxicities (0-1, n=599 vs. >2, n=120). Dominant genotypic model is assumed; Chi-square test, FDR and/or 10000 permutations were employed using SVS v8.3 and p Results and conclusions: We expect to reconfirm the associations of loci reported in candidate SNP and previous GWAS studies; XKR4 (rs4737264) for peripheral neuropathy, CYP3A5*3 (rs776746) with fatigue, and FACND2 (rs7637888) with myalgia in addition to the potential novel variants distinct from paclitaxel AE GWAS studies. Fine mapping around these loci may help identify potential causal variants. Both candidate SNP and GWAS identified variants may aid in developing risk stratification models. The GWAS identified loci and the flanking genes will be interrogated using the ingenuity pathway analysis for insights in to the biological roles in the drug metabolism. We expect to complete the analysis by mid-November 2015. Citation Format: Damaraju S, Gorbunova V, Gelmon K, García-Saenz J, Morales-Murillo S, AbiGerges D, Canon J-L, Kiselev I, Cohen GL, Jerusalem G, Thireau F, Fresco R, Houé V, Press MF, Kumaran M, Mackey JR. Genome wide association study (GWAS) of genetic variants associated with docetaxel toxicity in the ROSE/TRIO-012 trial. [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 P1-13-03.

Background: In a candidate single nucleotide polymorphism (SNP) association study, we previously identified VEGFR-1 and VEGFR-2 SNPs strongly associated with treatment emergent hypertension (HT) in the ramucirumab (RAM) and docetaxel (Doc) arm in the ROSE/TRIO-012 study, a double-blinded multinational phase III trial that randomized 1,144 patients with advanced breast cancer to receive first-line Doc in combination with RAM or placebo (Mackey et al, JCO Jan 10, 2015:141-148; Mackey et al, JCO, Volume 33, Issue 15_suppl, May 20, 2015: 547). However, candidate SNP studies limit the number of genes for interrogation and a more comprehensive genome wide search may identify critical variants associated with the phenotype of HT. Preliminary analysis indicated that patients experiencing HT with RAM showed better overall survival (Mackey JR, et al (2015). Reply to H. Lee, et al. JCO; in press). These observations provide the potential to identify those patients with genetic variants for predisposition to RAM-associated HT to inform therapeutic decisions. Methods: Genotyping of samples is underway using Affymetrix SNP 6.0 arrays. Genotype data will be filtered for deviations from Hardy Weinberg Equilibrium and minor allele frequency of >0.05. Study subjects (n=792) provided ethics-committee approved prospective consent for this genetic study of whom 478 subjects were allocated RAM + Doc arm. Toxicity grades 0-1 (n= 394 controls; low toxicity) vs. grade >2 (n= 84 cases, high toxicity) is our binary outcome. Dominant genotypic model is assumed. Chi-square test, FDR and/or 10000 permutation tests will be employed (Golden Helix-SVS v8.3) and p Results and conclusions: We expect up to 700,000 SNPs to be retained after filtering based on our previous breast cancer GWAS analyses (Damaraju et al. Cancer Research (suppl); Vol 70 (24), page 258s, 2010 and Sehrawat et al Hum Genet. 2011 Oct;130(4):529-37) and 30,000 SNPs to show significance at a nominal p-value (0.05); these will be analysed for regions of high linkage disequilibrium to narrow down potential loci showing association with HT to serve as candidate markers in further independent validation studies. Cumulative dose to adverse events will be considered in the analysis. Identified loci will be interrogated for potential genes in the flanking regions with biological relevance based on pathway analysis. Identified variants from candidate SNP and GWAS may allow developing predictive tools to enable stratification of patients for therapies. The analysis is expected to be completed by mid- November, 2015. Citation Format: Mackey JR, Lipatov O, Martín M, Webster M, Hegg R, Verma S, Ramos-Vázquez M, Fresco R, Thireau F, Houé V, Press MF, Kumaran M, Damaraju S. Genome wide association study (GWAS) to identify variants conferring ramucirumab-associated hypertension in the ROSE/TRIO-012 breast cancer trial. [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 P6-14-03.

Background: Facilitating healthy levels of physical activity (PA) during chemotherapy is important for the psychosocial and physical health of breast cancer survivors. The primary objective of this feasibility study was to examine the effects of a broad-reach PA behavior change intervention among women with breast cancer receiving adjuvant chemotherapy. Methods: Breast cancer patients receiving adjuvant chemotherapy (N = 95) were randomly assigned to receive a PA resource kit consisting of tailored print materials and a step pedometer (intervention) or a standard public health PA recommendation (standard recommendation). The primary outcome was daily pedometer steps. Secondary outcomes were self-reported light, moderate, and vigorous intensity PA, total moderate-to-vigorous PA, and sedentary time. Assessments were conducted before and after adjuvant chemotherapy. Results: Attrition was 19% (17 of 95). Intervention patients wore their step pedometer for 85 days (range, 35–144 days; SD = 26.4) for a 95% adherence rate. Analyses of covariance suggested that the intervention was not statistically superior to standard recommendation for daily average pedometer steps (−771; 95% CI = −2024 to 482; P = 0.22), total MVPA minutes (−4; 95% CI = −62 to 570; P = 0.90), or sedentary time (+160; 95% CI = −186 to 506; P = 0.42). Conclusion: This broach-reach and low intensive intervention was not more effective for promoting PA in breast cancer patients receiving chemotherapy than providing the standard public health guidelines for PA. Impact: Achieving physical activity behavior change during adjuvant breast cancer chemotherapy may require some level of supervised physical activity or more intensive (e.g., face-to-face, supervised) interventions. Cancer Epidemiol Biomarkers Prev; 25(2); 391–8. ©2015 AACR.

Myristoylation is the N-terminal modification of proteins with the fatty acid myristate. This process is mediated by two ubiquitously expressed N-myristoyltransferases, NMT1 and NMT2, and is critical for membrane targeting and cell signaling. Because NMT expression is increased in some cancers, we used three robotic screens to evaluate the potential of the potent pan-NMT inhibitor PCLX-001 on 300 cancer cell lines spanning the spectrum of human cancers. We discovered a marked increase in the sensitivity of hematological cancer cell lines, including B-cell lymphomas, to myristoylation inhibition. PCLX-001 consistently reduced both lymphoma cell proliferation and viability at concentrations lower than those needed to inhibit the growth of or to kill benign immortalized B cells. In lymphoma cell lines, PCLX-001 treatment inhibited early B-cell receptor (BCR) signaling events by disrupting membrane targeting of several myristoylated Src family kinases and promoted their ubiquitin-mediated degradation. Unexpectedly, PCLX-001 also promoted the degradation of non-myristoylated transcriptional activators P-ERK, c-Myc, NFκB and CREB downstream in the BCR signaling cascade, leading to loss of survival signals and apoptosis. Furthermore, compared to clinically approved drugs dasatinib and ibrutinib, PCLX-001 was more potent in vitro at inhibiting B-cell signaling, had a wider breadth of efficacy, and had greater selectivity thus sparing normal B cells. PCLX-001 treatment reduced tumor size in a time and concentration dependent manner in three B-cell lymphoma xenograft models and resulted in complete disease regression in two of these models, including an R-CHOP refractory lymphoma patient-derived xenograft. To investigate the potential mechanisms responsible for the sensitivity of hematological cancers to PCLX-001, we examined the NMT expression levels in cancer cells using publically available databases. Contrary to the reported NMT overexpression in some cancers, we found that hematological cancer cell lines and tumors both display significant reduction in NMT2 expression. The decreased NMT2 expression is significantly correlated with lower EC50 and poorer patient prognosis. Using the CRISPR-based genetic alteration Cancer Dependency Map, we discovered that cancer cells are highly dependent on functional NMT1, and that NMT1 dependency increases with low NMT2 expression. PCLX-001 treatment may mimic the effect of genetic alteration of NMT1 in hematological cancer cells low in NMT2 by pharmacologically inhibiting the remaining NMT1 in these cells. This results in an effect reminiscent of synthetic lethality since the vast majority of normal cells express both NMTs and PCLX-001 selectively kills NMT2-deficient cancer cells while sparing normal cells. Our findings support the ongoing development and eventual clinical trials of PCLX-001 as a therapy for hematological cancers. Citation Format: Erwan Beauchamp, Megan C. Yap, Maneka A. Perinpanayagam, Jay M. Gamma, Krista M. Vincent, Raymond Lai, Wei-Feng Dong, Manikandan Lakshmanan, Anandhkumar Raju, Vinay Tergaonkar, Soo Yong Tan, Soon Thye Lim, Lynne Postovit, Kevin D. Read, David W. Gray, Paul G. Wyatt, John R. Mackey, Luc G. Berthiaume. Targeting N-myristoylation in B-cell lymphomas as a therapeutic strategy [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5156.

N-myristoyltransferases (NMTs) catalyze the addition of 14-carbon fatty acids to the N-terminus of proteins. This activity regulates numerous membrane-bound signal transduction pathways important in cancer biology, and the pan-NMT inhibitor PCLX-001 is in clinical development as a cancer therapy. The physiologic distribution and relative contributions of the two human NMTs, NMT1 and NMT2, remain poorly understood as previous studies used polyclonal antibodies with potential cross-reactivity. We generated and validated mutually exclusive monoclonal antibodies (mAbs) specific to the human isotypes of NMT1 and NMT2. These mAbs were used to perform an immunohistochemical (IHC) analysis of the abundance and distribution of NMT1 and NMT2 in normal human breast epithelial samples and a large (n=703) cohort of primary breast adenocarcinomas from the BCIRG001 study. While NMT1 protein was readily identifiable in most normal and transformed breast epithelial tissue, NMT1 abundance was associated with higher overall histologic grade, higher Ki67, and lower hormone expression. While NMT2 protein was readily detected in normal breast epithelial tissue, NMT2 protein was undetectable in the majority of malignant breast cancers, but detectable NMT2 protein correlated with significantly poorer overall survival outcomes (hazard ratio for death 1.36; p < 0.029) and significantly worse biological features including younger age, higher histologic grade, lower hormone receptor expression, higher Ki67, and p53 positivity. NMT status was unrelated to HER2 status. NMT1 and NMT2 protein abundances were positively correlated with each other. Treatment of cultured breast cancer cells with the pan-NMT inhibitor PCLX-001 reduced cell viability in vitro. Daily oral administration of PCLX-001 to immunodeficient mice bearing human MDA-MB-231 breast cancer xenografts induced significant dose-dependent tumor growth inhibition in vivo. These results support the further evaluation of NMT immunohistochemistry for selection of patients for NMT inhibitor therapy, and clinical trials of NMT inhibition in breast cancer patients. Citation Format: John R. Mackey, Justine Lai, Utkarsh Chauhan, Wei-Feng Dong, Darryl Glubrect, Sunita Ghosh, Gilbert Bigras, Raymond Lai, Luc G. Berthiaume. N-myristoyltransferase proteins in breast cancer: Prognostic relevance and validation as a new drug target [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1782.

TPS3656 Background: Myristoylation regulates numerous membrane-bound signal transduction pathways important in cancer biology. This modification is catalyzed by N-myristoyltransferases 1 and 2 (NMT1 and NMT2). PCLX-001 is an oral small molecule with high affinity for both NMT proteins (IC50 of 5nM and 8nM, respectively) with high bioavailability and drug-like pharmacokinetic properties. In ex vivo sensitivity screening cell lines of hematologic cancer origin were exquisitely sensitive to PCLX-001, although high sensitivities and cell killing were also seen in some solid tumor lines derived from lung, pancreas, breast, colon, and bladder carcinomas. PCLX-001 demonstrated strong preclinical single-agent antitumor activity and tolerability in vivo in subcutaneous tumor xenograft models derived from lymphoma cell lines, lung cancer cell lines, a breast cancer cell line, as well as in a patient derived xenograft model from a patient with refractory DLBCL. The primary objective of this study is to determine the MTD and/or recommended phase II dose, safety, tolerability, and pharmacokinetics of PCLX-001 as a single agent, in patients with refractory lymphomas and advanced solid tumors. The secondary objective of the study is to evaluate the preliminary single agent anti-tumor activity of PCLX-001 in the patient populations studied. Methods: This is a multicenter, open-label, phase I dose-escalation study of oral PCLX-001 comprised of two parts (dose escalation and dose expansion). Eligible patients will have: histologically-confirmed advanced solid tumor or B-cell lymphomas who have failed prior therapy and/or are not eligible for therapies; ages ≥ 18 years; adequate organ function; life expectancy of at least 12 weeks; and measurable disease. Part A (dose-escalation) patients will be accrued in cohorts of 3 to 6 patients to each dose level, starting at 20 mg daily on a 28 day cycle. Dose escalation will follow a modified Fibonacci design such that the magnitude of escalation decreases as the dose level nears the human equivalent dose of the highest non-severely toxic dose in dogs and then escalate at 1.4 times the previous dose. Dose escalation and determination of the maximum tolerated dose will be based on the occurrence of dose limiting toxicities in cycle 1. Part B will have two single agent expansion cohorts (n = 20 each) in advanced solid malignancies and relapsed/refractory B-cell lymphoma, to determine the preliminary clinical activity of PCLX-001 to determine the recommended phase II dose. The first patient on study is planned for Q3 2020.

7001 Background: Docetaxel-cyclophosphamide (TC) adjuvant chemotherapy is commonly used in patients with early stage breast cancer (EBC). Due to the risk of febrile neutropenia (FN) with TC, primary prophylaxis with either ciprofloxacin (cipro) or granulocyte-colony stimulating factors (G-CSF) is recommended. Despite significant differences in costs (7-120 $US/course [cipro] vs. 2100-7000 $US/dose [G-CSF]) and toxicity profiles, optimal primary FN prophylaxis is unknown. We performed a pragmatic randomised trial comparing the superiority of G-CSF to cipro. Methods: EBC patients receiving TC chemo were randomized to receive cipro or G-CSF as primary FN prophylaxis. The primary outcome is a composite of either treatment-related hospitalisations or FN. Secondary outcomes included: chemo dose reductions, delays, discontinuations and incidence of C. difficile infections. Primary analysis was performed with the intention to treat (ITT) population. Results: 455 eligible patients were randomized to cipro (227) or G-CSF (228). 37/227 (16.3%) patients on cipro had a hospitalization, compared with 25/228 (11.0%) on G-CSF (Fisher’s exact test p-value=0.10). Relative risk (RR) of hospitalization for patients on G-CSF:0.68, 95%CI=0.42 to 1.09. Patients on cipro were statistically significantly more likely to be hospitalized for FN (30/227, 13.2%) vs 9/228 (4.0%) patients on G-CSF(p

Background Current clinical guidelines recommend mammography as the only imaging method for surveillance in asymptomatic survivors of early breast cancer (EBC). However, non-recommended tests are commonly used. We estimated the imaging radiation-induced malignancies (IRIM) risks in survivors of EBC undergoing different imaging surveillance models. Materials and Methods We built 5 theoretical models of imaging surveillance, from annual mammography only (model 1) to increasingly imaging-intensive approaches, including computed tomography (CT) scan, positron emission tomography-CT, bone scan, and multigated acquisition scan (models 2 through 5). Using the National Cancer Institute’s Radiation Risk Assessment Tool, we compared the excess lifetime attributable cancer risk (LAR) for hypothetical survivors of EBC starting surveillance at the ages of 30, 60, or 75 years and ending at 81 years. Results For all age groups analyzed, there is a statistically significant increase in LAR when comparing model 1 with more intensive models. As an example, in a patient beginning surveillance at the age of 60 years, there is a 28.5-fold increase in the IRIM risk when comparing mammography only versus a schedule with mammography plus CT scan of chest-abdomen and bone scan. We found no differences when comparing models 2 through 5. LAR is higher when surveillance starts at a younger age, although the age effect was only statistically significant in model 1. Conclusion Non-recommended imaging during EBC surveillance can be associated with a significant increase in LAR. In addition to the lack of survival benefit, additional tests may have significant IRIM risks and should be avoided.

Elevated growth in breast cancer (BC) activates hypoxia-inducible factor (HIF1α) and downstream, facilitative glucose transporter 1 (GLUT1), which can be visualized with 2-deoxy-2-[18F]fluoro-d-glucose ([18F]FDG). GLUT5 (fructose) and GLUT2 (glucose/fructose) might provide alternative targets for BC imaging as to why effects of hypoxia on GLUT1/2/5 levels and function were examined in human BC models. GLUT1/2/5 and HIF1α mRNA was analyzed in BC patient biopsies. In MCF10A, MCF7, and MDA-MB231 cells, [18F]FDG, 6-deoxy-6-[18F]fluoro-d-fructose (6-[18F]FDF) and [18F]-fluoroazomycin arabinoside were used in radiotracer experiments, whereas GLUT1/2/5 mRNA was analyzed with real-time PCR and protein levels determined via Western blot/immunohistochemistry. Positron emission tomography imaging was performed in MCF7 and MDA-MB231 tumor-bearing mice. Glucose/fructose/cytochalasin B reduced cellular 6-[18F]FDF uptake by 50%, indicating functional involvement of GLUT2. With GLUT5 staining lower than GLUT1, 6-[18F]FDF revealed lower uptake than [18F]FDG [standardized uptake value (SUV)6-[18F]FDF, 120 min 0.77 ± 0.06 vs. SUV[18F]FDG, 120 min 1.08 ± 0.07] in MDA-MB231 tumors and was blocked by 20% with cytochalasin B after 10 min. Whereas correspondence between 6-[18F]FDF uptake and GLUT5 protein was low, high GLUT2 levels were detected in all cell lines and tumor models. Besides GLUT1, GLUT5 seems to be regulated under hypoxia on the molecular and functional level. Additionally, results strongly support a functional involvement of GLUT2 in fructose metabolism, possibly by compensating for the weaker expression and function of GLUT5 in BC.-Hamann, I., Krys, D., Glubrecht, D., Bouvet, V., Marshall, A., Vos, L., Mackey, J. R., Wuest, M., Wuest, F. Expression and function of hexose transporters GLUT1, GLUT2, and GLUT5 in breast cancer-effects of hypoxia.

Estrogen receptor positive (ER+) breast cancer has been divided into two subtypes, luminal A and luminal B, which differ in their ER expression and response to hormone therapy. The absence of luminal A cell lines means the extensive amount ofin vitrowork studying the response to hormones in ER+ breast cancers is biased for the luminal B subtype. We have developed a luminal A like cell model by increasing the ER expression in the MCF-7 cell line. Our results show that increased ER expression promotes an anti-proliferative response to estrogen through regulation of genes involved in the G1/S-phase transition of the cell cycle. Furthermore, increased ER expression increases ER-DNA binding in the absence of estrogen and regulates basal gene transcription by promoting DNA looping. These results provide novel evidence that the characteristic increased ER expression of luminal A tumors may promote a novel chromatin configuration that enables growth of these tumors in the absence of estrogen and enables gene repression in the presence of hormones.

Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype with limited treatment options and poor prognosis. There is an urgent need to identify and understand the key factors and signalling pathways driving TNBC tumour progression, relapse, and treatment resistance. In this study, we report that gene copy numbers and expression levels of nuclear factor IB (NFIB), a recently identified oncogene in small cell lung cancer, are preferentially increased in TNBC compared to other breast cancer subtypes. Furthermore, increased levels of NFIB are significantly associated with high tumour grade, poor prognosis, and reduced chemotherapy response. Concurrent TP53 mutations and NFIB overexpression (z-scores0) were observed in 77.9% of TNBCs, in contrast to 28.5% in non-TNBCs. Depletion of NFIB in TP53-mutated TNBC cell lines promotes cell death, cell cycle arrest, and enhances sensitivity to docetaxel, a first-line chemotherapeutic drug in breast cancer treatment. Importantly, these alterations in growth properties were accompanied by induction of CDKN1A, the gene encoding p21, a downstream effector of p53. We show that NFIB directly interacts with the CDKN1A promoter in TNBC cells. Furthermore, knockdown of combined p21 and NFIB reverses the docetaxel-induced cell growth inhibition observed upon NFIB knockdown, indicating that NFIB's effect on chemotherapeutic drug response is mediated through p21. Our results indicate that NFIB is an important TNBC factor that drives tumour cell growth and drug resistance, leading to poor clinical outcomes. Thus, targeting NFIB in TP53-mutated TNBC may reverse oncogenic properties associated with mutant p53 by restoring p21 activity. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John WileySons, Ltd.

Background: Epirubicin dosing affects important clinical outcomes in breast cancer, with higher dose regimens improving efficacy but producing more myelosuppression. Epirubicin is metabolized by uridine glucuronosyltransferase 2B7 (UGT2B7). We previously reported relationships between UGT2B7’s promoter polymorphisms and epirubicin clearance and clinical outcomes in the (neo)adjuvant breast cancer setting; we identified a trend for increased grade 3/4 neutropenia but better efficacy outcomes in patients having at least one deficient allele (i.e. CT or CC) vs. patients who were wild type homozygotes. Patients homozygous for the deficient allele (CC) were at statistically significant increased risk for leucopenia compared to patients who were wild type homozygotes or heterozygotes. In this study we hypothesized patients with CT and TT genotypes would tolerate a higher epirubicin dose compared to CC genotype patients. We designed this study to determine the safety of pharmacogenetic-guided epirubicin dosing for each UGT2B7 genotype. Methods: Female breast cancer patients with histologically confirmed non-metastatic invasive breast cancer scheduled to receive at least three cycles of FE100C in the (neo)adjuvant setting were enrolled into the study. Peripheral blood was analyzed for UGT2B7 genotype. Patients received standard dose IV FE100C during the first 21 day cycle. Based on genotype, epirubicin dosing was escalated in the 2nd and 3rd cycles. Epirubicin Dose Escalation SchemeDose of Epirubicin per Cycle (mg/m2)CycleGenotype123CC100100100CT100115130TT100120140 Results: To date 32 patients are evaluable for pharmacogenetic guided epirubicin dosing (8 CC genotypes, 14 CT genotypes and 10 TT genotypes). All 32 patients received epirubicin100 mg/m2 in cycle one and a single patient in each of the CC and CT genotypes experienced grade 3 febrile neutropenia and were not dose escalated. All other patients with CT and TT genotypes were dose escalated in cycle 2 and all but two patients in the CT and TT genotypes were dose escalated in cycle 3. The incidence of febrile neutropenia was not dose dependent as all three genotypes had similar incidence in each cycle whereas leucopenia was genotype and dose dependent. The incidence of leukopenia increased in patients with CT and TT genotypes as their dose was increased and cycle 3 leukopenia rates were similar to patients with the CC genotype receiving standard dose epirubicin. Conclusions: Pharmacogenetic guided epirubicin dosing is well tolerated. This study is ongoing and updated data will be presented. Citation Format: John R Mackey, Edith Pituskin, Ann Vlahadamis, Katia Tonkin, Karen King, Sanraj Basi, Maria Ho, Judith Meza-Junco, Anil Joy, Dick Au, Sambasivarao Damaraju, Michael B Sawyer. Pharmacogenetic dosing of epirubicin in FEC chemotherapy [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P3-06-48.

Background : The distinction between HER2-positive (IHC 3+ or 2+ with FISH ratio >/= 2) and not overexpressing HER2 has been the focus of many diagnostic tests over the past years in association with development of HER2-targeted therapies. The paucity of therapies developed for the low to intermediate HER2 protein expression populations has resulted in limited attention to their diagnostic precision and accuracy. The development of NeuVaxTM (nelipepimut-S; Galena Biopharma, Inc.) in the defined population requires a HER2 IHC 1+/2+ diagnostic that precisely and accurately ensures identification of targeted patients. We describe discordance rates between local and central testing performed to identify tumors with HER2 IHC 1+/2+ expression that supports the development of a method to validate HER2 1+ and 2+ (FISH < 2.2) patients who receive nelipepimut-S adjuvant therapy. Methods : The Prevention of Recurrence in Early Stage, Node-Positive Breast Cancer with Low to Intermediate HER2 Expression with NeuVaxTM Treatment (PRESENT) study, is a multicenter, multinational, prospective, randomized, double-blind, controlled Phase 3 study assessing efficacy and safety of the peptide vaccine nelipepimut-S, in HLA A2 or A3 positive patients with early stage, node positive breast cancer expressing low and intermediate levels (IHC 1+/2+) of HER2 protein. PRESENT 2-step screening includes HLA testing and central lab confirmation of HER2 1+ or 2+ expression using the DAKO HercepTest. Results : As of 2 June 2014, 1454 patients underwent central IHC testing for HER2 and had a quantifiable result of 0, 1+, 2+, or 3+ for both local and central test. Per local testing, 61% (HER2 1+, n=612; HER2 2+, n=275) were eligible and 39% (HER2 0, n=468; HER2 3+, n=99) were ineligible. Of those eligible by local testing, 67.5% (n=599) were confirmed as eligible per central testing for a discordance rate of 32.5% (n=288). Of the 288 discordant samples tested centrally, 73.6% (n=212) and 26.4% (n=76) were reported as HER2 0 and 3+, respectively. 8.7% (76/877) of patients found to be HER2 1+ or 2+ by local testing were determined to be HER2+ (IHC3+) by central testing. Conclusions : Current tests for HER2 expression are defined by their ability to determine 3+ positivity, yet significant discordance still occurs with nearly 9% false negative rate in this trial. Similarly, marked discordance exists between local and central laboratory test results for HER2 by IHC at 1+/2+ levels of expression. The relatively high discordance rate observed may be due, in part, to the lack of a validated IHC assay for low-to-intermediate expression of HER2 (0, 1+, and 2+). In order to improve accuracy of testing and to develop a companion diagnostic for nelipepimut-S, the Leica Bond Oracle HER2 IHC System has been validated to determine samples across the IHC spectrum (0, 1+, 2+ and 3+) and is now incorporated into HER2 screening for the PRESENT trial as a companion diagnostic to increase accuracy, precision, and specificity in discerning HER2 1+ and 2+ patients. Citation Format: Michelle Melisko, Elizabeth A Mittendorf, Sufia Safina, Michael Schenker, Murray A Brunt, Maria Litwiniuk, John Mackey, Katarina Petrakova, Svitiana Alieva, Lacey Chance, Gavin S Choy, Mark Ahn, Adamm Hamm, Sonia Kumar, Hope S Rugo. HER2 discordant results in local vs. central testing in the phase 3 nelipepimut-S trial and implementation of Leica Bond Oracle HER2 Immunohistochemistry (IHC) System for low and intermediate levels (1+, 2+) of HER2 protein expression as a companion diagn [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P2-15-02.

Purpose: In pharmaceutical clinical trials, industrial sponsors pay for study drugs and related healthcare services. We conducted a study to determine industry’s economic contribution of these trials to the Alberta healthcare system. Methods: We used data from two trial centers for cancer and non-cancer trials at the University of Alberta. For each trial (cancer, non-cancer), we calculated the cost of drugs provided by the sponsors using the market price, the cost of clinical services, and the cost of administrative services that they paid. We extrapolated these results to all trials in Alberta based on information obtained from the registration website ClinicalTrials.gov. Results: Our sample consisted of 40 non-cancer and 39 cancer drug trials which were initiated in 2012. The monetary value of the industry sponsors’ contribution was $799,055 per non-cancer and $630,243 per cancer drug trial. Drugs (in-trial and post-trial) accounted for 84% of the total contribution of the non-cancer drug trials whereas it represented 93% of all trial-related contributions in the cancer category. The total province-wide contribution of industry-sponsored drug trials which were initiated in 2012 was estimated to be $101 million, including open-label drugs in the non-cancer category. Conclusions: Industry-sponsored pharmaceutical trials represent a major economic contributor to clinical research within the province.

Background: Dihydropyrimidine dehydrogenase (DPD) tumour expression may provide added value to human equilibrative nucleoside transporter-1 (hENT1) tumour expression in predicting survival following pyrimidine-based adjuvant chemotherapy. Methods: DPD and hENT1 immunohistochemistry and scoring was completed on tumour cores from 238 patients with pancreatic cancer in the ESPAC-3(v2) trial, randomised to either postoperative gemcitabine or 5-fluorouracil/folinic acid (5FU/FA). Results: DPD tumour expression was associated with reduced overall survival (hazard ratio, HR = 1.73 [95% confidence interval, CI = 1.21-2.49], p = 0.003). This was significant in the 5FU/FA arm (HR = 2.07 [95% CI = 1.22-3.53], p = 0.007), but not in the gemcitabine arm (HR = 1.47 [0.91-3.37], p = 0.119). High hENT1 tumour expression was associated with increased survival in gemcitabine treated (HR = 0.56 [0.38-0.82], p = 0.003) but not in 5FU/FA treated patients (HR = 1.19 [0.80-1.78], p = 0.390). In patients with low hENT1 tumour expression, high DPD tumour expression was associated with a worse median [95% CI] survival in the 5FU/FA arm (9.7 [5.3-30.4] vs 29.2 [19.5-41.9] months, p = 0.002) but not in the gemcitabine arm (14.0 [9.1-15.7] vs. 18.0 [7.6-15.3] months, p = 1.000). The interaction of treatment arm and DPD expression was not significant (p = 0.303), but the interaction of treatment arm and hENT1 expression was (p = 0.009). Conclusion: DPD tumour expression was a negative prognostic biomarker. Together with tumour expression of hENT1, DPD tumour expression defined patient subgroups that might benefit from either postoperative 5FU/FA or gemcitabine.

Background: Few studies have investigated the effects of exercise on modulation of host factors in cancer patients. We investigated the efficacy of chronic aerobic training on multiple host-related effector pathways in patients with solid tumours. Patients and Methods: Paired peripheral blood samples were obtained from 44 patients with solid tumours receiving cytotoxic therapy and synthetic erythropoietin (usual care; n=21) or usual care plus supervised aerobic training (n=23) for 12 weeks. Samples were characterised for changes in immune, cytokine and angiogenic factors, and metabolic intermediates. Aerobic training consisted of three supervised cycle ergometry sessions per week at 60% to 100% of peak oxygen consumption (VO2peak), 30–45 min per session, for 12 weeks following a nonlinear prescription. Results: The between-group delta change in cardiopulmonary function was +4.1 ml kg −1 min−1, favouring aerobic training (P

The purpose of this analysis was to assess the long-term impact of adding bevacizumab to adjuvant chemotherapy for early triple-negative breast cancer (TNBC).Patients eligible for the open-label randomized phase III BEATRICE trial had centrally confirmed triple-negative operable primary invasive breast cancer (pT1a-pT3). Investigators selected anthracycline- and/or taxane-based chemotherapy for each patient. After definitive surgery, patients were randomized 1:1 to receive ≥4 cycles of chemotherapy alone or with 1 year of bevacizumab (5 mg/kg/week equivalent). Stratification factors were nodal status, selected chemotherapy, hormone receptor status, and type of surgery. The primary end point was invasive disease-free survival (IDFS; previously reported). Secondary outcome measures included overall survival (OS) and safety.After 56 months' median follow-up, 293 of 2591 randomized patients had died. There was no statistically significant difference in OS between treatment arms in either the total population (hazard ratio 0.93, 95% confidence interval [CI] 0.74-1.17; P = 0.52) or pre-specified subgroups. The 5-year OS rate was 88% (95% CI 86-90%) in both treatment arms. Updated IDFS results were consistent with the primary IDFS analysis. Five-year IDFS rates were 77% (95% CI 75-79%) with chemotherapy alone versus 80% (95% CI 77-82%) with bevacizumab. From 18 months after first study dose to study end, new grade ≥3 adverse events occurred in 4.6% and 4.5% of patients in the two arms, respectively.Final OS results showed no significant benefit from bevacizumab therapy for early TNBC. Late-onset toxicities were rare in both groups. Five-year OS and IDFS rates suggest that the prognosis for patients with TNBC is better than previously thought.NCT00528567.

AB Observational studies suggest that physical activity after a breast cancer diagnosis is associated with improved cancer outcomes; however, no randomized data are available. Here, we report an exploratory follow-up of cancer outcomes from the Supervised Trial of Aerobic versus Resistance Training (START). Methods: The START was a Canadian multicenter trial that randomized 242 breast cancer patients between 2003 and 2005 to usual care (n = 82), supervised aerobic (n = 78), or resistance (n = 82) exercise during chemotherapy. The primary end point for this exploratory analysis was disease-free survival (DFS). Secondary end points were overall survival, distant DFS, and recurrence-free interval. The two exercise arms were combined for analysis (n = 160), and selected subgroups were explored. Results: After a median follow-up of 89 months, there were 25/160 (15.6%) DFS events in the exercise groups and 18/82 (22.0%) in the control group. Eight-year DFS was 82.7% for the exercise groups compared with 75.6% for the control group (HR, 0.68; 95% confidence interval (CI), 0.37-1.24; log-rank, P = 0.21). Slightly stronger effects were observed for overall survival (HR, 0.60; 95% CI, 0.27-1.33; log-rank, P = 0.21), distant DFS (HR, 0.62; 95% CI, 0.32-1.19; log-rank, P = 0.15), and recurrence-free interval (HR, 0.58; 95% CI, 0.30-1.11; Gray test, P = 0.095). Subgroup analyses suggested potentially stronger exercise effects on DFS for women who were overweight/obese (HR, 0.59; 95% CI, 0.27-1.27), had stage II/III cancer (HR, 0.61; 95% CI, 0.31-1.20), estrogen receptor-positive tumors (HR, 0.58; 95% CI, 0.26-1.29), human epidermal growth factor receptor 2-positive tumors (HR, 0.21; 95% CI, 0.04-1.02), received taxane-based chemotherapies (HR, 0.46; 95% CI, 0.19-1.15), and >=85% of their planned chemotherapy (HR, 0.50; 95% CI, 0.25-1.01). Conclusions: This exploratory follow-up of the START provides the first randomized data to suggest that adding exercise to standard chemotherapy may improve breast cancer outcomes. A definitive phase III trial is warranted

Aldehyde dehydrogenase (ALDH) 1A enzymes produce retinoic acid (RA), a transcription induction molecule. To investigate if ALDH1A1 or ALDH1A3‐mediated RA signaling has an active role in breast cancer tumorigenesis, we performed gene expression and tumor xenograft studies. Analysis of breast patient tumors revealed that high levels of ALDH1A3 correlated with expression of RA‐inducible genes with retinoic acid response elements (RAREs), poorer patient survival and triple‐negative breast cancers. This suggests a potential link between ALDH1A3 expression and RA signaling especially in aggressive and/or triple‐negative breast cancers. In MDA‐MB‐231, MDA‐MB‐468 and MDA‐MB‐435 cells, ALDH1A3 and RA increased expression of RA‐inducible genes. Interestingly, ALDH1A3 had opposing effects in tumor xenografts, increasing tumor growth and metastasis of MDA‐MB‐231 and MDA‐MB‐435 cells, but decreasing tumor growth of MDA‐MB‐468 cells. Exogenous RA replaced ALDH1A3 in inducing the same opposing tumor growth and metastasis effects, suggesting that ALDH1A3 mediates these effects by promoting RA signaling. Genome expression analysis revealed that ALDH1A3 induced largely divergent gene expression in MDA‐MB‐231 and MDA‐MB‐468 cells which likely resulted in the opposing tumor growth effects. Treatment with DNA methylation inhibitor 5‐aza‐2'deoxycytidine restored uniform RA‐inducibility of RARE‐containing HOXA1 and MUC4 in MDA‐MB‐231 and MDA‐MB‐468 cells, suggesting that differences in epigenetic modifications contribute to differential ALDH1A3/RA‐induced gene expression in breast cancer. In summary, ALDH1A3 induces differential RA signaling in breast cancer cells which affects the rate of breast cancer progression.

N-myristoylation, the addition of the 14-carbon fatty acid to proteins, plays a fundamental role in cell signaling. Over 200 proteins are myristoylated, including the Src Family Kinases (SFK) Src, Lyn, Lck, Hck, and Fgr, as well as c-Abl, Gα subunits, caspase truncated (ct-) Bid and ct-PAK2, regulating cell growth and apoptosis. Human myristoylation is performed by two ubiquitously expressed N-myristoyl-transferases NMT1 and NMT2. PCLX-001 is a new, orally bioavailable, small-molecule, dual NMT inhibitor that is under investigation as a novel and selective treatment for B-cell malignancies. In vitro, PCLX-001 inhibits NMT1 and NMT2 at IC50 of 5nM and 8nM, respectively, inhibits the growth of hematological cancer cells at concentrations 10-fold lower than dasatinib and ibrutinib, and inhibits SFK recruitment to B cell receptor signaling, reducing survival signals and triggering apoptosis. PCLX-001 causes complete tumor regression in NOD/SCID mouse xenograft models of Acute Myeloid Leukemia (AML), Burkitts Lymphoma (BL), and Diffuse Large B-cell Lymphoma (DLBCL), including drug-resistant human tumor in a PDX model. When screened in vitro for its ability to inhibit the activity of 468 kinases and kinase mutants in a KINOMEscan®, PCLX-001 did not inhibit any kinase at up to 10 µM (5380 ng/mL) and produced modest inhibition of only three kinases (MRCKA, PIP5K2B, and SRPK1) at 100 µM (53800 ng/mL). Male rats tolerated single oral doses of PCLX-001 at 100 mg/kg without clear effects, but one of three rats died after a single dose at 1000 mg/kg. Dogs tolerated single oral doses of PCLX-001 at 10 mg/kg without effects, but both dogs showed emesis and diarrhea and lost weight after a single dose at 50 mg/kg. In 21-day xenograft efficacy studies in mice, the maximum tolerated dose level (MTD) was 50mg/kg, which also produced complete tumor remission of most xenografts. Administered daily, the 14-day MTD is >75mg/kg (highest dose tested) in rats and between 5 and 25mg/kg in dogs. The dose-limiting toxicities in dogs involved the gastrointestinal tract and hematopoietic bone marrow. When administered orally, PCLX-001 is 26% bioavailable in rats and >90% bioavailable in mice and dogs. The plasma half-life was 5.7h in mice, 1-2h in rats, and 3.9h in dogs. With repeated daily administration, systemic exposure did not change in dogs but decreased in rats. Based on KINOMEscan® results, PCLX-001 is very unlikely to produce adverse effects in patients due to off-target kinase inhibition. The oral bioavailability and half-life in mice and dogs are consistent with a once-a-day dosing regimen. Repeat dosing studies of up to 21 days in mice and 14 days in rats and dogs determined preliminary MTDs. When extrapolated to human doses based on a body surface area scaling, these MTDs are equivalent to 150mg/m2 in mice, >450mg/m2 in rats, and between 100 and 500 mg/m2 in dogs. These results are consistent with a therapeutic window with potential for tumor response in humans, and support proceeding with 28-day GLP toxicology studies and an IND filing for first in human testing of oral PCLX-001. Disclosures Weickert: Pacylex Pharmaceuticals, Inc.,: Employment. Mackey:Pacylex Pharmaceuticals Inc.: Equity Ownership, Patents & Royalties; illumiSonics Inc: Equity Ownership, Other: Director Role; Pfizer Canada: Honoraria; CME: Honoraria; SMHeartCard Inc: Equity Ownership, Other: Director Role. Berthiaume:Pacylex Pharmaceuticals, Inc.,: Equity Ownership, Patents & Royalties.

Treatment of aggressive lymphoma is toxic, expensive, and a substantial proportion of patients relapse and die. There is an urgent need for more effective treatments. Myristoylation is required for biological activity of >200 intracellular proteins. N-myristoyltransferases (NMTs) transfer the fatty acid myristate to N-terminal glycine residue; there are two isoforms, NMT1 and 2. Since they are critical to intracellular signaling, NMTs are potential anti-cancer targets. We tested a novel potent pan-NMT inhibitor, PCLX-001, in B cell lymphoma cell lines. In vitro assays included cell viability, immunoblotting, and metabolic labeling of lymphoma cell lines. Immunohistochemistry was performed on formalin fixed paraffin embedded lymphoma specimens from patients. In vivo experiments included cell line derived murine xenografts and a patient derived mouse xenograft treated with increasing concentrations of PCLX-001. PCLX-001 selectively killed lymphoma cells, while sparing normal cells in vitro and in 3 mouse xenograft models, eradicating tumors in two of these models including a patient-derived xenograft from a R-CHOP refractory lymphoma patient. While NMT2 is overexpressed in some cancers, loss of NMT2 expression is common in numerous cancers and occurs at the highest prevalence in lymphomas, where it is independently linked to a worse prognosis. This NMT2 suppression occurred through epigenetic mechanisms and may account for lymphoma sensitivity to NMT inhibition. The global myristoylation of lymphoma cell proteins, including that of the protein tyrosine kinase oncogene Src, is profoundly inhibited by PCLX-001. Loss of Src myristoylation is accompanied by loss of Src activity and may account for loss of prosurvival signals causing lymphoma cell death. Targeting NMT2 deficient B cell lymphoma with a pan-NMT inhibitor suppresses the residual NMT1 function provides a novel, selective, and effective therapeutic strategy. Citation Format: John R. Mackey, Erwan Beauchamp, Megan C. Yap, Aishwarya Iyer, Maneka A. Perinpanayagam, Krista M. Vincent, Abass M. Al-Momany, Ryan J. Heit, Jacky Y. Sim, Raymond Lai, Wei-feng Dong, Manikandan Lakshmanan, Anandhkumar Raju, Vinay Tergaonkar, Soo Yong Tan, Soon Thye Lim, Lynne M. Postovit, Kevin D. Read, David W. Gray, Paul G. Wyatt, Luc G. Berthiaume. Targeting N-myristoylation in B cell lymphomas as a therapeutic strategy [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 3046.

Myristoylation is required for biological activity of >200 intracellular proteins. N-myristoyltransferases (NMTs) transfer the fatty acid myristate to N-terminal glycine residue; there are two isoforms, NMT1 and 2. In acute myeloid leukemia (AML), upregulation of Lyn and Src, important myristoylated proteins, contribute to cell survival and proliferation. The specific roles of NMT1 and NMT2 are unknown in this context. The relationships among NMT1/NMT2 expression and acute AML patient outcomes were studied using RNA-sequencing and microarray cohorts with over 350 patients. We found that high NMT1 and low NMT2 expression were associated with reduced overall and event-free survival in adult AML, which was independent of other prognostic markers on multivariate analysis. High NMT1, but not NMT2, expression was associated with proliferative gene sets in AML cell lines, indicating potential for distinct isozyme substrates. Given these results, we examined NMT1 and NMT2 levels in AML cell lines and AML patient blast cells using Western blotting and flow cytometry. We determined that NMT2 expression varied greatly among patients, but was markedly reduced in most cases, while NMT1 expression was relatively preserved. A potent small molecule NMT inhibitor, PCLX-001, preferentially induced apoptosis and reduced viability in NMT2-deficient AML cell lines cultured in vitro compared with normal lymphocytes and peripheral blood mononuclear cells. PCLX-001 also killed freshly isolated human AML blasts ex vivo with an IC50 of ~170nM regardless of their mutational background. In a murine AML xenograft model, subcutaneously delivered PCLX-001 monotherapy demonstrated dose-dependent anticancer activity and produced complete remissions after five daily 50 mg/kg doses. NMT expression provides independent prognostic information to refine existing clinical stratification, and NMT inhibition is a promising novel therapeutic strategy for AML. Citation Format: John R. Mackey, Aishwarya Iyer, Megan C. Yap, Zoulika Zak, Krista Vincent, Erwan Beauchamp, Lynne M. Postovit, Joseph Brandwein, Luc G. Berthiaume. Examination of NMT1 and NMT2 as independent prognostic markers and novel drug targets in adult acute myeloid leukemia [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 3043.

Key Points Questions How does one assess the status of HER2 ISH-equivocal breast cancers as either HER2 positive or HER2 negative for treatment purposes, and are the use of alternative controls, as recommended by the 2013/2014 American Society of Clinical Oncology and College of American Pathologists guidelines, appropriate? Findings In this study, chromosome 17 p-arm genomic sites had a high rate of heterozygous deletions, both in the publicly available Molecular Taxonomy of Breast Cancer International Consortium database and in the Breast Cancer International Research Group-005 clinical trial samples using fluorescence in situ hybridization. Meaning The indiscriminate use of alternative controls to assess HER2 status with HER2-to-control gene ratios by ISH may lead to false-positive determinations and should be avoided, as recommended by the 2018 clinical practice update., This study examines HER2 ISH-equivocal breast cancers to asses 2013/2014 American Society of Clinical Oncology and College of American Pathologists guidelines in determining HER2-positive or HER2-negative breast cancers and false-positive results., Importance The 2013/2014 American Society of Clinical Oncology and College of American Pathologists (ASCO-CAP) guidelines for HER2 testing by fluorescence in situ hybridization (FISH) designated an “equivocal” category (average HER2 copies per tumor cell ≥4-6 with HER2/CEP17 ratio

In pancreatic cancer, deoxycytidine kinase and the human equilibrative nucleoside transporter 1 have been validated as predictive markers for benefit from gemcitabine therapy. Gemcitabine is used with cisplatin or carboplatin as neoadjuvant chemotherapy for muscle invasive urothelial cancer of the bladder before radical cystectomy and patients rendered disease-free at surgery tend to have better outcomes. In this trial we examined if nucleoside transporter or deoxycytidine kinase protein abundance in biopsy specimens before chemotherapy is related to the response to neoadjuvant chemotherapy.A total of 62 consecutive patients undergoing neoadjuvant chemotherapy with platinum/gemcitabine at a single institution were accrued. Initial transurethral resection of bladder tumor specimens and cystectomy specimens were collected, and scored for nucleoside transporter and deoxycytidine kinase expression. Pathological response rates and survival data were collected.Of the 62 patients 17 (27%) achieved a complete pathological response (pT0) to neoadjuvant chemotherapy. Nucleoside transporter and deoxycytidine kinase protein expression in the transurethral resection of bladder tumor specimens did not predict for pT0 status to neoadjuvant chemotherapy. Median overall survival was not reached for the group achieving pT0 status and was 46 months for those with persistent cancer at definitive surgery (p = 0.07). Median followup for the cohort was 30 months.Nucleoside transporter and deoxycytidine kinase expression in transurethral resection of bladder tumor samples do not predict for response to gemcitabine and platinum neoadjuvant chemotherapy. Patients should continue to be offered neoadjuvant chemotherapy before radical cystectomy based on clinical and pathological staging.

Background: Observational studies suggest that physical activity following a diagnosis of breast cancer may be associated with a lower risk of recurrence and death. Some studies also suggest possible effect modification by disease stage, body mass index, and receptor status. To date, however, there are no randomized trials examining the effects of exercise on disease outcomes in any cancer patient group. Here, we report an exploratory follow-up of disease outcomes from the Supervised Trial of Aerobic versus Resistance Training (START). Patients and Methods: The START Trial was a Canadian multicenter trial that randomized 242 breast cancer patients starting adjuvant chemotherapy to either usual care (n = 82) or supervised aerobic (n = 78) or resistance (n = 82) exercise for the duration of their chemotherapy. The primary efficacy endpoint for this exploratory analysis was disease-free survival (DFS). Secondary endpoints were overall survival (OS), distant disease-free survival (DDFS), and recurrence-free interval (RFI). The two exercise arms were combined for the analysis (n = 160) and selected subgroups were explored. Results: After a median follow-up of 89 months (IQR 81 to 96), there were 25/160 (15.6%) DFS events in the exercise groups and 18/82 (22.0%) in the control group (log-rank p = 0.21). Eight-year DFS was 82.7% for the exercise groups compared with 75.6% for the control group (Hazard ratio [HR] = 0.68, 95% CI = 0.37-1.24). There were 13/160 (8.1%) deaths in the exercise groups and 11/82 (13.4%) in the control group (log-rank p = 0.21). Eight-year OS was 91.2% in the exercise groups compared with 82.7% in the control group (HR = 0.60, 95% CI = 0.27 to 1.33. There were 20/160 (12.5%) DDFS events in the exercise groups and 16/82 (19.5%) in the control group (log-rank p = 0.15). Eight-year DDFS was 86.7% in the exercise groups compared with 78.3% in the control group (HR = 0.62, 95% CI = 0.32 to 1.19). Finally, there were 20/160 (12.5%) RFI events in the exercise groups and 17/82 (20.7%) in the control group (Gray's p = 0.095). Eight-year cumulative incidence of RFI was 12.6% in the exercise groups compared with 21.6% in the control group (HR = 0.58, 95% CI = 0.30 to 1.11). Subgroup analyses for DFS and RFI suggested stronger effects for women who were overweight/obese, had stage II/III cancer, receptor positive tumors, HER2 positive tumors, received taxane-based chemotherapies, and received at least 85% of their intended chemotherapy dose-intensity. The most notable subgroup effect was for patients who received optimal chemotherapy dosing with a borderline significant effect for DFS (HR = 0.50, 95% CI = 0.25 to 1.01) and a significant effect for RFI (HR = 0.38, 95% CI = 0.18 to 0.81). Conclusions: In this exploratory follow-up of the START Trial, there was a suggestion that exercise during adjuvant chemotherapy may improve several efficacy endpoints although none achieved statistical significance. Nevertheless, the magnitude of the effects appear to be meaningful with absolute 8-year survival differences between 7% and 9% and relative rate reductions between 30% and 40%. The START Trial provides the first randomized data to suggest that adding exercise to standard chemotherapy for breast cancer may improve outcomes. A definitive phase III trial is warranted. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P4-08-01.

Background: Aerobic and resistance exercise, either separately or in combination, have been shown to improve physical functioning and manage some symptoms in breast cancer patients receiving chemotherapy. Few exercise trials, however, have compared different doses or types of exercise in breast cancer patients to identify the optimal exercise prescription. Here, we report the primary results of the Combined Aerobic and Resistance Exercise (CARE) Trial which compared two different doses and types of exercise for improving physical functioning and symptom management in breast cancer patients receiving chemotherapy. Methods: A multicenter trial in Canada randomized 301 breast cancer patients between 2008 and 2011 to thrice weekly, supervised exercise during chemotherapy consisting of either a standard dose of 25-30 minutes of aerobic exercise (STAN; n = 96), a higher dose of 50-60 minutes of aerobic exercise (HIGH; n = 101), or a combined dose of 50-60 minutes of aerobic and resistance exercise (COMB; n = 104). The primary endpoint was patient-reported physical functioning assessed by the Medical Outcomes Survey-Short Form (SF)-36. Secondary endpoints were other physical functioning scales, symptoms, physical fitness, and chemotherapy completion. Results: Between April 2008 and September 2011, we randomized 301 of 728 (41%) eligible patients. STAN, HIGH and COMB completed 88% (43/49), 82% (40/49), and 78% (39/50) of their prescribed aerobic exercise sessions. We obtained patient-reported outcome data from questionnaires on 99% of patients at each point during and after chemotherapy. Adjusted linear mixed-model analyses showed that neither HIGH (+0.8; 95% CI [-0.8 to 2.4]; p = 0.30) nor COMB (+0.5; 95% CI [-1.1 to 2.1]; p = 0.52) were statistically superior to STAN for the primary outcome. For important secondary outcomes, HIGH was superior to STAN for the SF-36 physical component summary (p = 0.041), SF-36 bodily pain (p = 0.015), and endocrine symptoms (p = 0.019). COMB was superior to STAN for endocrine symptoms (p = 0.009) and superior to STAN (p Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr PD2-6.

BACKGROUND: Exercise improves physical functioning and symptom management during breast cancer chemotherapy, but the effects of different doses and types of exercise are unknown. METHODS: A multicenter trial in Canada randomized 301 breast cancer patients to thrice-weekly supervised exercise during chemotherapy consisting of either a standard dose of 25 to 30 minutes of aerobic exercise (STAN; n = 96), a higher dose of 50 to 60 minutes of aerobic exercise (HIGH; n = 101), or a combined dose of 50 to 60 minutes of aerobic and resistance exercise (COMB; n = 104). The primary endpoint was physical functioning assessed by the Medical Outcomes Survey-Short Form (SF)-36. Secondary endpoints were other physical functioning scales, symptoms, fitness, and chemotherapy completion. All statistical tests were linear mixed model analyses, and the P values were two-sided. RESULTS: Follow-up assessment of patient-reported outcomes was 99.0%. Adjusted linear mixed-model analyses showed that neither HIGH (+0.8; 95% confidence interval [CI] = -0.8 to 2.4; P = .30) nor COMB (+0.5; 95% CI = -1.1 to 2.1; P = .52] were superior to STAN for the primary outcome. In secondary analyses not adjusted for multiple comparisons, HIGH was superior to STAN for the SF-36 physical component summary (P = .04), SF-36 bodily pain (P = .02), and endocrine symptoms (P = .02). COMB was superior to STAN for endocrine symptoms (P = .009) and superior to STAN (P < .001) and HIGH (P < .001) for muscular strength. HIGH was superior to COMB for the SF-36 bodily pain (P = .04) and aerobic fitness (P = .03). No differences emerged for body composition or chemotherapy completion. CONCLUSIONS: A higher volume of aerobic or combined exercise is achievable and safe during breast cancer chemotherapy and may manage declines in physical functioning and worsening symptoms better than standard volumes.

Tubulin is the target for very widely used anti-tumor drugs, including Vinca alkaloids, taxanes, and epothilones, which are an important component of chemotherapy in breast cancer and other malignancies. Paclitaxel and other tubulin-targeting drugs bind to the β subunit of tubulin, which is a heterodimer of α and β subunits. β-Tubulin exists in the form of multiple isotypes, which are differentially expressed in normal and neoplastic cells and differ in their ability to bind to drugs. Among them, the βIII isotype is overexpressed in many aggressive and metastatic cancers and may serve as a prognostic marker in certain types of cancer. The underpinning mechanisms accounting for the overexpression of this isotype in cancer cells are unclear. To better understand the role of β-tubulin isotypes in cancer, we analyzed over 1000 clones from 90 breast cancer patients, sequencing their β-tubulin isotypes, in search of novel mutations. We have elucidated two putative emerging molecular subgroups of invasive breast cancer, each of which involve mutations in the βI-, βIIA-, or βIVB isotypes of tubulin that increase their structural, and possibly functional, resemblance to the βIII isotype. A unifying feature of the first of the two subgroups is the mutation of the highly reactive C239 residue of βI- or βIVB-tubulin to L239, R239, Y239, or P239, culminating in probable conversion of these isotypes from ROS-sensitive to ROS-resistant species. In the second subgroup, βI, βIIA, and βIVB have up to seven mutations to the corresponding residues in βIII-tubulin. Given that βIII-tubulin has emerged as a pro-survival factor, overexpression of this isotype may confer survival advantages to certain cancer cell types. In this mini-review, we bring attention to a novel mechanism by which cancer cells may undergo adaptive mutational changes involving alternate β-tubulin isotypes to make them acquire some of the pro-survival properties of βIII-tubulin. These "hybrid" tubulins, combining the sequences and/or properties of two wild-type tubulins (βIII and either βI, βIIA, or βIVB), are novel isotypes expressed solely in cancer cells and may contribute to the molecular understanding and stratification of invasive breast cancer and provide novel molecular targets for rational drug development.

Molecularly targeted therapeutics (MTT) are the future of cancer systemic therapy. They have already moved from palliative therapy for advanced solid malignancies into the setting of curative-intent treatment for early-stage disease. Cardiotoxicity is a frequent and potentially serious adverse complication of some targeted therapies, leading to a broad range of potentially life-threatening complications, therapy discontinuation, and poor quality of life. Low-cost pleiotropic interventions are therefore urgently required to effectively prevent and/or treat MTT-induced cardiotoxicity. Aerobic exercise therapy has the unique capacity to modulate, without toxicity, multiple gene expression pathways in several organ systems, including a plethora of cardiac-specific molecular and cell-signaling pathways implicated in MTT-induced cardiac toxicity. In this review, we examine the molecular signaling of antiangiogenic and HER2-directed therapies that may underpin cardiac toxicity and the hypothesized molecular mechanisms underlying the cardioprotective properties of aerobic exercise. It is hoped that this knowledge can be used to maximize the benefits of small molecule inhibitors, while minimizing cardiac damage in patients with solid malignancies.

Summary Background We compared standard adjuvant anthracycline chemotherapy with anthracycline–taxane combination chemotherapy in women with operable node-positive breast cancer. Here we report the final, 10-year follow-up analysis of disease-free survival, overall survival, and long-term safety. Methods BCIRG 001 was an open label, phase 3, multicentre trial in which 1491 patients aged 18–70 years with node-positive, early breast cancer and a Karnofsky score of 80% or more were randomly assigned to adjuvant treatment with docetaxel, doxorubicin, and cyclophosphamide (TAC) or fluorouracil, doxorubicin, and cyclophosphamide (FAC) every 3 weeks for six cycles. Randomisation was stratified according to institution and number of involved axillary lymph nodes per patient (one to three vs four or more). Disease-free survival was the primary endpoint and was defined as the interval between randomisation and breast cancer relapse, second primary cancer, or death, whichever occurred first. Efficacy analyses were based on the intention-to-treat principle. BCIRG 001 is registered with ClinicalTrials.gov, number NCT00688740. Findings Enrolement took place between June 11, 1997 and June 3, 1999; 745 patients were assigned to receive TAC and 746 patients were assigned to receive FAC. After a median follow-up of 124 months (IQR 90–126), disease-free survival was 62% (95% CI 58–65) for patients in the TAC group and 55% (51–59) for patients in the FAC group (hazard ratio [HR] 0·80, 95% CI 0·68–0·93; log-rank p=0·0043). 10-year overall survival was 76% (95% CI 72–79) for patients in the TAC group and 69% (65–72) for patients in the FAC group (HR 0·74, 0·61–0·90; log-rank p=0·0020). TAC improved disease-free survival relative to FAC irrespective of nodal, hormone receptor, and HER2 status, although not all differences were significant in these subgroup analyses. Grade 3–4 heart failure occurred in 26 (3%) patients in the TAC group and 17 (2%) patients in the FAC group, and caused death in two patients in the TAC group and four patients in the FAC group. A substantial decrease in left ventricular ejection fraction (defined as a relative decrease from baseline of 20% or more) was seen in 58 (17%) patients who received TAC and 41 (15%) patients who received FAC. Six patients who received TAC developed leukaemia or myelodysplasia, as did three patients who received FAC. Interpretation Our results provide evidence that the initial therapeutic outcomes seen at the 5-year follow-up with a docetaxel-containing adjuvant regimen are maintained at 10 years. However, a substantial percentage of patients had a decrease in left ventricular ejection fraction, probably caused by anthracycline therapy, which warrants further investigation. Funding Sanofi.

Fascin 1 (FSCN1) is a cytoskeleton-associated protein recognized to function primarily in the regulation of cytoskeleton structure and formation of plasma membrane protrusions. Here we report a novel nuclear function for Fascin 1. Biochemical studies and genome wide localization using ChIP-seq identified phosphorylated Fascin 1 (pFascin) in complexes associated with transcription and that it co-localizes with histone H3 Lys4 trimethylation (H3K4me3) on chromatin. Gene expression profiling identified genes affected by Fascin 1 including SLC3A2, a gene encoding for a plasma membrane transporter that regulates intracellular amino acid levels. RbBP5, a subunit of the H3K4 histone methyltransferase (HMT) complex was found to interact with Fascin 1 supporting its role in H3K4me3 establishment at target genes. Moreover, we show that changes to SLC3A2 levels affect amino acid-mediated mTORC1 activation. These results reveal that Fascin 1 has a yet undiscovered nuclear function as an epigenetic modulator of genes essential for amino acid metabolism.

We have previously identified a novel intra-tumoral dichotomy in triple-negative breast cancer (TNBC) based on the differential responsiveness to a reporter containing the Sox2 regulatory region-2 (SRR2), with reporter responsive (RR) cells being more stem-like than reporter unresponsive (RU) cells. Using bioinformatics, we profiled the protein-DNA binding motifs of SRR2 and identified Myc as one of the potential transcription factors driving SRR2 activity. In support of its role, Myc was found to be highly expressed in RR cells as compared to RU cells. Enforced expression of MYC in RU cells resulted in a significant increase in SRR2 activity, Myc-DNA binding, proportion of cellsexpressing CD44+/CD24-, chemoresistance and mammosphere formation. Knockdown of Myc using siRNA in RR cells led to the opposite effects. We also found evidence that the relatively high ERK activation in RR cells contributes to their high expression of Myc and stem-like features. Using confocal microscopy and patient samples, we found a co-localization between Myc and CD44 in the same cell population. Lastly, a high proportion of Myc-positive cells in tumors significantly correlated with a short patient survival. In conclusion, inhibition of the MAPK/ERK/Myc axis may be an effective approach in eliminating stem-like cells in TNBC.

Transfer RNAs (tRNAs, key molecules in protein synthesis) have not been investigated as potential prognostic markers in breast cancer (BC), despite early findings of their dysregulation and diagnostic potential. We aim to comprehensively profile tRNAs from breast tissues and to evaluate their role as prognostic markers (Overall Survival, OS and Recurrence Free Survival, RFS). tRNAs were profiled from 11 normal breast and 104 breast tumor tissues using next generation sequencing. We adopted a Case-control (CC) and Case-Only (CO) association study designs. Risk scores constructed from tRNAs were subjected to univariate and multivariate Cox-proportional hazards regression to investigate their prognostic value. Of the 571 tRNAs profiled, 76 were differentially expressed (DE) and three were significant for OS in the CC approach. We identified an additional 11 tRNAs associated with OS and 14 tRNAs as significant for RFS in the CO approach, indicating that CC alone may not capture all discriminatory tRNAs in prognoses. In both the approaches, the risk scores were significant in the multivariate analysis as independent prognostic factors, and patients belonging to high-risk group were associated with poor prognosis. Our results confirmed global up-regulation of tRNAs in BC and identified tRNAs as potential novel prognostic markers for BC.

Background To our knowledge, the Alberta Moving Beyond Breast Cancer (AMBER) Study is the first and only prospective cohort study of breast cancer survivors that includes objectively-measured physical activity (PA), sedentary behavior, health-related fitness (HRF), and biologic mechanisms focused on understanding breast cancer outcomes. The purpose of the present study was to report on the feasibility of recruitment, baseline measurement completion, and the representativeness of the first 500 participants. Methods AMBER is enrolling newly diagnosed stage I (≥T1c) to IIIc breast cancer survivors in Alberta, Canada. Baseline assessments are completed soon after diagnosis and include cardiorespiratory fitness, musculoskeletal fitness, body composition, objective and self-reported PA and sedentary behavior, lymphedema, and blood collection. Results Between July 2012 and November 2014, AMBER recruited its first 500 participants from a pool of 1,447 (35 %) eligible breast cancer survivors. Baseline HRF assessments were completed on ≥85 % of participants with the exception of upper body strength. Collection of ≥4 days/week of monitoring for the Actigraph GT3X® and ActivPAL® were obtained from 90 % of participants. Completion rates were also high for blood (99 %), lymphedema (98 %), and questionnaires (95 %) including patient-reported outcomes and correlates of exercise. The first 500 participants in AMBER are an average age of 56 years, 60 % are overweight or obese, and 58 % have disease stage II or III. Conclusion Despite the modest recruitment rate and younger age, AMBER has demonstrated that many newly diagnosed breast cancer survivors are willing and able to complete a wide array of sophisticated and physically demanding HRF and PA assessments soon after diagnosis. AMBER is a unique breast cancer survivor cohort that may inform future randomized controlled trials on lifestyle and breast cancer outcomes as well as PA behavior change in breast cancer survivors. Moreover, AMBER may also inform guidelines on PA, sedentary behavior, and HRF for improving breast cancer outcomes and survivorship.

The purpose of this study was to conduct an exploratory analysis of the START examining the effects of resistance exercise training (RET) and aerobic exercise training (AET) on sarcopenia, dynapenia, and associated quality of life (QoL) changes in breast cancer (BC) patients receiving adjuvant chemotherapy. Participants were randomized to usual care (UC) (n = 70), AET (n = 64), or RET (n = 66) for the duration of chemotherapy. Measures of sarcopenia [skeletal muscle index (SMI)] and dynapenia [upper extremity (UE) and lower extremity (LE) muscle dysfunction (MD)] were normalized relative to age-/sex-based clinical cut-points. QoL was assessed by the Functional Assessment of Cancer Therapy-Anemia (FACT-An) scales. At baseline, 25.5 % of BC patients were sarcopenic and 54.5 % were dynapenic with both conditions associated with poorer QoL. ANCOVAs showed significant differences favoring RET over UC for SMI (0.32 kg/m(2); p = 0.017), UE-MD (0.12 kg/kg; p

Breast cancer is the leading cause of cancer-associated deaths in women worldwide. Clinical biomarkers give information on disease progression and identify relevant biological pathways. A confounding factor that uncouples markers from disease outcome is the ability of tumor cells to mutate and evade clinical intervention. Therefore, we focussed on apoptotic genes that modulate tumor regression. Using gene and tissue microarray analyses, we identified an association of Bcl-2 interacting killer (Bik) with poor breast cancer prognosis. Bik prognostic ability was independent of Estrogen Receptor/Progesterone Receptor and Her2 status. Additionally, Bik was independent of anti-apoptotic Bcl-2, Bcl-xL, Mcl-1 and Bcl-w suggesting a complex mechanism of tumor promotion identified by Bik high tumors. Bik also stimulates autophagy, which can contribute to enhanced tumor fitness. We found a significant association between the autophagy marker ATG5 and Bik. Combined high expression level of ATG5 and Bik was a stronger predictor of outcome than either alone. Thus, our study identifies Bik as a novel, independent prognostic biomarker for poor outcomes in breast cancer and suggests that Bik-mediated autophagy contributes to disease recurrence.

Exercise is beneficial for breast cancer patients during chemotherapy, but their motivation to perform different types and doses of exercise is unknown. The purpose of this study was to examine the anticipated and experienced motivation of breast cancer patients before and after three different exercise programs during chemotherapy. Breast cancer patients initiating chemotherapy (N = 301) were randomized to a standard dose of 25–30 min of aerobic exercise, a higher dose of 50–60 min of aerobic exercise, or a combined dose of 50–60 min of aerobic and resistance exercise. Patient preference and motivational outcomes from the theory of planned behavior (i.e., perceived benefit, enjoyment, support, difficulty, and motivation) were assessed before and after the interventions. At pre-randomization, breast cancer patients were significantly (p

Background: Several candidate BMs have been explored in randomized trials of BEV across tumor types with the aim of identifying patients (pts) deriving the most substantial benefit from BEV therapy. In phase III trials, baseline pVEGF-A and pVEGFR-2 showed potential predictive value in metastatic BC (AVADO, AVEREL), pancreatic cancer (AViTA), and gastric cancer (AVAGAST; VEGF-A only). The randomized phase III BEATRICE trial, evaluating the addition of BEV to adjuvant chemotherapy in pts with triple-negative early BC, includes a comprehensive program to identify potential BMs predicting efficacy and toxicity of BEV therapy. We report results for baseline pVEGF-A and pVEGFR-2. Methods: After selection of chemotherapy (anthracycline and/or taxane), pts with T1a-T3 operable BC were randomized 1:1 to receive ≥4 cycles of chemotherapy either alone or with 1 year of BEV 5 mg/kg/wk equivalent. The primary endpoint is invasive disease-free survival (IDFS). pEDTA samples were collected from consenting pts at baseline (before treatment, after surgery), during study treatment, and at relapse. Pts were dichotomized using the median baseline concentration of each marker as the cut-off between high and low cohorts; further exploratory analyses were also performed by quartile. Results: Between Dec 2007 and Mar 2010, 2591 pts were enrolled. Of these, 1273 (49%) consented to the BM study and 1178 (45%) were included in the BM-evaluable population (BEP). Overall, the BEP was representative of the ITT population except for lower proportions of Asian pts (12% vs 24%). IDFS was similar in the BEP and ITT populations. Baseline characteristics were balanced between arms in the BEP. Baseline pVEGF-A showed neither prognostic nor predictive value using the median as the cut-off, although with a third quartile (Q3) cut-off there was a more pronounced but non-significant differentiation between treatments (HR 0.92 [low] vs 0.64 [high]). High baseline pVEGFR-2 showed potential predictive value for BEV efficacy (HR 1.24 [low] vs 0.61 [high]; p=0.029). Conclusions: Unlike trials in metastatic BC (AVADO, AVEREL), in the adjuvant setting, baseline pVEGF-A concentration did not show predictive value for BEV efficacy with a median cut-off. However, analyses using the Q3 cut-off suggest a trend toward predictive value. High baseline pVEGFR-2 was associated with greater BEV treatment effect, consistent with previous results in AVADO and AVEREL. The impact of differing biology in the adjuvant setting, lower median VEGF-A concentration than in the metastatic setting (77.0 vs 125.0–129.1 pg/mL), and the possible influence of surgery immediately before treatment require further investigation. Additional exploratory analyses are ongoing to provide better understanding of the BEATRICE dataset and the complex biology of angiogenesis, including additional markers, changes over time, and combination signatures. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P3-06-34.

Purpose Angiogenesis is critical for tumor growth and a promising therapeutic target. This review will summarize and analyze data from clinical trials of anti-angiogenic agents in the treatment of breast cancer (BC). Design A systematic search of PubMed and conference databases was performed to identify reports of randomized clinical trials investigating specific anti-angiogenic agents in the treatment of BC. Results and discussion Phase III trials in advanced BC have demonstrated a reduction in the risk of disease progression (22–52%), improved response rates and net improvements in progression-free survival of 1.2 to 5.5 months, but no significant improvements in overall survival with the addition of bevacizumab to chemotherapy. Results of phase III trials in early breast cancer have been inconsistent. Bevacizumab-containing regimens have also been associated with higher overall adverse event rates compared to chemotherapy alone. Phase III trials of the tyrosine kinase inhibitor sunitinib were negative, while randomized phase II trials of sorafenib and pazopanib have improved some outcomes when combined with chemotherapy or targeted therapy compared to controls. In addition to expected vascular class safety signals, tyrosine kinase inhibitors show “off-target” side effects. Ongoing clinical trials evaluating combinatorial strategies based on biological synergies and translational studies identifying biological predictors of response will be crucial to establish meaningful clinical benefits in selected BC populations. Conclusion Most trials of anti-angiogenic agents in BC have reported improved response rate and progression-free survival but no increase in overall survival compared to chemotherapy alone. Optimizing the therapeutic indices of these agents is a focus of ongoing research and will be critical to their future development.

Purpose: The mechanisms by which trastuzumab imparts clinical benefit remain incompletely understood. Antibody-dependent cellular cytotoxicity via interactions with Fcγ receptors (FcγR) on leukocytes may contribute to its antitumor effects. Single-nucleotide polymorphisms (SNP) in FCGR3A and FCGR2A genes lead to amino acid substitutions at positions 158 and 131, respectively, and affect binding of antibodies to FcγR such that 158V/V and 131H/H bind with highest affinity. This study aimed to determine whether high-affinity SNPs are associated with disease-free survival (DFS) among patients with HER2-positive nonmetastatic breast cancer. Experimental Design: Genomic DNA was isolated from 1,286 patients enrolled in a trial of adjuvant trastuzumab-based chemotherapy. Genotyping was conducted using Sanger sequencing and Sequenom mass spectrometry. Results: Patient samples (N = 1,189) were successfully genotyped for FCGR3A and 1,218 for FCGR2A. Compared with the overall results of the BCIRG006 study, in the subset of patients genotyped in this analysis, a less robust improvement in DFS was observed for the trastuzumab arms than control arm (HR, 0.842; P = 0.1925). When stratified for prognostic features, the HR in favor of trastuzumab was consistent with that of the overall study (HR, 0.74; P = 0.036). No correlation between DFS and FCGR3A/2A genotypes was seen for trastuzumab-treated patients (158V/V vs. V/F vs. F/F, P = 0.98; 131H/H vs. H/R vs. R/R, P = 0.76; 158V/V and/or 131H/H vs. others, P = 0.67). Conclusion: This analysis evaluating the association between FCGR3A/2A genotypes and trastuzumab efficacy in HER2-positive breast cancer did not show a correlation between FCGR3A-V/F and FCGR2A-H/R SNPs and DFS in patients treated with trastuzumab. Clin Cancer Res; 18(12); 3478–86. ©2012 AACR.

FABP7 has been implicated in tumour cell proliferation, cell migration, and poor prognosis in patients with high-grade astrocytoma and melanoma. In this study, we examine FABP7 expression in a cohort of 176 primary breast cancers by gene profiling and tissue microarray immunostaining. We show that FABP7 is significantly up-regulated in triple-negative breast cancer. Elevated FABP7 levels are associated with poor prognosis, absence of oestrogen and progesterone hormone receptors (ER, PR) and HER2, increased cell proliferation, and high tumour grade. Depletion of FABP7 in the ER/PR-negative cell line, MDA-MB-435S, significantly reduced cell growth rate and sensitized the cells to growth inhibition by omega-3 docosahexaenoic acid (DHA). A target of DHA-bound FABP7 in the nucleus is RXRβ, a retinoid-activated nuclear receptor that functions as a transcription factor by either homodimerizing or heterodimerizing with other nuclear receptors such as PPARs. Based on our microarray data, RXRβ, like FABP7, is an adverse prognostic factor for breast cancer. We propose that the DHA-FABP7-RXRβ pathway promotes cell survival/proliferation in triple-negative breast cancer. Targeting this pathway may thus provide an alternate route for the treatment of triple-negative breast cancer.