bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract A81: Targeting of free fatty acid signaling in ovarian cancer may serve as a potential therapeutic approach.
Authors: Munkarah Adnan , Hamid Suhail , Chhina Jasdeep , Mert Ismail , Jackson LaToya , Hensley-Alford Sharon , Chitale Dhananjay , Giri Shailendra , Rattan Ramandeep
Published: May 2016

Recent studies have revealed an association between adipocyte driven free fatty acids (FFA) and the aggressiveness of epithelial ovarian cancer (EOC). Adipocyte derived free fatty acids (FFA) seem to promote epithelial ovarian cancer (EOC) growth and progression by acting as mitochondrial fuel source to support the energy requirements of EOC cells. Apart from acting as a fuel source, FFA may also enhance tumor growth through other signaling pathways that can promote proliferation and regulate metabolism. Recently, a family of FFA activated G-protein coupled receptors (FFAR/GPCRs) was identified, including: FFAR1/GPR40, FFAR2/GPR42, FFAR3/GPR41, FFAR4/GPR120 and GPR84. Using RT-PCR, we have found that exposing EOC cells to adipocytes results in overexpression of FFAR1/GPR40 in multiple ovarian cancer cell lines (ID8, A2780, C200, OVCAR3 and SKOV3). In mRNA extracted from formalin-fixed paraffin embedded tumor specimens, approximately 80% of high-grade serous ovarian carcinomas exhibited increased FFAR1 expression. Additionally, analysis of TCGA database revealed that patients with tumors exhibiting increased mRNA expression of FFAR1/GPR40 had poor overall survival and progression free survival. We have also found that targeting FFAR1 using its specific antagonist GW1100 inhibited the proliferation of A2780, ID8, C200, OVCAR3 and SKOV3 cell lines and resulted in downstream inhibition of insulin secretion and the Akt signaling pathway. On the other hand, treatment of the same cells with the FFAR1 agonist ,CAY10587, had no effect on proliferation. Thus, we conclude that FFAR1 may play an important role in the regulation of FFA mediated EOC cell proliferation. Targeting of FFAR1/GPR40 may be an attractive strategy in EOC, particularly for EOC patients presenting with high adiposity.

Citation Format: Adnan Munkarah, Suhail Hamid, Jasdeep Chhina, Ismail Mert, LaToya Jackson, Sharon Hensley-Alford, Dhananjay Chitale, Shailendra Giri, Ramandeep Rattan. Targeting of free fatty acid signaling in ovarian cancer may serve as a potential therapeutic approach. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: Exploiting Vulnerabilities; Oct 17-20, 2015; Orlando, FL. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(2 Suppl):Abstract nr A81.
Abstract B11: Mutational heterogeneity in non-serous ovarian cancers.
Authors: Teer Jaime , Yoder Sean , Gjyshi Anxhela , Zhang Chaomei , Monteiro Alvaro
Published: May 2016

Background: The mutational landscape of non-serous ovarian carcinomas is underexplored despite their substantial contribution to ovarian cancer morbidity and mortality. In this study we report whole exome sequencing analysis of 10 non-serous ovarian tumors and matched normal samples. We also report targeted gene sequencing results of an additional 28 non-serous tumor samples targeting 1,321 genes selected for their involvement in cancer.

Methods: Whole exome capture was performed using Roche NimbleGen SeqCap EZ, followed by sequencing of captured libraries using an Illumina HiScan platform. Target gene capture was performed using SureSelect custom designs, using GAIIx sequencing technology. Sequences from both experimental procedures were aligned to the human genome using the Burrows-Wheeler Aligner and the Genome Analysis ToolKit was used for insertion/deletion realignment, quality score calibration, and variant identification. Mutations were detected with MuTect or Strelka (whole exome) or GATK (target gene).

Results: We annotated 32,965 somatic mutations (829-15,737 per tumor) for an overall rate of mutation 12.2 per Mb. Interestingly, a BRCA2 frameshift mutation was observed in mucinous tumors; although inherited BRCA mutations have not been previously observed in mucinous histology. TP53 mutations were much less common than observed in high-grade serous ovarian cancers. PTEN stood out as a gene containing truncating mutations in many samples. We confirmed FOXL2 importance in granulosa, KRAS in mucinous histology and identified POLE mutations in endometrioid ovarian samples. Finally, we report comparative analysis of our results to The Cancer Genome Atlas (TCGA) database.

Conclusion: In summary, we report that non-serous ovarian tumors are mutationally distinct from serous ovarian tumors, having a mutational profile more similar to non-serous endometrial cancers.

Citation Format: Jaime Teer, Sean Yoder, Anxhela Gjyshi, Chaomei Zhang, Alvaro Monteiro. Mutational heterogeneity in non-serous ovarian cancers. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: Exploiting Vulnerabilities; Oct 17-20, 2015; Orlando, FL. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(2 Suppl):Abstract nr B11.
Abstract B35: Ovarian cancer incidence projections in the United States: Expected increases through 2030.
Authors: Stewart Sherri L. , Thompson Trevor D. , Weir Hannah K. , Rim Sun Hee
Published: May 2016

Background: Several reports have shown that age-standardized ovarian cancer rates are decreasing in recent years in the United States. While these reported rates are useful to compare trends across certain groups, they do not always accurately convey the burden of ovarian cancer, nor do they provide information on the actual numbers of ovarian cases diagnosed each year. Since age is the greatest risk factor for ovarian cancer, and the older population of the Unites States is growing, the accurate assessment of future ovarian cancer trends is important in order to plan for clinical care needs and allocate resources effectively.

Objective: To assess future trends in the United States ovarian cancer burden by projecting the number of new cases of ovarian and fallopian tube cancer cases diagnosed through the year 2030.

Methods: We used observed incidence data from 1982-2011 from the SEER program, and population projections based on the 2010 US Census to predict ovarian and fallopian tube cancer cases and rates through 2030. Cases of both ovarian and fallopian tube cancers were included to account for misclassification that often occurs between the two cancer sites and also because some ovarian cases are thought to originate in the fallopian tube. Predictions were made using age-period-cohort regression models implemented in the Nordpred software.

Results: The number of ovarian cancer cases is estimated to increase from 23,215 in 2012 to 27,666 in 2030, for a 19.2% total increase in new cases of ovarian cancer in 2030. Fallopian tube cancer cases are expected to increase from 1,621 in 2012 to 3,686 in 2030. Together, 31,352 cases of ovarian and fallopian tube cancers are projected to be diagnosed in 2030, for an overall expected increase of 26.2% for these cancers.

Discussion: Given the expected increase in cancer cases, prioritizing resources for ovarian cancer control is warranted. Continued training and outreach of gynecologic oncologists to ensure ovarian cancer patients receive optimal treatment is necessary. Systems changes that ensure referral and accessibility to gynecologic oncologists are also needed. Additional research into genetic and other factors that predispose individuals to ovarian cancer, and the continued characterization of symptoms may help to identify cases earlier, potentially resulting in more treatment options for ovarian cancer patients.

Citation Format: Sherri L. Stewart, Trevor D. Thompson, Hannah K. Weir, Sun Hee Rim. Ovarian cancer incidence projections in the United States: Expected increases through 2030. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: Exploiting Vulnerabilities; Oct 17-20, 2015; Orlando, FL. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(2 Suppl):Abstract nr B35.
Abstract B37: Menstrual cycle characteristics, PCOS, and ovarian cancer risk.
Authors: Harris Holly R. , Titus Linda J. , Cramer Daniel W. , Terry Kathryn L.
Published: May 2016

Background: Long and irregular menstrual cycles, a hallmark of polycystic ovary syndrome (PCOS), have been associated with higher androgen and lower sex hormone binding globulin levels (SHBG) and this altered hormonal environment may increase the risk of specific histologic subtypes of ovarian cancer.

Methods: We investigated the association between menstrual cycle characteristics and PCOS among 2071 women with epithelial ovarian cancer and 2100 controls in the New England Case-Control Study (1992-2008). Women self-reported menstrual cycle irregularity, menstrual cycle length, and previous diagnosis of PCOS. We used unconditional logistic regression models to calculate odds ratios (OR) and 95% confidence intervals (95% CIs) for ovarian cancer risk overall, and polytomous logistic regression to evaluate whether the associations differed across histologic subtypes.

Results: Overall, we observed no difference in ovarian cancer risk between women who reported periods that were never regular and women who did not report menstrual cycle irregularity (multivariable OR=0.87; 95% CI=0.69-1.10). A similar association was observed for those reporting an average menstrual cycle length of greater than 35 days with a multivariable OR of 0.87 (95% CI=0.47-1.60). In addition, no association was observed with self-reported PCOS and ovarian cancer (multivariable OR=0.96; 95% CI=0.60-1.54). When we examined the associations by histologic subtype, we observed significant differences in the association with menstrual cycle irregularity as the exposure (pheterogeneity=0.007) but not for cycle length >35 days or self-reported PCOS. Menstrual cycle irregularity was protective for serous invasive tumors (OR=0.66; 95% CI=0.47-0.91) but was associated with a non-significant increase in risk of serous borderline tumors (OR=1.32; 95% CI=0.86-2.03).

Conclusions: Our results suggest that menstrual cycle characteristics may have influences on ovarian cancer risk that vary by histologic subtype. Future research in a large collaborative consortium may help clarify these associations.

Citation Format: Holly R. Harris, Linda J. Titus, Daniel W. Cramer, Kathryn L. Terry. Menstrual cycle characteristics, PCOS, and ovarian cancer risk. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: Exploiting Vulnerabilities; Oct 17-20, 2015; Orlando, FL. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(2 Suppl):Abstract nr B37.
Abstract B45: Characterization of human fallopian tube epithelial stem-like cells.
Authors: Ding Dah-Ching , Chu Tang-Yuan
Published: May 2016

Purpose: Recent studies have suggested a large part of ovarian high grade serous carcinomas arise from the fallopian tube epithelium (FTE). The purpose of this study was to derive the stem-like cells of fallopian tube epithelium (FTESC).

Results: Immunohistochemistry of human fallopian tube revealed expression of Pax8 in secretory cells, Tubb4 in ciliated cells and Epcam in all the epithelial cells, whereas Lgr5 was expressed in a small subset of cells in the epithelial layer. The presumed FTESC were cultured from the fallopian tube fimbria epithelium exhibiting a cuboidal cell morphology and could be maintained at a constant proliferation rate up to P9. When cultured on matrigel, these cells formed spheroids with Pax8, Tubb4 and Lgr5 expression. Flowcytometry revealed decreasing expression of EpCAM, CD73, CD133 and CD105, and increasing of Pax 8 secretory cell marker (87% in P6, 78% in P3) with the advance of passage. Adding of estrogen to the culture resulted in an enlargement of cell body (4 nM) and differentiation toward ciliated cells (400 nM). When these FTESC were cultured on the mouse embryonic fibroblast feeder, or mixe-cultured with fallopian tube stromal cells and HUVECs, they formed tubal-like structures mimicking the fallopian tube.

Conclusion: We successfully derived a presumed FTESC with self-renewal and differentiation capability. These cells may provide new insight to investigate how FTE regenerate, respond to hormone and ROS injury and subjected to cancer initiation.

Citation Format: Dah-Ching Ding, Tang-Yuan Chu. Characterization of human fallopian tube epithelial stem-like cells. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: Exploiting Vulnerabilities; Oct 17-20, 2015; Orlando, FL. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(2 Suppl):Abstract nr B45.
Abstract IA01: The origins of ovarian cancer.
Authors: Cho Kathleen R.
Published: May 2016

Ovarian cancer is a generic term that can be used for any malignant ovarian tumor. The majority of ovarian cancers are carcinomas, i.e., malignancies with epithelial differentiation. Ovarian carcinomas represent a heterogeneous group of tumors that can be divided into five major types based largely on their light microscopic appearance. Notably, the origins of these different tumor types (high-grade serous carcinoma, low-grade serous carcinoma, endometrioid carcinoma, clear cell carcinoma, and mucinous carcinoma) remain incompletely understood. Indeed, many ovarian cancers may not actually originate in the ovaries, but instead involve the ovaries secondarily. Even those that develop in the ovaries may arise from cell types, for example endometrial- or fallopian tube-type epithelium, that are not considered intrinsic to normal ovaries.

Potential origins and putative precursors of the five major types of ovarian carcinoma will be reviewed and experimental data from mouse models supporting a role for the ovarian surface epithelium and/or fallopian tube epithelium will be presented. A more complete understanding of the origin(s) of each type of ovarian carcinoma is needed to inform the development of effective ovarian cancer prevention, early detection and treatment strategies.

Citation Format: Kathleen R. Cho. The origins of ovarian cancer. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: Exploiting Vulnerabilities; Oct 17-20, 2015; Orlando, FL. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(2 Suppl):Abstract nr IA01.
Abstract IA23: Ovarian cancer screening.
Authors: Menon Usha
Published: May 2016

Ovarian cancer is the 5th most common cause of cancer death among women in the UK, with ten-year survival rates of 35%. The continued high case fatality ratio has underpinned efforts to develop a screening strategy that could impact on mortality. In the UK large ovarian cancer screening trials have been completed both in the general (United Kingdom Collaborative Trial of Ovarian Cancer Screening - UKCTOCS) and high-risk (UK Familial Ovarian Cancer Screening Study UKFOCSS) population. Emerging evidence from UKCTOCS is encouraging with the multimodal strategy appearing to have detected double the number of invasive epithelial ovarian/fallopian tube cancer during incidence screening compared to a fixed cut-off for serum CA125 used previously. The key issue is impact on ovarian cancer mortality, which will be available from UKCTOCS in December 2015. Meanwhile new insights into carcinogenesis with a better understanding of the target lesion, improved design of biomarker discovery studies, a focus on detecting low volume disease using cancer specific markers, novel biospecimens such as endocervical samples, targeted imaging and time series algorithms for interpreting marker profile suggests that a new era in ovarian cancer screening is underway.

Citation Format: Usha Menon. Ovarian cancer screening. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: Exploiting Vulnerabilities; Oct 17-20, 2015; Orlando, FL. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(2 Suppl):Abstract nr IA23.
Abstract 20: Mutation profiling reveals distinct signatures in hypopharyngeal squamous cell carcinoma
Authors: Liu Zhiye , Wei Lai , Zhao Dewei , Wang Ruoyu , Tong Kai , Zhang Yuewei , Yang Bin , Cao Hong , Li Nina , Liang Shanshan , Gu Jiahuan , Wang Zhiqiang , Ding Yan
Published: May 2016

The mutational profiles of hypopharyngeal squamous cell carcinoma (HSCC), an anatomical and clinical subtype of head and neck squamous cell carcinoma (HNSCC) was characterized in this study. Whole exome sequencing of HSCC tumors (n=15) and normal pairs (n=15) reveals distinct somatic gene mutations to HSCC (KDR, ABL-1, ATM, and HNF1A), while some others are common to HNSCC significantly mutated genes (e.g. TP53, HRAS, NOTCH1 and PIK3CA). Copy number analysis shows distinct signatures of HSCC gene amplifications(e.g. CCND1) or deletions(e.g. GSTT1). Furthermore, pathway analysis identified several statistically significant enriched pathways (e.g. VEGFR signaling) for genomic alterations specific to HSCC. We then performed targeted therapies to individual patients corresponding to their mutation profiles, and monitored their responses to the personalized treatment by enumerating the circulating tumor cells. These findings offer new insights into understanding fundamental tumorigenic mechanisms of HSCC and exploring precise therapies.

Citation Format: Zhiye Liu, Lai Wei, Dewei Zhao, Ruoyu Wang, Kai Tong, Yuewei Zhang, Bin Yang, Hong Cao, Nina Li, Shanshan Liang, Jiahuan Gu, Zhiqiang Wang, Yan Ding. Mutation profiling reveals distinct signatures in hypopharyngeal squamous cell carcinoma. [abstract]. In: Proceedings of the AACR Precision Medicine Series: Integrating Clinical Genomics and Cancer Therapy; Jun 13-16, 2015; Salt Lake City, UT. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(1_Suppl):Abstract nr 20.
Abstract 36: Functional characterization of ERBB2 mutations and response to targeted therapies
Authors: Johnsen Hannah , Hanrahan Aphrothiti , Jones Alexis , Solit David
Published: May 2016

Amplification of HER2 (ERBB2) has been well characterized as an oncogenic driver, but only recently have mutations in ERBB2 also been shown to have activating and tumorigenic potential. We sought to characterize a large number of ERBB2 mutations seen in sequencing data from patients at MSKCC as well as the cBioPortal for Cancer Genomics, and determine the therapeutic response of these mutations to HER2 targeted therapies. HER2 is a transmembrane tyrosine kinase that is responsible for regulating cell growth and proliferation, among other cell processes. The ERBB2 gene is amplified in approximately 25% of breast cancers and in this disease, HER2 directed therapies are now a component of standard care. More recently, ERBB2 mutations were identified in a variety of cancers, most commonly cancers of the urinary tract but also at lower frequency in cancers of the breast, stomach, colon and lung, among others. Recent data has shown that select ERBB2 mutations activate HER kinase signaling and promote anchorage independent growth in vitro and tumor formation in mice. However, only a small number of ERBB2 mutations have been reported and biologically characterized as oncogenic in preclinical models. Moreover, the therapeutic impact of ERBB2 mutations has not been validated. To screen for activating potential of a large number of ERBB2 mutations, cells were transiently transfected to express ERBB2 mutants generated using site-directed mutagenesis. Expression of several mutations led to increased receptor activation and downstream signaling, as determined by levels of phosphorylated HER2, AKT, ERK, and S6 protein, whereas other mutations had little impact on activity compared to a wild type HER2 control. Stable cell lines were then generated to express the activating mutations and tested for transforming ability in soft agar. Mutations promoted colony formation, with certain mutations producing more colonies. Cells were also grown in the presence of the dual EGFR/HER2 inhibitor, neratinib. While certain mutations were more sensitive to lower doses of neratinib, addition of neratinib at higher doses abrogated colony formation in soft agar for all mutations. While a number of ERRB2 mutations have been shown to have tumorigenic potential, not all mutations respond similarly to HER2 targeted therapies. We hope to create an annotated database of alterations so that in the future, patients can be screened for specific mutations to minimize ineffective and unnecessary treatment options.

Citation Format: Hannah Johnsen, Aphrothiti Hanrahan, Alexis Jones, David Solit. Functional characterization of ERBB2 mutations and response to targeted therapies. [abstract]. In: Proceedings of the AACR Precision Medicine Series: Integrating Clinical Genomics and Cancer Therapy; Jun 13-16, 2015; Salt Lake City, UT. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(1_Suppl):Abstract nr 36.
Abstract 41: XI-006 induces potent p53-independent apoptosis in Ewing sarcoma
Authors: Pishas Kathleen , Neuhaus Susan , Clayer Mark , Farshid Gelareh , Staudacher Alexander , Callen David
Published: May 2016

Despite aggressive multi-modal treatment strategies, limited improvement in overall survival rates for patients diagnosed with Ewing sarcoma, the second most prevalent solid bone tumor observed in children and young adolescents, has been achieved over the past thirty years. As such, new targeted therapies are urgently required particularly for those sarcoma patients with recurrent or metastatic disease. Overexpression of MDM4, a major transcriptional regulator of the TP53 tumour suppressor has been documented in a substantial proportion of Ewing, Synovial and Osteosarcomas (>30%). For this reason, we have examined the molecular and cellular responses of cultured Ewing and Osteosarcoma cell lines following treatment with XI-006 (NSC207895), a small molecule 4-nitrobenzofuroxan derivative MDM4 transcriptional inhibitor.

XI-0006 treatment of Ewing and osteosarcoma cell lines (n=11) resulted in rapid and potent apoptosis at low nano-molar concentrations specifically in Ewing sarcoma cell lines (48hr IC50: 0.098-1.61µM). Importantly, the viability of normal human fibroblasts (IMR90) remained unaffected at these low nano-molar concentrations (48hr IC50: 6.80µM). Unexpectedly, apoptotic response was not dependent on MDMX mRNA and protein levels or TP53 status. Alkaline/neutral comet and γH2AX immunofluorescence assays revealed that the cytotoxic effects of XI-006 at low nano-molar concentrations could not be attributed to the induction of DNA damage. RNA expression analysis of XI-006 treated cell lines revealed that the mechanism of action of XI-006 could be attributed to the inhibition of cell division and cycle regulators such as KIF20A and GPSM2. XI-006 was also shown to synergise with current Ewing sarcoma chemotherapeutic protocols, Doxorubicin, Etoposide and Actinomycin D in vitro. Furthermore, combinatorial treatment with Olaparib led to a maximum 56.3% increase in apoptosis compared to XI-006 alone, due to the down-regulation of Mre11.

Our pre-clinical findings suggest that XI-006 represents a novel therapeutic intervention for the treatment of Ewing sarcoma, and warrants further pharmacokinetic and pharmacodynamic investigations of XI-006 in vivo.

Citation Format: Kathleen Pishas, Susan Neuhaus, Mark Clayer, Gelareh Farshid, Alexander Staudacher, David Callen. XI-006 induces potent p53-independent apoptosis in Ewing sarcoma. [abstract]. In: Proceedings of the AACR Precision Medicine Series: Integrating Clinical Genomics and Cancer Therapy; Jun 13-16, 2015; Salt Lake City, UT. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(1_Suppl):Abstract nr 41.
Abstract IA12: Open-source, crowdsourcing-capable tools for N of 1 cancer genomics
Authors: Mardis Elaine R.
Published: May 2016

There are multiple challenges to implementing cancer genomics in a clinical setting to identify for individual patients and their oncologist, the potentially druggable targets in their disease. Our group has introduced several open-source informatics tools for the identification of known driver mutations, for identifying their vulnerabilities regarding small molecule- and/or antibody-based inhibitors. These are meant not only to negotiate databased information that resides in different formats and locations, but also to serve as crowdsourcing-based information intake points that permit the community of practicing cancer care physicians to participate in knowledge sharing. My talk will highlight these tools and their functionality in the context of several “N of 1” examples from our work.

Citation Format: Elaine R. Mardis. Open-source, crowdsourcing-capable tools for N of 1 cancer genomics. [abstract]. In: Proceedings of the AACR Precision Medicine Series: Integrating Clinical Genomics and Cancer Therapy; Jun 13-16, 2015; Salt Lake City, UT. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(1_Suppl):Abstract nr IA12.
Abstract A063: Cancer cell expression of CCR2 regulates the PD-L1/ PD-1 immune checkpoint in breast cancer
Authors: Fein Miriam R. , Almeida Ana S. , Eberhardt Anaïs , Egeblad Mikala
Published: May 2016

C-C chemokine receptor type 2 (CCR2) is highly expressed in human breast cancer tissue by both cancer cells and infiltrating immune cells. High expression of CCR2 is correlated with advanced tumor stage, metastasis, and early relapse. Studies in mice have suggested that CCR2 expression on infiltrating inflammatory monocytes promotes primary tumor growth and metastasis. However, the role of CCR2 expression on cancer cells has not been well studied.

To determine how CCR2 directly impacts tumor growth and metastasis, we used a murine model of luminal breast cancer, the mouse mammary tumor virus (MMTV) promoter polyoma middle T antigen (PyMT) model. We crossed MMTV-PyMT mice to Ccr2-/- mice and found that loss of CCR2 did not alter tumor onset. However, primary tumors of MMTV-PyMT;Ccr2+/+ mice grew significantly faster than tumors of MMTV-PyMT;Ccr2-/- mice. Lung metastases were also larger in MMTV-PyMT;Ccr2+/+ mice than in MMTV-PyMT;Ccr2-/- mice.

We determined by RNA in situ hybridization that CCR2 was indeed expressed by both host cells and cancer cells in MMTV-PyMT tumors. To differentiate between the contributions of CCR2 expression in the tumor and host, we transplanted primary cancer cells derived from MMTV-PyMT;Ccr2+/+ and MMTV-PyMT;Ccr2-/- mice to syngeneic Ccr2+/+ and Ccr2-/- hosts. Surprisingly, CCR2 expression in the host had no significant effect on tumor growth. However, deletion of CCR2 from cancer cells resulted in delayed tumor growth and increased survival. Furthermore, when Ccr2+/+ and Ccr2-/- cancer cells were transplanted to athymic nude mice (lacking T cells), the resulting tumors no longer showed a delayed growth. Consistent with this finding, flow cytometry of infiltrating leukocytes in tumors transplanted to syngeneic mice revealed increased levels of CD8+ cytotoxic T cells and antigen-presenting CD103+ dendritic cells into Ccr2-/- tumors compared to wild type tumors. These data suggest that CCR2 expression on cancer cells regulates an immune response to the tumors. Supporting this conclusion, expression of programmed cell death ligand 1 (PD-L1) on Ccr2-/- cancer cells was reduced to just 30% of the level on wild type cancer cells, indicating that CCR2 expression on cancer cells suppresses the anti-tumor immune response. Together, our results establish a novel role for cancer cell CCR2 expression in breast cancer progression by suppression of cytotoxic T cells through an immune checkpoint.

Citation Format: Miriam R. Fein, Ana S. Almeida, Anaïs Eberhardt, Mikala Egeblad. Cancer cell expression of CCR2 regulates the PD-L1/ PD-1 immune checkpoint in breast cancer. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr A063.
Abstract A163: Tertiary lymphoid structures: Prognostic significance and relationship with tumor-infiltrating lymphocytes in triple-negative breast cancer
Authors: Park In Ah , Lee Hee Jin , Song In Hye , Kim Joo Young , Yu Jong Han , Gong Gyungyub
Published: May 2016

Tumor-infiltrating lymphocytes (TILs) have a strong prognostic and predictive significance, particularly in triple-negative breast cancer (TNBC). One important source of TILs in breast cancer is tertiary lymphoid structures (TLSs). Here, we performed histologic analysis of surgically resected TNBC to identify the location of TLSs, the relationship between TLSs and TILs, and their prognostic significance in TNBC. We retrospectively analyzed 769 patients with TNBC. TILs were defined as the percentage of stroma of invasive carcinoma infiltrated by lymphocytes. TLSs were mainly present within adjacent terminal duct lobular units and around in situ components. TNBC with higher levels of TILs showed a higher nuclear grade, lower lymphovascular invasion rate, less accompanying in situ component, a homogeneous growth pattern, necrosis in invasive areas, low levels of tumor stroma, high levels of peritumoral lymphocytic infiltration, and moderate to abundant TLSs in adjacent tissue. TILs, degree of peritumoral lymphocytic infiltration, and adjacent TLSs were prognostic factors for disease-free and overall survival outcomes. Although the TIL level did not have a prognostic value in stage I, it added significant prognostic information for stages II and III. Conversely, patients with high levels of TILs did not show prognostic differences according to the pTNM stage. Patients with high levels of TILs (>60%) and moderate to abundant TLSs had significantly better disease-free survival outcomes than those with high levels of TILs but none or few TLSs. TLSs are frequently present in TNBC and are closely associated with TILs. TILs provide additional prognostic information in TNBC patients with a higher pTNM stage.

Citation Format: In Ah Park, Hee Jin Lee, In Hye Song, Joo Young Kim, Jong Han Yu, Gyungyub Gong. Tertiary lymphoid structures: Prognostic significance and relationship with tumor-infiltrating lymphocytes in triple-negative breast cancer. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr A163.
Abstract A172: Allogeneic dendritic cells (AlloDCs) transduced with an infection-enhanced adenovirus as adjuvant for cancer immunotherapy
Authors: Fotaki Grammatiki , Jin Chuan , Karlsson-Parra Alex , Yu Di , Essand Magnus
Published: May 2016

AlloDC cancer immunotherapy utilizes the allogeneic reaction as an immunostimulatory adjuvant. Intratumoral alloDC vaccination reverses the immune suppressed tumor micro-environment and activates anti-tumor immune responses. Herein, we aim to examine the use of Ad5f35PTD, an infection-enhanced serotype 5 adenovirus containing fibers from serotype 35 and cell-penetrating peptides on the hexons, as a vector to deliver tumor-associated antigens to monocyte-derived immature DCs in combination with an established DC maturation cocktail. Ad5f35PTD-transduced, cocktail-matured DCs expressed co-stimulatory and activation molecules and secreted the Th-1 type cytokine IL-12, in vitro. Natural killer (NK) cells migrated better towards medium from Ad5f35PTD-transduced, cocktail-matured DCs than medium from only cocktail-matured DCs, which was in accordance with a higher CXCL-10 chemokine level. The alloDC effect was examined in vivo using mature murine DCs of BALB/c origin (D2SC/I), transduced with Ad5f35PTD encoding the melanoma-associated antigen gp100, injected subcutaneously into C57BL/6 mice. They induced a higher migration of host DCs in the draining lymph node, in comparison to the D2SC/I cells alone. The migrated host DCs efficiently presented the captured gp100 and activated gp100-specific CD8+ T-cells. Moreover, alloDCs transduced with the gp100-encoding Ad5f35PTD reduced the immunosuppressive environment when injected intratumoraly by reducing the ratio of monocytic to granulocytic myeloid-derived suppressor cell compartment and delay tumor growth in a B16 melanoma model, in comparison with the D2SC/I cells alone. Concluding, the Ad5f35PTD transduced alloDCs expressing gp100 showed potency in eliciting tumor-specific immune responses in comparison to the alloDCs alone.

Citation Format: Grammatiki Fotaki, Chuan Jin, Alex Karlsson-Parra, Di Yu, Magnus Essand. Allogeneic dendritic cells (AlloDCs) transduced with an infection-enhanced adenovirus as adjuvant for cancer immunotherapy. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr A172.
Abstract B017: A soluble form of CD80 inhibits PD-L1 immune suppression and stimulates T cells through CD28-specific pathways indicating potential for increased therapeutic activity over checkpoint inhibition alone
Authors: Long Tiha M. , Ostrand-Rosenberg Suzanne
Published: May 2016

Antibodies that block the PD-1/PD-L1 pathway have demonstrated therapeutic success against several cancer types, but do not induce a response in the majority of patients. The development of treatments that provide activation signals to T cells by targeting co-stimulatory receptors, in addition to checkpoint inhibition, may increase anti-tumor immunity and the percent of responding patients. CD80/B7-1 is a co-stimulatory molecule that also interacts with PD-L1. CD80 is expressed by antigen presenting cells and ligates to CD28 providing co-stimulation during T cell activation. Furthermore, PD-L1 interacts with T cell CD80 in a reverse signaling pathway that promotes T cell anergy. The ability of CD80 to block inhibitory and promote stimulatory signaling pathways in T cell activation suggests potential therapeutic value. We have shown that soluble CD80 fused to the Fc region of IgG (CD80-Fc) blocks PD-1/PD-L1 interactions thus restoring activation of CD4+ and CD8+ T cells. Moreover, CD80-Fc was more effective in vitro than PD-1/PD-L1 monoclonal antibodies at restoring T cell activation, which suggests that CD80-Fc may be stimulating CD28 in addition to blocking PD-1/PD-L1 signaling. Stimulation of T cell CD28 in the absence of other signals induces downstream transcription, and signaling pathway activation. In fact, agonist CD28 antibody can induce functions in T cells in both naïve and activated cells, even in the absence of TCR activation. We now demonstrate that CD80-Fc does indeed stimulate T cells via CD28 activation by demonstrating comparable downstream results of CD80-Fc and agonist CD28 antibody treatment of both human and mouse T cells. Therefore, CD80-Fc may provide a novel therapeutic approach to target T cell checkpoint inhibition in addition to providing stimulatory signaling to improve T cell activation and anti-tumor immunity.

Citation Format: Tiha M. Long, Suzanne Ostrand-Rosenberg. A soluble form of CD80 inhibits PD-L1 immune suppression and stimulates T cells through CD28-specific pathways indicating potential for increased therapeutic activity over checkpoint inhibition alone. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr B017.
Abstract B030: Structure-function studies of cytosolic DNA sensing pathway
Authors: Gao Pu , Ascano Manuel , Zillinger Thomas , Barchet Winfried , Tuschl Thomas , Patel Dinshaw J.
Published: May 2016

The appearance of DNA in the cytoplasm is a very dangerous signal to cells. This could be caused by viral or bacterial infection, or as a result of cellular damage that leaks host DNA into the cytoplasm. Cytosolic DNA activates the host immune response, which is critically dependent on the newly identified cGAS-cGAMP-STING pathway. Structure-function studies from our laboratories have demonstrated that upon dsDNA binding, cGAS is activated through conformational transitions, resulting in formation of a catalytically competent and accessible nucleotide-binding pocket for generation of c[G(2',5')pA(3',5')p]. Further, cyclization occurs in a stepwise manner through initial generation of 5'-pppG(2',5')A prior to cyclization to c[G(2',5')pA(3',5')p], with the latter positioned precisely in the catalytic pocket. These studies established c[G(2',5')pA(3',5')p] as a founding member of a family of metazoan 2',5'-containing cyclic heteronucleotide second messengers distinct from bacterial 3',5' cyclic dinucleotides. Follow up structure-function studies from our laboratories have shown that human and mouse STING undergo an ‘open’ to ‘closed’ conformational transition upon binding c[G(2',5')pA(3',5')p] and the anti-viral agent DMXAA. Comparing hSTING to mSTING in cellular assays, 2',5'-linkage-containing cGAMP isomers were more specific triggers of the IFN pathway compared to the all 3',5'-linkage isomer counterparts. Our results highlight the critical role of the lid residue at position 230 (Gly in hSTING; Ile in mSTING) for the DMXAA species selectivity. Our structural and functional results also shed light on strategies to restore an efficient DMXAA-response of hSTING based on the binding pocket S162A and Q266I substitutions.

Citation Format: Pu Gao, Manuel Ascano, Thomas Zillinger, Winfried Barchet, Thomas Tuschl, Dinshaw J. Patel. Structure-function studies of cytosolic DNA sensing pathway. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr B030.
Abstract B043: Suppression of T cell responses by androgen deprivation drug leads to early relapse
Authors: Pu Yang
Published: May 2016

Physical or chemical androgen deprivation therapy (ADT) is the cornerstone treatment for metastatic prostate cancer, but relapse eventually occurs and the results of combination of ADT with immunotherapy are disappointing. Here, we show that physical castration can synergize with subsequent immunotherapy while the more commonly used chemical ADT suppresses the immunological protection induced by immunotherapy. Furthermore, chemical ADT can broadly inhibit immune responses. Mechanistically, the inhibition of immune responses by chemical ADT targets T cells by interfering with initial T cell priming, independently of the androgen receptor. We have shown that this off-target immune suppression is mediated by a receptor shared by γ-aminobutyric acid. Further investigation of chemical ADT administered with immunotherapy has revealed that timing and dosing critically control anti-tumor effects. Therefore, this study highlights an underappreciated mechanism of ADT immune suppression and provides a new strategy to enhance immunity and prevent the relapse of advanced prostate cancer.

Citation Format: Yang Pu. Suppression of T cell responses by androgen deprivation drug leads to early relapse. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr B043.
Abstract B073: Structural basis of the 12-23 rule in V(D)J recombination
Authors: Ru Heng , Chambers Melissa G. , Liao Maofu , Wu Hao
Published: May 2016

The V, D and J segment recombination (V(D)J recombination) is the key event in the development and maturation of T and B lymphocytes, which generates a diverse repertoire of T and B cell clones for the further selection of useful specificities. V(D)J recombination is also the basis for antibody diversity. The initiation step of V(D)J recombination is carried out by a pair of lymphocyte specific enzymes, which are called recombination-activating gene 1 and 2 (RAG1 and RAG2). RAG1 and RAG2 associate with each other and with two recognition signal sequences (RSS, 12RSS and 23RSS) adjacent to the coding ends of the V, D and J segments to catalyze the double stranded breaks required for joining. The reaction goes through several catalytic steps that result in formation of the single recombination complex (SC), the paired complex (PC), the cleaved signal complex (CSC) as well as the signal end complex (SEC).

In order to understand the mechanisms of how RAG complex functions on DNA, we solved the structures PC and CSC by cryo-electron microscopy near the atomic resolution. Compared with the Apo-RAG complex that constitutes an open conformation, the PC and CSC reveal a closed conformation with RAG1 in one protomer interacts with RAG2 in another protomer. Furthermore, the first nucleotide of the heptamer in the non-transfer strand is flipped out to avoid the steric hindrance of the 3'-OH to attack the phosphodiester bond in the transfer strand. Upon nucleophilic attack and hairpin formation, the nucleotide adjacent to the 5'-position of the attacked phosphate is also flipped out to accommodate the tensile force in the hairpin DNA. In addition, we found that the conformation changes in the nonamer-binding domain (NBD), which is distorted toward to the 12RSS, and the distorsion requires two turns for the nonamer DNA in the 23RSS to reach the NBD. Finally, we found that two HMGB1 molecules, rather than one, are required to bind and bend both 12RSS and 23RSS DNA to adjust the conformational change of NBD on each side. In summary, our structural models not only uncover the mechanisms of how RAG protein catalyzes the hairpin formation, but also provide the structural basis of the 12-23 rule in V(D)J recombination.

Citation Format: Heng Ru, Melissa G. Chambers, Maofu Liao, Hao Wu. Structural basis of the 12-23 rule in V(D)J recombination. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr B073.
Abstract B104: Characterization and specificity analysis of tumor infiltrating lymphocytes in ovarian carcinoma
Authors: Peper Janet Kerstin , Röhle Kevin , Schuster Heiko , Wagner Philipp , Rammensee Hans-Georg , Stevanovic Stefan
Published: May 2016

Introduction: The great majority of patients suffering from ovarian cancer (OvCa) relapse after initial therapy, making OvCa the most lethal gynecological malignancy. The immunogenic nature of OvCa is highlighted by frequent infiltration with immune cells, which represent an independent prognostic factor in OvCa patients and argues in favor for the development of immunotherapies, including peptide-based cancer vaccines. The phenotype of OvCa infiltrating T cells as well as their specificity and expression of inhibitory receptors is so far unknown. This information is however critically important for the design of novel immunotherapies including the application of checkpoint inhibitor therapy.

Materials and Methods: Tumor infiltrating lymphocytes (TILs) were isolated from fresh tumor tissue of OvCa patients obtained during tumor debulking surgery. The phenotype and expression of inhibitory molecules LAG3, CTLA4, PD1 and TIM3 was assessed by flow cytometry. Peptide specificity of infiltrating T cells was analyzed by screening for reactivity against naturally presented HLA ligands employing IFNγ ELISPOT and flow cytometry. These natural presented HLA ligands have been originally identified by mass spectrometry in an extensive HLA ligandome analysis of OvCa samples.

Results: TILs from each analyzed OvCa sample demonstrated expression of at least one of the investigated inhibitory receptors. While LAG3 and PD1 were most prominent during this analysis, CTLA4 expression was only weak or absent. Furthermore, high amounts of effector memory, but few naive or central memory T cells were present. Notably, OvCa infiltrating CD4+ T cells and CD8+ T cells shared a similar expression pattern of inhibitory surface markers within each single patient. We further obtained first results in peptide-specificity of OvCa-derived T cells by screening them for reactivity against naturally presented HLA ligands identified on OvCa samples.

Conclusion: In summary, we provide first insight into the expression of various inhibitory receptors on OvCa infiltrating T cells and their peptide specificities.

Citation Format: Janet Kerstin Peper, Kevin Röhle, Heiko Schuster, Philipp Wagner, Hans-Georg Rammensee, Stefan Stevanovic. Characterization and specificity analysis of tumor infiltrating lymphocytes in ovarian carcinoma. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr B104.
Abstract B119: Deciphering CD20 immunogenicity to generate protective anti-tumor CD4+ and CD8+ T cells
Authors: Siberil Sophie , Milcent Benoit , Riller Quentin , Deligne Claire , Hamieh Mohamad , Toutirais Olivier , Latouche Jean-Baptiste , Teillaud Jean-Luc
Published: May 2016

We have previously shown that anti-tumor T cell response is triggered by anti-CD20-mAb in mice bearing EL4-huCD20 tumor cells, a murine tumor expressing human CD20+ (huCD20) molecule. To define huCD20-derived epitopes that might be targeted by mAb-induced adaptive responses, we have selected different candidate peptides restricted to Kb, Db, and I-Ab, based on in silico prediction and in vitro binding to recombinant MHC molecules. Naive C57Bl/6 mice were then immunized with different mixtures of these peptides and subsequently challenged with EL4-huCD20 tumor cells. A longer survival could be achieved in mice immunized with MHCI and MHCII huCD20-derived peptide mixtures corresponding to various regions of the human CD20 molecule. Then, we defined in the same regions twenty-three huCD20-derived peptides with high in vitro binding to human MHCI and MHCII molecules frequent in Caucasian populations. The use of humanized HLA-DR1/HLA-A2.1 transgenic mice injected with EL4-huCD20 tumor cells and of NIH/3T3 cells expressing accessory and human HLA-DR1 or A2.1 molecules loaded with the candidate peptides made it possible to define 4 MHCI-restricted and 13 MHCII-restricted CD20-derived peptides capable of stimulating IFN-gamma production by specific CD4+ and CD8+ T cells. These findings support further studies on the identification and characterization of CD20-specific T cells in anti-CD20-treated follicular lymphoma patients. The definition of CD20-derived immunogenic epitopes also paves the way for the search of new biomarkers correlating with the clinical outcome of patients.

Citation Format: Sophie Siberil, Benoit Milcent, Quentin Riller, Claire Deligne, Mohamad Hamieh, Olivier Toutirais, Jean-Baptiste Latouche, Jean-Luc Teillaud. Deciphering CD20 immunogenicity to generate protective anti-tumor CD4+ and CD8+ T cells. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr B119.