bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract P6-17-06: Relevance of the hyaluronidase-1 (HYAL1) in brain metastasis formation of breast cancer patients
Authors: Oliveira-Ferrer L , Anna M , Wickman H , Matschke J , Schumacher U , Milde-Langosch K , Müller V , Witzel I
Published: May 2016

The incidence of brain metastases in breast cancer (BMBC) patients has increased in the last years and represents the major life-limiting problem for metastatic breast cancer (BC) patients. However, the knowledge about tumor cell invasion in the brain is still very limited and new markers for brain metastasis incidence are urgently needed in order to early detect high risk patients.

In a recent study based on cDNA microarray data of primary breast tumors, we could show that mRNA expression of certain glycosylation enzymes significantly correlates with an organ-specific metastatic spread. Interestingly, hyaluronan synthase 2 (HAS2) and hyaluronidase-1 (HYAL1), both genes involved in hyaluronan (HA) metabolism, showed an independent prognostic value and a significant correlation with brain metastasis formation.

In order to corroborate the role of these enzymes at protein level, we examined the expression of HYAL1 and HAS2 on a tissue microarray including 200 primary BC samples. Here, the prognostic impact of HAS2 could not be validated, whereas for HYAL1 shorter disease free survival (DFS) was observed for patients with high HYAL1-expression levels. This trend could be additionally verified in a second cohort of 107 BC samples, using western blot analysis. Moreover, significantly higher HYAL1 expression was detected among primary tumors with subsequent brain metastases compared with those without brain metastases using immunohistochemistry (IHC).

No impact of HYAL1 expression on disease progression of BCBM patients could be observed after analysis of 87 brain metastasis samples. Here, quantification of tumor-associated HA revealed a significant positive correlation with triple negative tumors and a trend towards shorter progression-free survival.

Taken together, our data suggest a role of the enzyme HYAL1 for progression and especially for the development of brain metastases in breast cancer patients.

Citation Format: Oliveira-Ferrer L, Anna M, Wickman H, Matschke J, Schumacher U, Milde-Langosch K, Müller V, Witzel I. Relevance of the hyaluronidase-1 (HYAL1) in brain metastasis formation of breast cancer patients. [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 P6-17-06.
Abstract PD2-08: Potential role of prolactin signaling in development and growth of the lobular subtype of breast cancer
Authors: Oesterreich S , Katz TA , Logan G , Levine K , Nagle A , Huo Z , Tseng GC , Rui H , Lee AV , Butler LM
Published: May 2016

Invasive lobular carcinoma (ILC) is the eighth most frequently diagnosed cancer in any organ, and accounts for 8-11% of breast cancer. This histological subtype is characterized by loss of E-cadherin, and favorable prognostic factors, such as low Ki67 and high rates of ER/PR-positive tumors. Only recently is the lobular subtype gaining recognition as a distinct disease, displaying a unique growth pattern, unique molecular changes in addition to loss of E-cadherin, and evidence for late recurrences and reduced response to targeted endocrine therapy. It is widely accepted that a late age at first full term birth (FFTB) increases a women's risk for breast cancer. Interestingly, several published epidemiological studies have shown that the increased risk after a late age at FFTB is preferential for the lobular subtype of breast cancer compared to the ductal subtype. We therefore hypothesized that pregnancy hormones like prolactin play an integral role in the development and progression of ILC. Interrogation of the Cancer Genome Atlas (TCGA) data revealed a high expression of milk protein genes as well as prolactin signaling molecules, specifically Stat5a and Stat5b in lobular carcinomas compared to ductal carcinomas. We developed a lactation score including 7 milk protein genes and found that in the TCGA data set ILC tumors have a significantly higher lactation score than IDC tumors. Additionally, we found that ILC cell lines express increased prolactin receptor mRNA and protein levels compared to IDC cell lines. Prolactin treatment in ILC and IDC cells reveals divergent signaling pathways - prolactin stimulates ERK activation in IDC but not ILC cells. We are currently further delineating the prolactin signaling pathways, and resulting phenotypes, comparing ILC and IDC cells. We expect these experiments to move the field forward by establishing a relationship between prolactin and lobular carcinoma.

Citation Format: Oesterreich S, Katz TA, Logan G, Levine K, Nagle A, Huo Z, Tseng GC, Rui H, Lee AV, Butler LM. Potential role of prolactin signaling in development and growth of the lobular subtype of 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 PD2-08.
Abstract PD3-03: Inhibition of the autocrine IL6-JAK2-STAT3-calprotectin axis as targeted therapy for HR-/HER2+ breast cancers
Authors: Silva J , Rodriguez-Barrueco R , Yu J , Kalinsky K , Maurer M , Califano A
Published: May 2016

Although HER2+ tumors are commonly considered as a single entity there is increasing evidence indicating that important intrinsic differences associated to hormone receptor status (HR) exist. Indeed, while HR+/HER2+ patients benefit from anti-hormonal and HER2 targeted therapies, the outcome of HR-/HER2+ patients strongly depends on their response to chemotherapy as well as anti-HER2 therapy. To identify genes that represent novel mechanistic dependencies in HR-/HER2+ breast cancer cells, we designed an integrative approach that combines functional genomic (RNAi screens) and computational algorithms to interrogate regulatory networks reverse engineered.

Our analysis identified STAT3 as a de novo master regulator (MR) gene associated to HER2 mediated transformation in HR- breast cancer cells. Importantly, we demonstrate that aberrant STAT3 activity is necessary to maintain HR-/HER2+ tumor state, thus representing a non-oncogene dependency. Mechanistically we found that HR-/HER2+ breast tumors secrete high levels of IL-6. This acts in an autocrine way to induce the activation of STAT3 via the canonical JAK2/STAT3 pathway. Aberrant STAT3 activity induces upregulation and secretion of the S100A8/9 complex (Calprotectin) triggering a second autocrine stimulus that enhances proliferation and survival via ERK1/2 and AKT. As a result, disruption of the "IL6-JAK2-STAT3-S100A8/9 cascade" compromises HR-/HER2+ cell viability suggesting that the components of this pathway represent putative therapeutic targets in HR-/HER2 tumors.

Importantly, small molecule inhibitors and blocking antibodies for components of this double autocrine loop are already FDA approved or in clinical trials. Here, we demonstrate in vivo that blocking the IL-6 receptor (IL-6R) with the humanized monoclonal antibody Tocilizumab, STAT3 inactivation with the dual JAK1/2 inhibitor Ruxolitinib or Calprotectin inhibition with the small molecule inhibitor Tasquinimod), alone or in combination with anti HER2 therapies, compromises the viability of HR-/HER2+breast cancer cells. Availability of FDA-approved inhibitors to target this novel mechanism represents an exciting opportunity for rapid translation of these findings to the clinics.

Citation Format: Silva J, Rodriguez-Barrueco R, Yu J, Kalinsky K, Maurer M, Califano A. Inhibition of the autocrine IL6-JAK2-STAT3-calprotectin axis as targeted therapy for HR-/HER2+ breast cancers. [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 PD3-03.
Abstract PD7-02: Multiplex Identification of genetic etiologies among women with bilateral breast cancer using a 25-gene hereditary cancer panel
Authors: Weitzel JN , Blazer KR , Nehoray B , Kidd J , Slavin TP , Solomon I , Niell-Swiller M , Rybak C , Saam J , Lancaster J
Published: May 2016

Background: Multiple primary cancers within an individual is one of the hallmarks of hereditary cancer predisposition. Technical advances in sequencing and identification of additional cancer susceptibility genes have led to multi-gene panel approaches to determine if patient cancers have a heritable cause. Simultaneous testing of multiple breast cancer associated genes to determine the prevalence, spectrum and combinations of mutations has not yet been evaluated in a large set of patients with two breast cancers.

Methods: Patients with two breast cancer diagnoses were identified from 80,748 consecutive cases that underwent a 25-gene hereditary cancer panel test at a commercial diagnostic laboratory. Patient clinical data were obtained by healthcare provider report on test requisition forms.

Results: Of 3,182 patients with two breast cancers, 13.3 % (n=424) had a pathogenic variant (PV) in at least one of 19 genes on the panel. PVs were most prevalent within BRCA1, representing 26.3% of the PVs, followed by BRCA2 (21.9%), CHEK2 (13.5%), PALB2 (11%) and ATM (11%). Fourteen patients had PVs in two different genes, including five with CHEK2/PALB2 in combination, which was the only combination observed more than once. PVs were more common among the 33.8% of patients with synchronous breast cancers compared to those with metachronous diagnoses (10.8% vs. 15%) and in the 58.4% who were first diagnosed younger than 50 years old (15.7% vs. 10%).

Conclusion: Multiplex testing in women with two primary breast cancers identifies a relatively high percentage with a PV, including those whose first diagnosis was older than 50. While still the most prevalent and clear cause, fewer than half of the PVs identified were in BRCA1 or BRCA2. The observation of 5 cases with double CHEK2/PALB2 PVs is statistically unexpected given their individual prevalence in other studies of high risk women with breast cancer, suggesting a possible synergistic or additive effect. This study adds to our understanding of susceptibility to multiple primary breast cancers, and reaffirms the importance of this circumstance as a prompt for genetic testing. Nonetheless, as 66% of cases had metachronous tumors, it would be preferable to identify those at risk with heritable cancer syndromes at their first diagnosis and enable subsequent prevention or early detection.

Citation Format: Weitzel JN, Blazer KR, Nehoray B, Kidd J, Slavin Jr TP, Solomon I, Niell-Swiller M, Rybak C, Saam J, Lancaster J. Multiplex Identification of genetic etiologies among women with bilateral breast cancer using a 25-gene hereditary cancer panel. [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 PD7-02.
Abstract PL2: Clonal Dynamics and Breast Cancer Subtypes
Authors: Aparicio S
Published: May 2016

Breast cancers exhibit inter-patient and intra-tumoural genomic variability which underpins our understanding of intrinsic drivers of the disease. The advent of next generation sequencing methods and informatics approaches has redefined the landscape of primary breast cancer subtypes into many more molecularly defined patient subgroups than had been previously appreciated, thus redefining our understanding of inter-patient variation. These studies have implications for the understanding of driver mutation context in primary cancers and should pattern future clinical and molecular studies of primary breast cancers. Closely linked to inter-patient genomic variability is the notion that most cancers are ecosystems of evolving cellular clones has implications for biological understanding and clinical application. The evolution of clonal composition has particular significance when evidence of positive or negative selection can be associated with the clonal genotype or epigenotype. Over the last 5 years next generation sequencing of tumours and methods for single cell analysis have opened up this approach for solid epithelial malignancies. I will discuss the implications of clonal evolution for cancer medicine and biological studies of cancer with reference to breast cancers. We have developed informatics approaches to population level clonal analysis and extended these to single cell measurements of genotypes. I shall discuss our more recent data from single cell sequencing and clonal analysis applied to clonal evolution of patient derived tumour xenografts, to illustrate the impact of clonal evolution on biological studies of breast cancer in model systems.

Citation Format: Aparicio S. Clonal Dynamics and Breast Cancer Subtypes. [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 PL2.
Abstract S6-03: Anastrozole versus tamoxifen for the prevention of loco-regional and contralateral breast cancer in postmenopausal women with locally excised ductal carcinoma in-situ (IBIS-II DCIS)
Authors: Cuzick J , Forbes JF , Sestak I , Howell A , Bonanni B , Bundred N , Levy C , von Minckwitz G , Eiermann W , Neven P , Stierer M , Holcombe C , Coleman RE , Jones LJ , Ellis I
Published: May 2016

Background: Third generation aromatase inhibitors are a more effective treatment option than tamoxifen for hormone receptor positive invasive breast cancer in postmenopausal women. However, it is not known whether anastrozole is more effective than tamoxifen in preventing the recurrence of breast cancer in women with hormone receptor (HR) positive ductal carcinoma in situ (DCIS). Here, we compare the efficacy of anastrozole versus tamoxifen in postmenopausal women with HR positive DCIS.

Methods: A multi-centre randomised placebo-controlled trial of 1mg/day anastrozole (oral) vs. 20mg/day tamoxifen (oral) for five years was conducted in 2980 postmenopausal women with locally excised HR positive DCIS. The primary endpoint was to determine if anastrozole is at least as effective as tamoxifen in loco-regional control and prevention of contralateral disease. Secondary endpoints included breast cancer mortality, other cancers, cardiovascular disease, fractures, adverse events and non-breast cancer deaths. All analyses were done on an intention-to-treat basis and Cox proportional hazard were used to compute hazard ratios and corresponding confidence intervals for recurrence.

Results: Between 2003 and 2012, a total of 2980 postmenopausal women were recruited into the IBIS-II DCIS trial. 1471 women were randomly assigned to receive anastrozole and 1509 women tamoxifen. Median follow-up for this first analysis is 6.8 years and 131 breast cancer recurrences have been recorded. Median age was 60.3 years (56.1-64.6), median BMI was 26.7 (23.6-30.7), and 45.6% of women had used hormone replacement therapy (HRT) before joining the trial. Of the 131 women with recurrent disease, 77 had a loco-regional recurrence and 51 reported contralateral disease. A total of 61 deaths were recorded. We will present a comprehensive analysis of the efficacy of anastrozole and tamoxifen for preventing loco-regional/contralateral breast cancer and major adverse events by intention to treat (ITT).

Conclusions: To follow.

Citation Format: Cuzick J, Forbes JF, Sestak I, Howell A, Bonanni B, Bundred N, Levy C, von Minckwitz G, Eiermann W, Neven P, Stierer M, Holcombe C, Coleman RE, Jones LJ, Ellis I. Anastrozole versus tamoxifen for the prevention of loco-regional and contralateral breast cancer in postmenopausal women with locally excised ductal carcinoma in-situ (IBIS-II DCIS). [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 S6-03.
Abstract B26: Leptin affects proliferation, stem cells and chemotherapeutic treatment outcome of pancreatic cancer: A link to health disparity
Authors: Harbuzariu Adriana , Daley-Brown Danielle S. , Harmon Tia L. , Garrison Robin C. , Beech Derrick J. , Cason Frederick D. , Klug Christopher , Gonzalez-Perez Ruben R.
Published: May 2016

Background: Pancreatic cancer (PC) is an aggressive disease that is in most cases advanced at the time of diagnosis. The overall 5-year survival rate for PC patients is less than 5 %. African-Americans show the highest incidence of obesity and PC as well as mortality rate. How obesity is associated to PC incidence and health disparity is still an unanswered question. High levels of leptin shown by obese African American patients could impact negatively on PC. We hypothesize that leptin signaling induces PC proliferation, stem cells (PCSC) and drug resistance.

Methods: Human PC cell lines derived from primary pancreatic adenocarcinoma (Panc-1 and MiaPaCa-2-highly aggressive, BxPC-3-less aggressive) and from metastatic tumor (AsPC-1) were challenged with leptin and chemotherapeutic agents. Cell proliferation, apoptosis rate, and the expression levels of molecular markers were analyzed. PCSC markers were determined in cell lines and tumor biopsies. Effects of leptin and chemotherapeutics on PC-tumorsphere formation were also determined.

Results: Leptin stimulated proliferation of PC cells, induced the expression of PCSC markers and increased tumorsphere formation. Moreover, leptin impaired the efficacy of chemotherapeutics on PC cells.

Conclusions: Present data suggest leptin is a proliferative and survival factor for PC that reduces chemotherapeutic effectiveness. These observations are of paramount importance for obese African American PC patients, who show the highest levels of leptin, poor prognosis and outcome of chemotherapeutic treatment.

Acknowledgements: this work was supported by the DOD W81XWH-13-1-0382; NIH/SBIR 1R41CA183399-01A1; Pilot Project Award from MSM/Tuskegee University/UAB Cancer Center Partnership grant 5U54CA118638; PC SPORE Grant from UAB to RRGP, and facilities and support services at Morehouse School of Medicine (1G12RR026250-03; NIH RR03034 and 1C06 RR18386).

Citation Format: Adriana Harbuzariu, Danielle S. Daley-Brown, Tia L. Harmon, Robin C. Garrison, Derrick J. Beech, Frederick D. Cason, Christopher Klug, Ruben R. Gonzalez-Perez. Leptin affects proliferation, stem cells and chemotherapeutic treatment outcome of pancreatic cancer: A link to health disparity. [abstract]. In: Proceedings of the Eighth AACR Conference on The Science of Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; Nov 13-16, 2015; Atlanta, GA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2016;25(3 Suppl):Abstract nr B26.
Abstract B82: Predictors of intention to obtain colorectal cancer screening among African American men in a state fair setting
Authors: Rogers Charles R. , Goodson Patricia , Dietz Lindsey R. , Okuyemi Kola S.
Published: May 2016

Racial disparities in health among African American men in the United States are appalling: African American men have the highest mortality and incidence rates from colorectal cancer (CRC) compared to all other ethnic, racial, and gender groups. Juxtaposed to their white counterparts, African American men have CRC incidence and mortality rates 25% and 50% higher, respectively. CRC is a treatable and preventable condition when detected early, yet the intricate factors influencing African American men's intention to screen remain understudied. Employing a non-experimental, on-line survey research design at the Minnesota State Fair, the purpose of this study was to explore whether male role norms affect knowledge, attitudes, and intention to screen for CRC directly, or mediated by perceived barriers, among 297 African American men. As hypothesized, these Minnesota men (ages 18-65) lacked appropriate CRC knowledge: only 33% of the sample received a “passing” knowledge score (85% or better). In a logistic regression model, the three factors significantly associated with a higher probability of obtaining CRC screening were age, perceived barriers, and perceived subjective norms. Findings from this study provide a solid basis for informing health policy and designing health promotion and early-intervention CRC prevention programs that are responsive to the needs of African American men in Minnesota and beyond.

Citation Format: Charles R. Rogers, Patricia Goodson, Lindsey R. Dietz, Kola S. Okuyemi. Predictors of intention to obtain colorectal cancer screening among African American men in a state fair setting. [abstract]. In: Proceedings of the Eighth AACR Conference on The Science of Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; Nov 13-16, 2015; Atlanta, GA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2016;25(3 Suppl):Abstract nr B82.
Abstract C60: Injectable, biodegradable micro- and nano-particles loaded with prodigiosin-based drug for localized anticancer drug delivery
Authors: Obayemi John D. , Malatesta Karen A. , Odusanya Olushola S. , Yiporo Danyuo , Yu Wei , Uhrich Kathryn E. , Soboyejo Winston O.
Published: May 2016

This paper presents the synthesis and physicochemical characterization of injectable, multi-functional, biodegradable poly (D,L-lactide-co-glycolide) (PLGA)-loaded micro- and nano-particles. These particles were loaded with an anticancer drug from prodigiosin (PG), which was obtained from bacteria, Serratia marcescens subsp. Marcescens. The release of paclitaxel (PT) was also tested as a control. The PG and PT were encapsulated using a single-emulsion solvent evaporation technique with PLGA as a polymer matrix and poly-(vinyl alcohol) (PVA) as an emulsifier. The dependence of particle size and morphology on processing conditions was also evaluated. In vitro release studies were used to elucidate drug loading efficiency, encapsulation efficiency and microparticle morphology using a combination of UV-visible (UV-Vis) spectrophotometry, optical Microscopy, scanning electron microscopy (SEM), differential scanning calorimetry (DSC), and atomic force microscopy (AFM). The implications of the results are then explored using MDA-MB-231 cells (breast cancer cells) for the development of injectable, multi-functional, polymeric micro- and nano-particles for the controlled release of cancer drugs and the localized treatment of cancer.

Citation Format: John D. Obayemi, Karen A. Malatesta, Olushola S. Odusanya, Danyuo Yiporo, Wei Yu, Kathryn E. Uhrich, Winston O. Soboyejo. Injectable, biodegradable micro- and nano-particles loaded with prodigiosin-based drug for localized anticancer drug delivery. [abstract]. In: Proceedings of the Eighth AACR Conference on The Science of Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; Nov 13-16, 2015; Atlanta, GA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2016;25(3 Suppl):Abstract nr C60.
Abstract A14: Hedgehog inhibition delays cancer growth in C3(1)/Tag transgenic murine mammary carcinoma model
Authors: Makarova Anastasia M. , KC Subheksha , Dolorito John A. , Khaimskiy Yefim I. , Epstein Ervin H.
Published: May 2016

In 1941 Apperly hypothesized that differential sunlight exposure is the cause of the latitudinal gradient in breast cancer mortality, and subsequently the effect was postulated to be due to the tumor inhibitory properties of skin produced vitamin D3. We first tested this latitudinal hypotheses using the C3(1)/Tag transgenic murine mammary carcinoma model and found that ultraviolet radiation (UVR) significantly prolongs tumor-free and overall survival but that dietary D3 has no such effect. The mechanism of this anti-mammary carcinogenesis effect of UVR is unclear but there is evidence that D3 is a potent inhibitor of the hedgehog (HH) - signaling pathway. To test this mechanistic hypothesis we compared the effects of the small molecule Smo-acting HH pathway inhibitor (HHI) BMS-833923/XL139 with those of UVR on breast cancer development in C3(1)/Tag mice. We have found that oral administration of the pharmacologic HHI delays Tag mammary carcinogenesis with an efficacy similar to that of UVR treatment (350 mJ/cm2, 3x/week), thus, time to first palpable tumor was delayed by 9 days by UVR treatment (p=0.016) and by 14 days by HHI treatment (p=0.009), compared control mice. Overall survival was delayed by 9 days by UVR treatment (p=0.19) and by 15 days by HHI treatment (p=0.011). And, we have found that HHI, like UVR, acts at an early stage of carcinogenesis (i.e. reduction of histologic changes at age 12 weeks), and initial studies of UVR + HHI treatments suggest that HHI and UVR have overlapping effects. Taken together our results are consistent with (albeit by no means proof of) our hypothesis that inhibition of mammary carcinogenesis by UVR-produced vitamin D3 underlies the latitudinal gradient and is mediated via HH inhibition. Moreover, our results suggest more broadly that HHI treatment before tumors develop may have more of an anti-cancer effect than does HHI treatment after tumors already have become clinically apparent.

Note: This abstract was not presented at the conference.

Citation Format: Anastasia M. Makarova, Subheksha KC, John A. Dolorito, Yefim I. Khaimskiy, Ervin H. Epstein, Jr. Hedgehog inhibition delays cancer growth in C3(1)/Tag transgenic murine mammary carcinoma model. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research; Oct 17-20, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(2_Suppl):Abstract nr A14.
Abstract A32: MCL1 gene amplification in breast cancer is associated with TNBC status and can respond to a sorafenib/vorinostat regimen
Authors: Ross JS , Wang K , Johnson A , Watson J , Hatzis C , Pusztai L , Chmielecki J , Yelensky R , Lipson D , Elvin JA , Vergilio J , Suh J , Miller VA , Dicke K , Stephens PJ , Ali SM
Published: May 2016

Background: MCL1 encodes the induced myeloid leukemia cell differentiation protein Mcl-1, a member of the BCL-2 family which functions to inhibit apoptosis. Mcl-1 over-expression has been associated with high tumor grade and adverse prognosis in triple negative breast cancer (TNBC) but therapies specifically leading to inhibition of MCL-1 have not been identified.

Methods: Comprehensive genomic profiling (CGP) using hybridization capture of 3,769 exons from 315 cancer-related genes and 47 introns of 19 genes commonly rearranged in cancer was applied to ≥50ng of DNA extracted from 2,824 consecutive BC and sequenced to high, uniform median coverage (>600X). The original primary BC was assayed in 44% of cases and a sample from a metastatic focus was assayed in 56% of cases.

Results: Of 2824 consecutive BC cases, 200 (7.1%) cases harbored MCL1 amplification. Of these MCL1-amplified cases, 146 (73%) were TNBC and 54 were non-TNBC (p<0.0001). Twelve of the latter cases (22%) were ERBB2 (HER2) amplified and slide-based HER2 status concordance with CGP was 99%. MCL1 amplification was also observed in the TCGA dataset for 32/123 (26%) of TNBC (p=0.008). Of the MCL1 amplified TNBC cases, 88% were high grade and 98% were stage IV at the time of CGP. Genes co-altered within MCL1 amplified TNBC included TP53 (86%), MYC (41%), MYST3 (21%), LYN (20%), CCNE1 (19%), PIK3CA (18%), and AKT3 (15%). Two MCL1 amplified TNBC patients were treated with a multi-drug regimen based on sorafenib and vorinostat and experienced significant clinical benefit.

Conclusions: MCL1 amplification is a frequent feature in advanced stage and high grade TNBC, and correspondingly such MCL1 amplified tumors very seldom harbor co-amplifications of ERBB2. Clinical observation suggests that treatment with sorafenib and vorinostat in heavily pre-treated MCL1 amplified patients may be correlated with clinical benefit, consistent with historic preclinical investigation. These preliminary findings suggest that MCL1 amplified TNBC may be able to benefit from combination targeted therapy, and warrant further systematic investigation.

Citation Format: JS Ross, K Wang, A Johnson, J Watson, C Hatzis, L Pusztai, J Chmielecki, R Yelensky, D Lipson, JA Elvin, J Vergilio, J Suh, VA Miller, K Dicke, PJ Stephens, SM Ali. MCL1 gene amplification in breast cancer is associated with TNBC status and can respond to a sorafenib/vorinostat regimen. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research; Oct 17-20, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(2_Suppl):Abstract nr A32.
Abstract B10: mTORC1 directly phosphorylates and activates ERα upon estrogen stimulation
Authors: Alayev Anya , Holz Marina K.
Published: May 2016

Approximately 75% of breast cancers are estrogen receptor (ERα) positive, underscoring the dependence of cancer cells on estrogen for growth and survival. Patients treated with endocrine therapy often develop resistance, either de novo or acquired, which is caused by aberrations within the growth factor signaling pathways. The mechanistic target of rapamycin complex 1 (mTORC1) has emerged as a critical node in estrogenic signaling. We have previously shown that mTORC1 can phosphorylate and activate ERα on S167 via its effector the 40S ribosomal S6 kinase 1 (S6K1). Presently, we have uncovered a direct link between mTORC1 and ERα. We found that ERα binds to regulatory-associated protein of mTOR (Raptor) and causes it to translocate to the nucleus upon estrogen stimulation. Additionally, we identified mTOR as the kinase that directly phosphorylates ERα and activates transcription of ER target genes. Our findings show a direct link between mTORC1 and ERα, which further implicates mTORC1 signaling in the pathogenesis of ER-positive breast cancer and provides additionale rationale for the FDA-approved use of mTORC1 inhibitors in combination with endocrine agents for treatment of this disease .

Citation Format: Anya Alayev, Marina K. Holz. mTORC1 directly phosphorylates and activates ERα upon estrogen stimulation. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research; Oct 17-20, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(2_Suppl):Abstract nr B10.
Abstract B30: Ex vivo culture of breast cancer patient-derived xenografts in modified 3-D hyaluronan hydrogels
Authors: Sablatura Lindsey K. , Kavuri MeghaShyam , Richards Priya , Harrington Daniel A. , Ellis Matthew J. , Farach-Carson Mary C.
Published: May 2016

The high attrition rate of potential anti-cancer drugs entering clinical trials indicates the need for more predictive pre-clinical model systems. Standard compound screening is high-throughput and relatively inexpensive, but conducted with non-representative cells in a non-physiological environment. Three-dimensional (3D) culture systems provide cells a more physiological microenvironment including relevant extracellular matrix (ECM) cues and cell-cell interactions. Patient-derived xenograft (PDX) culture preserves tumor heterogeneity while mitigating harsh selective pressures induced by traditional 2D culture methods. However, in vivo mouse models are slow and expensive, often with low “take” rates that limit the numbers of drugs and drug combinations that can be tested. Within the present study, we established the direct culture of breast cancer PDX cells in a hyaluronan-based hydrogel matrix that is amenable to automated dispensing. Washington University Human in Mouse (WHIM) breast cancer PDX tumors were dissociated and PDX cell were encapsulated in 3D hydrogels for in vitro culture. In vitro cultured PDX cells showed high viability over 7 days, as assessed by standard live/dead assay. Cell morphology and marker retention were also examined using immunofluorescence staining for markers including ER, EpCAM, and vimentin, and were found to be subtype-consistent. Ongoing studies seek to expand the PDX tumors in hydrogel matrices such that they may be useful for a variety of applications including drug screening, proteome and kinome activation analysis, and other applications that will enable the development of personalized therapies suited to individual breast cancer patients presenting with various disease subtypes.

Citation Format: Lindsey K. Sablatura, MeghaShyam Kavuri, Priya Richards, Daniel A. Harrington, Matthew J. Ellis, Mary C. Farach-Carson. Ex vivo culture of breast cancer patient-derived xenografts in modified 3-D hyaluronan hydrogels. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research; Oct 17-20, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(2_Suppl):Abstract nr B30.
Abstract B45: De novo generation of highly aggressive human breast tumors by rapid serial passaging of oncogene-transduced starting populations of purified normal cells
Authors: Lefort Sylvain , Pellacani Davide , Nguyen Long , Eaves Connie J.
Published: May 2016

Human breast cancers are clonal outgrowths that have diversified genetically and biologically by the time they are clinically evident. However, early events that predicate malignant transformation of normal human mammary epithelial cells and the importance of the specific cell type to first be altered has remained elusive. We have developed an in vivo system that allows analysis of the initial steps of tumorigenesis from defined subsets of normal human mammary cells within 8 weeks using a single (KRASG12D) or multiple oncogenes. Using a protocol in which we first isolate biologically, transcriptionally and phenotypically different human mammary cell types by FACS using EpCAM and CD49f or CD10 as distinguishing markers, and then transduce the cells with lentiviral constructs, and immediately transplant them into highly immunodeficient NOD/RAG1-/-IL2rγ-/- (NRG) female mice, we have found that invasive ductal carcinomas expressing a variety of basal and luminal-associated markers (such as CK5, CK8/18, EGFR and MUC1) can be obtained at high frequency from purified starting populations of both basal cells and luminal progenitors within 8 weeks. Many tumours could be passaged both in vivo and under multiple conditions in vitro (on collagen, in suspension “mammopshere” culture or 3D matrigel cultures). After only 4passages in vivo, an aggressive tumor cell line that is tumorigenic within 6 weeks in vivo at unit efficiency has emerged. Intravenous injections shown that these cells can colonize multiple organs including the lung, kidney and spleen. Molecular characterization of these aggressive tumors has established that they are EpCAM+ and CD44+. Additional examples of de novo tumors able to be serially passaged are now being characterized. This new tumorigenesis model will allow us to compare the sequential changes that accompany and underlie the full process of malignant progression and should thus enable key molecular determinants that drive cancer initiation and progression to be identified.

Citation Format: Sylvain Lefort, Davide Pellacani, Long Nguyen, Connie J. Eaves. De novo generation of highly aggressive human breast tumors by rapid serial passaging of oncogene-transduced starting populations of purified normal cells. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research; Oct 17-20, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(2_Suppl):Abstract nr B45.
Abstract B52: Overcoming phenotypic heterogeneity and plasticity in basal-like breast cancer through targeting adaptive pathway use
Authors: Risom Tyler , Langer Ellen , Rantala Juha , Alvarez Mariano , Johnson-Camacho Katie , Pelz Carl , Wang Nicholas , Spellman Paul , Califano Andrea , Gray Joe , Sears Rosalie
Published: May 2016

The use of small-molecule kinase inhibitors is a promising therapeutic strategy for the management of breast cancer. Positive responses to these agents, however, are often transient, and acquired resistance arises in the course of weeks to months. Recent studies have demonstrated that phenotypic plasticity in cancer cell populations can provide adaptive resistance to small-molecule kinase inhibitors, whereby cancer cells transition to drug-tolerant phenotypic states, reliant on compensatory survival and proliferative signaling pathways. Drug combinations and sequences can prevent this adaptive resistance to targeted therapy, however, nominating effective drug pairs is challenging. In this study, we seek to identify such drug combinations through first identifying single agent therapeutics that reduce phenotypic heterogeneity and enrich distinct cell-states with common pathway reliance. To do so, we focus on phenotypic heterogeneity in tumor-cell lineage-state; using immunofluorescent staining against markers of the luminal, basal, and mesenchymal lineages, we combine high-throughput drug screening with high-content imaging and pursue small-molecule inhibitors that reduce phenotypic heterogeneity and promote the accumulation of particular lineage-states in residual cell populations. We observe pronounced lineage-state heterogeneity in Triple-Negative tumors and Basal-Like breast cancer cell lines, and find that the lineage-state distributions are greatly influenced by numerous therapeutics. MEK and PI3K/mTOR inhibitors in particular induce robust time- and dose-dependent alterations is lineage-state distribution in residual cell populations, selecting for a cell population enriched in, or depleted of a basal lineage-state, respectively. Through gene expression profiling and master-regulator analysis of active transcriptional states in the residual cell populations, we are able to identify compensatory-signaling pathways. We demonstrate that combining MEK and PI3K/mTOR inhibitors with agents targeting these compensatory pathways induces synergistic antiproliferative effects.

Citation Format: Tyler Risom, Ellen Langer, Juha Rantala, Mariano Alvarez, Katie Johnson-Camacho, Carl Pelz, Nicholas Wang, Paul Spellman, Andrea Califano, Joe Gray, Rosalie Sears. Overcoming phenotypic heterogeneity and plasticity in basal-like breast cancer through targeting adaptive pathway use. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research; Oct 17-20, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(2_Suppl):Abstract nr B52.
Abstract B53: Genomic redefinition of Her2+ breast shows role of estrogen and androgen signaling
Authors: Daemen Anneleen , Rinaldi Jacob , Manning Gerard
Published: May 2016

Many breast cancers are driven by amplification and overexpression of the Her2 receptor tyrosine kinase. We searched the TCGA and Metabric genomic datasets to define and better understand the genetic drivers of Her2+ cancer. We defined Her2+ tumors as those with a ploidy-corrected copy number for Her2 of 4 or more, encompassing 12% of breast tumors in both cohorts. This population showed high concordance with Her2 IHC/FISH status, but weak concordance with intrinsic PAM50 subtype: 45% of Her2+ breast tumors were Her2-enriched, 48% luminal A or B, and 7% basal-like. We characterized this transcriptionally heterogeneous Her2+ population using a meta-consensus clustering approach. Her2 amplification--though a driver--had limited impact on the transcriptional profile of each tumor. Her2+ tumors were rather additionally driven by estrogen or androgen signaling, with their transcriptional subtype defined by ER, PR and AR status. In terms of copy number, Her2+ tumors showed biologic selectivity for oncogenes and putative driver genes located in 8 regions outside of the Her2 amplicon that were selectively aberrated in presence of Her2 amplification. We also identified a small population of breast tumors with substantial over-expression of Her2 and neighboring genes that lack amplification but show consistent hypo-methylation. In conclusion, considering the transcriptional profile of Her2+ breast tumors provides combination strategies and viable treatment options for patients who developed resistance to Her2-targeted treatments, with endocrine therapies for Her2+/ER+ tumors, and antiandrogen strategies for Her2+/AR+ tumors. Furthermore, co-ordinated regulation of Her2 and neighboring genes supports the notion that the effect of the Her2 amplicon may be more than through overexpression of Her2.

Citation Format: Anneleen Daemen, Jacob Rinaldi, Gerard Manning. Genomic redefinition of Her2+ breast shows role of estrogen and androgen signaling. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research; Oct 17-20, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(2_Suppl):Abstract nr B53.
Abstract B67: CD4 Th1 cytokines synergize with HER-2/HER-3 blockade to induce tumor senescence and apoptosis in both HER-2 sensitive and resistant breast cancer cells
Authors: Rosemblit Cinthia , Czerniecki Brian J.
Published: May 2016

Background: We have demonstrated there is a loss of anti-HER-2 Th1 response during tumorigenesis in HER-2pos breast cancer. Maintenance of this response is associated with complete response to neoadjuvant therapy and prolonged disease free survival. The CD4 Th1 or Th1 cytokines IFN-γ and TNF-α induce apoptosis and tumor senescence in HER-2pos breast cancer cells. In the current study we investigated whether combining Th1 cytokines with HER-2/HER-3 blockade would impact both HER-2 sensitive and resistant breast cancer cells.

Results: SK-BR-3 breast cancer cells exposed to HER2 Class II peptide-primed CD4+ T-cells cells in a transwell culture system confirmed that the cytokines produced by the CD4+ T cells induced senescence and apoptosis of the HER-2pos cells as measured by SA-β-galactosidase (gal) staining, increased expression of p15INK4b and cleaved caspase 3 detected by western blot. CD4+ T-cells primed either with immature dendritic cells (DCs) or mature DCs plus irrelevant Class II (BRAF or survivin) peptides were not able to induce senescence, nor apoptosis. The senescence and apoptosis observed was dramatically augmented in HER-2pos cells exposed to HER2 Class II peptide-primed CD4+ T-cells in presence of trastuzumab and pertuzumab in the transwell. HER-2 resistant cells HCC-1419 and JIMT-1 treated with IFN-γ and TNF-α demonstrated a dose dependent increase in senescence and apoptosis suggesting HER-2 resistant cell lines are sensitive to Th1 cytokine effects. Both breast cancer cell lines tested express IFN-γ and TNF-α receptors as measured by western blot analysis. Although trastuzumab and pertuzumab had little effect on induction of senescence or apoptosis in the resistant cells, the combination of Th1 cytokines with trastuzumab and pertuzumab induced similar levels of senescence measured by SA-β-gal assay and p15INK4b as well as effectively induced cell death by apoptosis to levels similar to those seen in HER-2 sensitive cell lines.

Conclusions: Our results establish a role for Th1 cytokines IFN-γ and TNF-α in inducing tumor senescence and apoptosis in both sensitive and resistant HER-2 expressing cells that is dramatically augmented by HER-2/HER-3 blockade. An effective CD4 Th1 response, combined with HER-2 and HER-3 blockade can significantly drive tumor senescence and apoptosis and should be readily explored as non-cross-reactive therapy to effectively eliminate residual cancer cells and prevent recurrence in HER-2pos breast cancer.

Citation Format: Cinthia Rosemblit, Brian J. Czerniecki. CD4 Th1 cytokines synergize with HER-2/HER-3 blockade to induce tumor senescence and apoptosis in both HER-2 sensitive and resistant breast cancer cells. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research; Oct 17-20, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(2_Suppl):Abstract nr B67.
Abstract IA15: Immune control of breast cancer metastasis
Authors: Welm Alana L.
Published: May 2016

Twenty to thirty percent of breast cancer patients eventually develop metastasis despite receiving state-of-the-art therapies. Clearly, identifying better ways to eliminate micro-metastatic tumor cells is paramount to eradicating death from breast cancer. We now know that every tumor is unique and that each tumor is heterogenous. Tumors hijack multiple, redundant pathways to survive and grow, and evolve resistance under selective pressure from therapy. Moreover, micro-metastatic cancer cells may be dormant and resistant to therapy. Thus, traditional approaches to treat cancer and prevent metastatic recurrences don't work for everyone.

Stimulating the immune system to kill cancer cells (known as immunotherapy) carries strong appeal: the response can be individualized; the immune system is effective against diverse pathogens (in this case, tumor antigens); and immunity can evolve and retain memory for long-term control of disease. The challenge is that by the time tumors are clinically detectable they are already ‘invisible’ to the immune system due to a number of factors: (1) natural selection for tumor cells that can escape immune control; (2) immune tolerance to self- and tumor-antigens; and (3) the strongly immunosuppressive environment in tumors, which renders effector cells inactive. A better understanding of how tumors achieve immune evasion will allow development of strategies to promote more effective anti-tumor immunity.

We have determined that Ron protein kinase activity in the host immune system is a major contributor to metastasis. This finding was unanticipated; the expected result was that blocking Ron signaling in tumor cells would inhibit tumor progression, which was the basis for development of Ron inhibitors by the pharmaceutical industry. Our work now shows that Ron inhibitors reduce metastasis through dual effects on tumor cells and on the immune system: Approximately 40% of human breast tumors upregulate the Ron ligand, which is secreted from the tumor, activates Ron on macrophages, and polarizes them to an immune suppressive state. Inhibiting Ron, either genetically or with drugs, blocks metastasis by activating an anti-tumor immune response mediated by CD8+ cytotoxic T lymphocytes (CTLs). Thus, inhibition of Ron boosts anti-tumor immunity and prevents metastasis. These data suggest that inhibition of Ron in the tumor microenvironment might provide a new, complementary strategy to ongoing immunotherapy. Thus, Ron inhibitors might serve as macrophage ‘checkpoint inhibitors’ to complement checkpoint inhibition in T-cells.

Citation Format: Alana L. Welm. Immune control of breast cancer metastasis. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research; Oct 17-20, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(2_Suppl):Abstract nr IA15.