bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract 234A: Aberrant sodium current contributes to constitutive mTOR activity in malignant melanoma cells
Authors: Gallant Benjamin , Lizza Nicole , Garrity Ryan , Madigan Audrey , Jost Zachary , Gonzalez Alfredo , Justiniano Jeanine , Xie An , Wan Yinsheng 
Published: July 2016

Abstract
Transformation from disciplined melanocytes to untamed melanoma cells remains an enigma. Our previous studies have demonstrated that melanoma cells are more resistant to oxidative stress and melanoma cells exhibit constitutive mTOR activity. Surprisingly, further studies have failed to present the expression and activity of EGFR using conventional anti-EGFR antibodies. We hypothesized that constitutive mTOR activity in melanoma cells may be due to mutated EGFR variants and membrane channel activities. Using patch clamping technique, we have shown that melanoma cells (WM 266-4) but not human skin melanocytes exhibit Na+ current which is blocked by TTX. Interestingly, mTOR inhibitor, rapamycin, blocks Na+ current. Western blot and confocal microscopy data further revealed that Na+/Ca2+ exchanger blocker KB-R7943 (KBR), and L-type Ca2+ channel blocker Nifedipine (NIF) inhibits mTOR activity in melanoma cells in a dose dependent manner. To further characterize Na+ channels, we used commercially available channel antibodies. Western blot analysis data showed that melanoma cells but not human melanocytes express Na+ v1.5 and Na+ v1.6 and NCX3. Functional studies also indicated that KBR and NIF inhibit melanoma cell proliferation and migration. Taken all together, our data suggest that aberrant Na+ current contributes to constitutive mTOR activity in melanoma cells and channel blockers may be potential for the treatment of melanoma.
Citation Format: Benjamin Gallant, Nicole Lizza, Ryan Garrity, Audrey Madigan, Zachary Jost, Alfredo Gonzalez, Jeanine Justiniano, An Xie, Yinsheng Wan. Aberrant sodium current contributes to constitutive mTOR activity in malignant melanoma cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 234A.
Abstract 2356: Erythrocytes used as tumor antigen delivery system to target antigen-presenting cells embody an innovative approach for in situ cancer immunotherapy
Authors: Cremel Magali , Guerin Nathalie , Barthe Quitterie , Bourgeaux Vanessa , Berlier Willy , Horand Françoise , Godfrin Yann 
Published: July 2016

Abstract
Introduction: In current vaccine therapy, efficient delivery of tumor antigens (TA) to antigen-presenting cells (APC) represents a major step toward the development of strong TA-specific immune response including cytotoxic T lymphocyte (CTL) induction. Our innovative approach is to use the property of erythrocytes to be naturally phagocytosed by APC when senescent. Indeed, TA can be encapsulated into erythrocytes that will be used as carrier to specifically deliver the TA to APC, which will ensure their degradation and presentation to T cells.
Materials and Methods: The proof of concept was established with Ovalbumin (OVA), tyrosinase-related protein 2 (TRP2 (melanoma TA) and prostate specific antigen (PSA, prostate TA). These TA were encapsulated into erythrocytes by a hypotonic lysis process. To stimulate the erythrophagocytosis by APC, the membranes of erythrocytes were coated with antibodies. Then, erythrocytes containing TA were intravenously injected to mice concomitantly with Poly(I:C) adjuvant. The CTL response was analyzed by IFNγ ELISPOT and in vivo lysis of TA-expressing target cells injected to immunized mice. Humoral response was assessed by the determination of TA-specific immunoglobulin titer in mouse serum.
Results: All TA were efficiently encapsulated in erythrocytes in a dose-dependent manner. A strong T-cell response was induced after 2 injections of TA encapsulated in erythrocytes: 2μg of OVA and 20μg of TRP2 induced the in vivo lysis of 97% and 96% of TA-target cells, respectively. By comparison, injections of free antigen induced less than 5% of lysis. Furthermore, a significant number of TA-specific IFNγ-secreting cells was generated (1585 and 286 cells / 106 cells for TRP2 and PSA, respectively) compared to the free TA (&lt; 80 cells / 106 cells for both TA). In addition, we established a dose-dependent PSA-specific humoral response. Finally, a significant delay of tumor growth was obtained in mice more than twenty days after the subcutaneous implantation of EG7-OVA or B16F10 (melanoma) tumor cell lines, compared to injections with free antigen or control vehicle.
Conclusion: The use of erythrocytes as TA carrier to specifically deliver antigen to APC and induce efficient immune response against tumor can be a very promising strategy in cancer immunotherapy.
Citation Format: Magali Cremel, Nathalie Guerin, Quitterie Barthe, Vanessa Bourgeaux, Willy Berlier, Françoise Horand, Yann Godfrin. Erythrocytes used as tumor antigen delivery system to target antigen-presenting cells embody an innovative approach for in situ cancer immunotherapy. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2356.
Abstract 2364: Spontaneous and vaccine-induced clearance of Mus Musculus Papillomavirus1 (MusPV1) infection
Authors: Jiang Rosie T. , Wang Joshua W. , Peng Shiwen , Hung Chien-Fu , Roden. Richard B.S 
Published: July 2016

Abstract
Human papilloma virus (HPV) high risk (hr) types cause several cancers, including cervical, vaginal, penile, and oropharyngeal, whereas low risk types cause benign diseases such as skin warts, anogenital warts and recurrent respiratory papillomatosis. Although hrHPV infection is one of the most common sexually transmitted diseases most infections remain asymptomatic and are cleared while a sub-population develops persistent HPV-related pathologies including cancer. Further, immune-compromised populations, notably organ transplant and HIV-positive patients, are particularly vulnerable to HPV-related pathologies. Here, we describe the first establishment of mouse papillomavirus-1 (MusPV1) disease in outbred and immune competent SKH1-Elite mice (Crl:SKH1-Hrhr). Challenge of SKH1 mice with MusPV1 resulted in three clinical outcomes: 1) persistent (&gt;2 months) papillomas (∼15%), 2) transient papillomas that spontaneously regress typically within 2 months (∼10%), 3) no visible papillomas and viral clearance (∼75%). SKH1 mice with persisitent papillomas were treated using a potentially preventive and therapeutic DNA vaccine composed of human calreticulin (hCRT) linked to MusPV1 early proteins, mE6 and mE7 and the late protein mL2 (hCRTmE6/mE7/mL2). Three intramuscular DNA vaccinations (15 μg in PBS) via in vivo electroporation were administered weekly and immune responses were measured, including mE6/mE7/mL2 serum antibody response and T-cell epitope mapping studies. Previously persistent papillomas disappeared within 2 months after final vaccination. Clearance of MusPV1 was confirmed by loss of mE6/mE7 transcript staining using RNA-CISH on formalin-fixed, paraffin-embedded tail sections. Vaccination induced a strong mE6 and mE7 CD8+ T cell response in all mice. Interestingly, mL2 antibody responses were faster and stronger in asymptomatic challenged mice and in those that spontaneously cleared their papillomas after challenge as compared to mice with persistent papillomas. In sum, MusPV1 challenge of outbred, immune-competent SKH1 mice represents a promising model to further study potential immunotherapies for HPV-related disease and the relationship between spontaneous clearance of papillomavirus and host genetics.
Citation Format: Rosie T. Jiang, Joshua W. Wang, Shiwen Peng, Chien-Fu Hung, Richard B.S Roden. Spontaneous and vaccine-induced clearance of Mus Musculus Papillomavirus1 (MusPV1) infection. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2364.
Abstract 237: Enrichment of AXL-high/ MITF-low melanoma cells in the presence of MAPK inhibitors in vitro
Authors: Dutton-Regester Ken , Garraway Levi 
Published: July 2016

Abstract
BRAF and MEK inhibitors have become a standard of care for patients with metastatic BRAF mutant (V600) melanoma. Despite their success, 10- 20% of patients exhibit ‘intrinsic’ resistance and fail to respond to treatment. Recently, distinct transcriptional profiles have been associated with sensitivity to these drugs with intrinsically resistant melanomas exhibiting an AXL-high/ MITF-low phenotype (1). Using melanoma cell lines from the Cancer Cell Line Encyclopedia with matched gene expression data, we confirmed that AXL-high/ MITF-low cell lines had increased resistance to both BRAF (Dabrafenib) and MEK (Trametinib) inhibitors, either singly or in combination. Using flow cytometry with an AXL antibody, we observed that AXL-high/ MITF-low resistant cell lines frequently exhibited a high percentage of AXL+ cells (≥90%) while AXL-low/ MITF-high sensitive cell lines showed the opposite (≤5%). We hypothesized that the small percentage of AXL+ cells in the sensitive cell lines may be responsible for mechanisms of adaptive or acquired resistance (a feature that is frequently observed with the use of BRAF and MEK inhibitors in vitro and in the clinic). To explore this, we cultured sensitive cell lines in vitro for 5 days in the presence of combined Dabrafenib and Trametinib (DT) and performed flow cytometry to determine if there was a change in the percentage of AXL+ cells. All sensitive cell lines exhibited an enrichment of AXL+ cells with increasing concentrations of DT (maximum increase compared to DMSO control across individual cell lines ranged from 10% to 85%). It is unknown whether this phenomenon could be explained by a clonal selection or ‘plastic’ epigenetic reprogramming process and as such, we are currently performing experiments to determine this. Looking to the future, identifying potential dependencies of AXL-low/ MITF-high melanomas will deepen our understanding of the biology supporting this resistant state and provide a platform for the design of future clinical intervention strategies in this subset of patients.
1) Konieczkowski et al. 2014. Cancer Discovery. A melanoma cell state distinction influences sensitivity to MAPK pathway inhibitors. 4(7):816-27.
Citation Format: Ken Dutton-Regester, Levi Garraway. Enrichment of AXL-high/ MITF-low melanoma cells in the presence of MAPK inhibitors in vitro. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 237.
Abstract 2375: Multiclonal invasion in breast cancer identified by single cell DNA sequencing
Authors: Casasent Anna K. , Long Annalyssa N. , Schalck Aislyn , Sei Emi , Gao Ruli , Davis Alexander , Wang Yong , Edgerton Mary E. , Navin Nicholas E. 
Published: July 2016

Abstract
Ductal carcinoma in situ (DCIS) is the most common form of early stage breast cancer and is frequently detected by mammography. Only 10% of low-grade and 30% of high-grade DCIS patients will later present with an invasive carcinoma, making it difficult to determine which patients to treat aggressively. Although DCIS is often considered a precursor to invasive breast carcinoma, a major question in the field is whether invasive subpopulations evolve directly from the in situ populations or independently. We hypothesize that early tumor cells evolve over an extended period of time in the ducts, generating multiple subpopulations that migrate out of the ducts together. To test this hypothesis, we developed an approach that combines laser-capture microdissection (LCM) with single cell sequencing to perform DNA copy number profiling while preserving the topographic location of cells. We applied this method to 10 high-grade breast cancer patients with defined in situ, invasive, and normal regions. Using these data, we delineated the clonal substructure and location of the in situ and invasive subpopulations. We found that most high-grade tumors are composed of 1-4 major clonal subpopulations that are intermixed in both the in situ and invasive regions. These subpopulations share a common evolutionary lineage or cell of origin. We also performed deep-exome sequencing (100X) of matched microdissected regions of in situ, invasive and adjacent normal tissue from these breast cancer patients. In each tumor we identified a large number of shared somatic mutations between the in situ and invasive regions, further supporting our direct genomic lineage hypothesis. We also identified a few mutations confined to the invasive tumor regions that may play an important role in the invasive phenotype. These data support a multiclonal invasion model in which distinct clones evolve in the ducts and subsequently migrate into the adjacent tissues to establish invasive tumors. This model has important implications for the diagnosis and therapeutic treatment of DCIS breast cancer patients since multiple clones may need to be targeted to inhibit invasion.
Citation Format: Anna K. Casasent, Annalyssa N. Long, Aislyn Schalck, Emi Sei, Ruli Gao, Alexander Davis, Yong Wang, Mary E. Edgerton, Nicholas E. Navin. Multiclonal invasion in breast cancer identified by single cell DNA sequencing. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2375.
Abstract 2386: Microfluidic single cell exome-seq and RNA-seq analysis of tumor composition
Authors: Ragoussis Ioannis , Savage Paul , Wang Yu-Chang , Revil Timothee , Badescu Dunarel , Saleh Sadiq , Iacucci Ernesto , Bertos Nicolas , Monast Anie , Omeroglou Attila , Zuo Dongmei , Park Morag 
Published: July 2016

Abstract
Human breast tumors have been shown to exhibit extensive inter- and intra-tumor heterogeneity. While recent advances in genomic technologies have allowed us to deconvolute this heterogeneity, few studies have addressed the functional consequences of diversity within tumor populations. Here, we identified an index case for which we have derived a patient-derived xenograft (PDX) as a renewable tissue source to identify subpopulations and perform functional assays. On pathology, the tumor was an invasive ductal carcinoma which was hormone receptor-negative, HER2-positive (IHC 2+, FISH average HER2/CEP17 2.4), though the FISH signal was noted to be heterogeneous. On gene expression profiling of bulk samples, the primary tumor and PDX were classified as basal-like. We performed single cell RNA and exome sequencing of the PDX to identify population structure. Using a single sample predictor of breast cancer subtype, we have identified single basal-like, HER2-enriched and normal-like cells co-existing within the PDX tumor, a finding replicated in several independent experiments. Genes differentially expressed between these subpopulations are involved in proliferation and differentiation. One population was characterized by high MYC and the other high EGFR/KRT14. These findings were validated using immunohistochemistry on PDX and primary tumor material. Further functional experiments showed that that EGFR subpopulation has stem cell character istics and forms metastasis in the animal model. Microfluidic whole genome amplification followed by whole exome capture of 81
single cells, along with exome sequencing of the germline, primary and post treatment tumor and whole PDX showed that BRCA1 and TP53 are mutated in all single cells, as well as a number of sub-clonal mutations that are being investigated further. Loss of heterozygocity was observed in 16 TCGA cancer driver genes and novel mutations in 7 known cancer driver genes. Careful comparison of the exome sequencing data allowed the association of driver gene mutation prevalence with tumor progression. These findings are important in our understanding the functional consequences of intra-tumor heterogeneity with respect to clinically important phenotypes such as invasion, metastasis and drug-resistance.
Citation Format: Ioannis Ragoussis, Paul Savage, Yu-Chang Wang, Timothee Revil, Dunarel Badescu, Sadiq Saleh, Ernesto Iacucci, Nicolas Bertos, Anie Monast, Attila Omeroglou, Dongmei Zuo, Morag Park. Microfluidic single cell exome-seq and RNA-seq analysis of tumor composition. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2386.
Abstract 2396: Heterogeneity of colorectal cancers and multiple liver metastases by mutation and copy number profiling
Authors: Alagaratnam Sharmini , Høst Brunsell Tuva , Aske Danielsen Stine , Sveen Anita , Eknæs Mette , Hektoen Merete , Røsok Bård , Bjørnbeth Bjørn Atle , Nesbakken Arild , Lothe Ragnhild 
Published: July 2016

Abstract
To investigate genetic and genomic heterogeneity in liver metastases of colorectal cancer, multiple tumor foci in the liver per patient were identified on the basis of radiological images and samples from these systematically biobanked. For a total of 67 patients, 258 metastatic tumor samples were collected (minimum of 2 and maximum of 9 samples per patient, median of 4 samples per patient). Additionally, for n = 14 patients, tumor samples from the corresponding primary colorectal cancer were also available. DNA isolated from these samples was subjected to mutation profiling by Sanger sequencing for the genes BRAF, KRAS, PIK3CA and TP53, as well as allele-specific copy number profiling by CytoscanHD SNP array. For the four genes investigated for mutations, 39/67 (58%) of the patients were homogeneous in their mutation profile, while 21/67 (32%) were heterogeneous (one or more samples had mutations not detected in all samples). For 7/67 patients (10%), no mutations were detected in any of these four genes. For 9/14 patients with matching primary tumors, there was complete concordance between the primary tumor and all liver metastases profiled for mutations, while the remaining 5/14 patients showed discordance in mutation profile between the primary tumor and at least one of the liver metastases profiled. High resolution copy number profiles of this cohort detected many of the aberrations previously described in primary colorectal cancer, specifically gain of chromosomes 7, 8q, 13q and 20q, and loss of chromosomes 5q, 8p, 15q, 17p and 18q. The extent of heterogeneity observed in patient-wise mutation profiles was largely mirrored in the copy number profiles of the same tumors. 16/27 (60%) of the patients for which both mutation and copy number profiles were available showed concordance in the degree of heterogeneity per patient. In summary, intra-individual heterogeneity varied from patient to patient, with results from the genetic and genomic data types largely concurring.
Citation Format: Sharmini Alagaratnam, Tuva Høst Brunsell, Stine Aske Danielsen, Anita Sveen, Mette Eknæs, Merete Hektoen, Bård Røsok, Bjørn Atle Bjørnbeth, Arild Nesbakken, Ragnhild Lothe. Heterogeneity of colorectal cancers and multiple liver metastases by mutation and copy number profiling. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2396.
Abstract 24: Loss of ubiquitin-specific peptidase 18 (USP18) causes cold sensitive mice by destabilizing the critical regulator of thermogenesis: uncoupling protein-1 (UCP-1)
Authors: Liu Xi , Lu Yun , Zheng Lin , Sekula David J. , Freemantle Sarah J. , Dmitrovsky Ethan 
Published: July 2016

Abstract
USP18 is the major ISG15 (Interferon-Stimulated Gene 15) deconjugase that removes ISG15 from substrate proteins. We recently reported that USP18 null mice spontaneously develop leiomyosarcomas. Intriguingly, these USP18 null mice are also markedly cold sensitive as compared to their wild-type littermates. When briefly exposed to cold stimuli, USP18 null mice significantly (P &lt; 0.05) decreased their body temperature. In contrast, wild-type mice exposed to the same experimental conditions exhibited transient changes in their body temperature. We sought to elucidate the engaged mechanism. Expression profiles of critical thermogenic regulatory proteins were examined by immunoblot analyses of brown fat tissues of USP18 null mice. Strikingly, UCP-1 expression was substantially reduced in these tissues of USP18 null versus wild-type mice. To independently confirm that reduced UCP-1 expression was caused by loss of USP18, stable knock-down of USP18 was independently achieved in a panel of murine cell lines by use of transfected small hairpin RNAs (shRNAs). The obtained results were compared to that of transfected control vectors. USP18 down-regulation by different shRNAs markedly reduced UCP-1 protein as compared to controls. As expected, engineered gain of USP18 expression in the same murine cell lines stabilized UCP-1 protein. We explored if UCP-1 destabilization was due to its complex formation with the ubiquitin-like protein ISG15. This was the expected experimental outcome since USP18 is the distinct deconjugase that removes ISG15 from complexed proteins. Immunoprecipitation assays were performed to establish this direct association. These studies revealed a complex formed between ISG15 and UCP-1 protein. Thus, engineered loss of USP18 in mice led to cold sensitive mice from repression of UCP-1. As a consequence of its complex with ISG15, the thermogenic regulator UCP-1 was destabilized. Taken together, this study extends prior work by showing a previously unrecognized link between USP18 and regulation of thermogenesis. This defines a novel role for the USP18 protease in metabolism.
Citation Format: Xi Liu, Yun Lu, Lin Zheng, David J. Sekula, Sarah J. Freemantle, Ethan Dmitrovsky. Loss of ubiquitin-specific peptidase 18 (USP18) causes cold sensitive mice by destabilizing the critical regulator of thermogenesis: uncoupling protein-1 (UCP-1). [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 24.