bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3616: Hypoxia determines the fate of myeloid cell differentiation by controlling STAT3 activation in tumor site
Authors: Kumar Vinit
Published: May 2015

The abnormal differentiation of myeloid cells is the key element of altered immune response in cancer. Myeloid derived suppressor cells (MDSC) are a major component of immune suppressive network. MDSC accumulate in large numbers in peripheral lymphoid organs and inside tumors. Recent data indicated that the fate of MDSC depends on the localization of these cells. In contrast to peripheral lymphoid organs, inside tumors, MDSC rapidly differentiate to tumor associated macrophages with potent immune suppressive activity. Hypoxia is intricate part of tumor microenvironment. We have found that hypoxia plays a major role in regulation of MDSC differentiation to macrophages in tumor site. Hypoxia caused dramatic reduction in phosphorylated STAT3 in MDSC. These results were unexpected, since high level of STAT3 activity is a hallmark of MDSC present in blood and lymphoid organs. In our experiments spleen MDSC in tumor-bearing mice had much higher level of STAT3 phosphorylation than their counterparts in tumor site from the same mice. In cancer patients MDSC in tumor site also had substantially lower level of pSTAT3 than MDSC in peripheral blood. In vitro culture of spleen MDSC in hypoxia led to the suppression of STAT3 activation. Furthermore, mice with constitutively active STAT3 had shown the dramatic expansion of MDSC and down-regulation of macrophage population in tumor microenvironment, suggesting that STAT3 is a downstream target for hypoxia driven myeloid cells differentiation in cancer. We found that CD45 protein tyrosine phosphatase was a molecule that regulated STAT3 activation in hypoxia. The activity but not expression of CD45 phosphatase was up-regulated in tumor MDSC as compared to MDSC in spleens. The exposure of spleen MDSC to hypoxia led to the significant increase in CD45 activity. The siRNA mediated silencing of CD45 abrogated the effect of hypoxia on the down-regulation of STAT3 activity in MDSC. This suggests that hypoxia driven up-regulation of CD45 phosphatase activity suppresses STAT3 activation, which in turn plays a key role in differentiation of myeloid cells in tumor site. These unexpected findings may provide rationale for re-evaluation of therapeutic strategies targeting myeloid cells in cancer.

Citation Format: Vinit Kumar. Hypoxia determines the fate of myeloid cell differentiation by controlling STAT3 activation in tumor site. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3616. doi:10.1158/1538-7445.AM2014-3616
Abstract 3618: Ikkα modulates primary sclerosing cholangitis and intrahepatic cholangiocarcinoma
Authors: Jiang Qun , Xiao Zuoxiang , Back Timothy , Scarzello Anthony , Roan Scott , Willette-Brown Jami , Zhu Feng , Hu Yinling , Wiltrout Robert H.
Published: May 2015

Intrahepatic cholangiocarcinoma (ICC) is the second most common primary liver cancer after hepatocellular carcinoma (HCC). The risk of ICC is higher in patients with primary sclerosing cholangitis (PSC). To date, the cellular and molecular mechanism underlying the pathological progression of PSC and ICC is poorly understood. IKKα is part of the IκB kinase (IKK) complex, which plays an important role in regulating inflammation- associated carcinogenesis through both NF-κB-dependent and independent pathways. Here, we show that IKKα mutant mice developed very serious PSC as early as four weeks of age. The ALT/AST and bilirubin levels were significantly increased in the serum of IKKα mutant mice along with lymphocytic and eosinophilic infiltration into the liver. Liver inflammation in the IKKα mutant mice, mediated by macrophages, neutrophils and CD4+ T cells, was associated with the death of cholangiocytes and hepatocytes, and obstruction of intrahepatic and extrahepatic bile ducts, which impeded bile flow and ultimately led to biliary fibrosis and cirrhosis. Additionally, upon activation of NOTCH signaling in the liver via hydrodynamic shear, we observed that NOTCH-induced ICC, with the PSC, developed significantly faster in IKKα mutant mice. To identify whether intrinsic IKKα dysfunction in hepatocytes promotes the NOTCH-induced ICC in IKKα mutant mice, we generated IKKα hep KO mice, in which IKKα is conditionally knocked out in hepatocytes. No biliary disease or liver injury was observed in these mice. We then established an accelerated ICC model utilizing hydrodynamic delivery of NICD and AKT expression vectors. Unexpectedly, ICC development was remarkably slower in the IKKα hep KO mice compared to the IKKα floxed control mice, and this delayed ICC development was associated with reduced activation or levels of AKT, NOTCH, MAPK/ERK and c-Myc. These data suggest that IKKα may play a protective role in PSC, while promote ICC derived from hepatocytes. In contrast to ICC, we also established the cMET/β-catenin-induced HCC model in the IKKα hep KO mice, which did not reveal any difference in tumor development between IKKα hep KO and control mice. Taken together, our findings suggest that IKKα plays complicated and important roles in PSC and ICC pathological progression.

Citation Format: Qun Jiang, Zuoxiang Xiao, Timothy Back, Anthony Scarzello, Scott Roan, Jami Willette-Brown, Feng Zhu, Yinling Hu, Robert H. Wiltrout. Ikkα modulates primary sclerosing cholangitis and intrahepatic cholangiocarcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3618. doi:10.1158/1538-7445.AM2014-3618
Abstract 3620: Neuropeptide signaling activates Type-1 immunity through the NK1 and NK2 receptors on human dendritic cells
Authors: Kitamura Hidemitsu , Ohtake Junya , Kaneumi Shun , Masuko Kazutaka , Sumida Kentaro , Terada Satoshi , Kishikawa Takuto , Ono Yosuke , Kita Toshiyuki , Kobayashi Hiroya
Published: May 2015

Dendritic cells (DCs), representative antigen-presenting cells, effectively induce antigen-specific immune responses through activation of CD4+T and CD8+T cells. In tumor microenvironments, DCs uptake tumor-derived antigens, and generate helper T cells and cytotoxic T lymphocytes (CTLs), which recognize and kill the target tumor cells in response to the antigens. Thus, proper regulation of DC function is important for tumor immunology.

Substance P (SP) and Neurokinin A (NKA), neurotransmitters, are widely distributed in both central and peripheral nervous system. Recently, we demonstrated that IFN-γ remarkably induced neurokinin-2 receptor (NK2R) expression human monocyte-derived dendritic cells. These findings strongly suggested that neuropeptide-signaling cascade might be closely related with regulation of DC-mediated immune responses. To elucidate the precise role of such neuro-immune crosstalk, we further investigated the effect of neuropeptide signaling on function of human DCs.

At first, we found that both neurokinin-1 receptor (NK2R) and NK2R mRNA expressions were significantly enhanced by IFN-β, IFN-γ, LPS, or poly I:C stimulation with human DCs generated from peripheral blood mononuclear cells (PBMCs). In addition, we confirmed that the upregulation of NK1R and NK2R mRNA expressions was induced in a STAT-1-dependent manner. Surface expression levels of HLA-DR and costimulatory molecules on DCs, augmented by poly I:C, were modulated in the presence of specific inhibitors against NK1R or NK2R. On the other hand, we confirmed that blockade of NK1R- and NK2R-mediated signaling cascade significantly suppressed cytokine productions by antigen-specific CD4+ T cells after the antigen stimulation with DCs. Finally, we found that human DCs also enhanced expression level of TAC-1 gene, which encodes SP and NKA, after the IFN-γ stimulation.

Thus, these findings indicate that NK1R- and NK2R-dependent neuropeptide signaling regulate Type-1 immunity through the activation of DC function, suggesting that such neuro-immune cross-talk through SP-NK1R and NKA-NK2R cascade might be involved in various diseases caused by excessive Type-1-dominant immunity including cancer and infection with chronic inflammation.

Citation Format: Hidemitsu Kitamura, Junya Ohtake, Shun Kaneumi, Kazutaka Masuko, Kentaro Sumida, Satoshi Terada, Takuto Kishikawa, Yosuke Ono, Toshiyuki Kita, Hiroya Kobayashi. Neuropeptide signaling activates Type-1 immunity through the NK1 and NK2 receptors on human dendritic cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3620. doi:10.1158/1538-7445.AM2014-3620
Abstract 3622: Total loss of HLA class I expression by two sarcomatoid hepatocellular carcinoma cell lines sHCC29 and sHCC63 was caused by a ∼49-kbp deletion at chromosome 15q15 across the β2m gene locus
Authors: Lei Wei-Yi , Hsieh Chin-Hsuan , Chang Chien-Chung , Wen Shao-Hsuan , Hu Chi-Tan , Liao Shuen-Kuei
Published: May 2015

Scanty information is available concerning the abnormalities in HLA class I antigen presentation in sarcomatoid hepatocellular carcinoma (sHCC), although this information can improve our understanding of sHCC-host interactions and assist in a rational design of T cell immunotherapy. In this study, we have established three sarcomatoid HCC cell lines, namely sHCC29, sHCC63, and sHCC74, from the tumor lesions in three patients; two of them were HBV carriers from which sHCC29 and sHCC63 were derived, while the other was a HCV carrier from which sHCC74 was generated. No viral DNA was detected in the culture supernatants or cell extracts from any of the three HCC cell lines by COBAS AmpliPre/TaqMan qPCR. In monolayer cell culture, both sHCC29 and sHCC74 showed similar elongated fibroblastic morphology, while sHCC63 exhibited a short-spindle shape and expanded in a density-sensitive manner. HLA class I molecules were abundantly expressed on sHCC74, but were not detected on sHCC29 and sHCC63 by cytofluorometric analysis. Cytoplasmic β2-microglobulin (β2m) was also undetected in the latter two cell lines, in spite of their baseline level of low-moderate cytoplasmic HLA class I heavy chain expression, which was 33-50% of that expressed by sHCC74 in terms of % positive cells. The observed HLA class I-loss phenotype was IFN-γ (300 μg/ml, 48 h)-uncorrectable. Inactivation of the β2m genes in both sHCC29 and sHCC63 was supported by the failure to detect their β2m mRNA transcripts and by the identification of a large (∼49,187 bp) deletion across the β2m gene locus at chromosome 15q15. This deletion, the largest reported to date, explains the loss of β2m protein and lack of assembly of HLA class I molecules in the sHCC29 and sHCC63 cell lines. Mapping of the deletion breaking point and validation by immunohistochemistry of β2m loss in the corresponding tumor lesions from which sHCC29 and sHCC63 were derived are currently underway.

Citation Format: Wei-Yi Lei, Chin-Hsuan Hsieh, Chien-Chung Chang, Shao-Hsuan Wen, Chi-Tan Hu, Shuen-Kuei Liao. Total loss of HLA class I expression by two sarcomatoid hepatocellular carcinoma cell lines sHCC29 and sHCC63 was caused by a ∼49-kbp deletion at chromosome 15q15 across the β2m gene locus. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3622. doi:10.1158/1538-7445.AM2014-3622
Abstract 3634: Postoperative neutrophil-lymphocyte ratio as a surrogate marker for recurrence of gastric cancer
Authors: Tanaka Hiroaki , Muguruma Kazuya , Sakurai Katsunobu , Toyokawa Takahiro , Kubo Naoshi , Ohira Masaichi , Hirakawa Kosei
Published: May 2015

Tumor markers such as carcinoembryonic antigen (CEA) and CA 19-9 are both widely used in the follow up of patients with gastric cancer. However, there are no definitive predictive markers for postopetrative recurrence. It has been reported that preoperative neutrophil lymphocyte ratio (NLR) was associated with prognosis of patients with gastrointestinal cancer. The aim of this study was to examine the impact of postoperative NLR on prediction for postoperative recurrence of gastric cancer. We retrospective reviewed data from 133 patients underwent curative surgical resection for gastric cancer between 2006 and 2010. Patients treated with preoperative chemotherapy or multiple cancers were excluded. We found 52 recurrent diseases which mainly included 29 cases with peritoneal dissemination, 13 with distant lymph node metastasis and 7 with liver metastasis. In univariate analysis, histological diffuse type, mascroscopical type 4 cancer, regional lymph node metastasis, lymphatic invasion in primary tumor, tumor diameter, elevated CEA or CA19-9, and elevated NLR. By ultivariate analysis, we found that elevated CA19-9 and NLR were independent predictive markers. Increased NLR was associated with diffuse type, peritoneal ddissemination, elevated CEA and preoperative NLR but not the presence or absence of adjuvant chemotherapy. In conclusion, postoperative NLR might be one of the surrogate markers for recurrence after curative surgery for gastric cancer.

Citation Format: Hiroaki Tanaka, Kazuya Muguruma, Katsunobu Sakurai, Takahiro Toyokawa, Naoshi Kubo, Masaichi Ohira, Kosei Hirakawa. Postoperative neutrophil-lymphocyte ratio as a surrogate marker for recurrence of gastric cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3634. doi:10.1158/1538-7445.AM2014-3634
Abstract 3638: Vaccine-draining lymph nodes of cancer patients for generating anticancer antibodies
Authors: Shukla Girja S. , Pero Stephanie C. , Sun Yu-jing , Carman Chelsea L. , Olson Walter C. , Slingluff Craig L. , Krag David N.
Published: May 2015

Human vaccine studies regularly demonstrate vaccine-induced antibodies in the blood. Characterization of these antibodies shows that they exhibit an extensive range of bioactive mechanisms. Unfortunately, there is not yet a reliable method by which to produce these important antibodies. Human antibodies may not require any molecular modification for therapeutic application and have a much faster pipeline to clinical studies than animal-derived antibodies. Lymph nodes are the primary destination of tumor or vaccine antigens. In the node, B cells undergo clonal expansion and somatic hypermutation, leading to the affinity-matured populations of effector B cells secreting antibodies that bind to the tumor or vaccine. The cancer vaccine-draining node is the ideal source of B cells that produce anticancer antibodies. Despite multiple human cancer vaccine studies, very little research has been done to recover the B cells responsible for vaccine-induced anticancer antibodies. In the present study, we used mononuclear cells from surgically removed vaccine-draining lymph nodes of melanoma patients vaccinated with 6 melanoma peptides derived from cancer-testis antigens and from melanocytic differentiation proteins for generating anti-vaccine peptide antibodies. The lymph node draining the vaccination site was localized by lymphatic mapping with a radiotracer. The development and maturity of B cells were assessed by determining phenotypic characters of lymph node cells using multicolor flow cytometry. The results showed that these vaccine-draining nodes contain high numbers of class-switched (CD19+CD27+IgD-IgM-) B cells and plasmablasts (CD19+CD38+IgM-). B-cell ELISpot assay was used to quantify the proportion of B cells in vaccine-draining lymph nodes that secrete anti-melanoma peptide antibodies. Positive ELISpot responses were observed in patients who also showed serum antibody-reactivity towards the vaccine peptides. The identification of lymph node cell samples exhibiting strong B-cell responses allows efficient generation and screening of hybridomas that secrete antibodies against cancer vaccine antigens. This study establishes a step-wise protocol for generating anti-cancer antibodies from vaccine-draining lymph nodes. We anticipate that the ability to reliably generate in vitro the same antibodies observed in the blood of vaccinated patients will further stimulate research to understand mechanisms of human antibody activity.

Citation Format: Girja S. Shukla, Stephanie C. Pero, Yu-jing Sun, Chelsea L. Carman, Walter C. Olson, Craig L. Slingluff, David N. Krag. Vaccine-draining lymph nodes of cancer patients for generating anticancer antibodies. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3638. doi:10.1158/1538-7445.AM2014-3638
Abstract 364: Mechanical properties of growing melanocytic nevi and the progression to melanoma
Authors: Taloni Alessandro , Alemi Alexander A. , Ciusani Emilio , Sethna James , Zaperi Stefano , Laporta Caterina Anna Maria
Published: May 2015

Melanocytic nevi are benign proliferations that sometimes turn into malignant melanoma in a way that is still unclear from the biochemical and genetic point of view. Diagnostic and prognostic tools are then mostly based on dermoscopic examination and morphological analysis of histological tissues. To investigate the role of mechanics and geometry in the morpholgical dynamics of melanocytic nevi, we study a computation model for cell proliferation in a layered non-linear elastic tissue. Numerical simulations show that the morphology of the nevus is correlated to the initial location of the proliferating cell starting the growth process and to the mechanical properties of the tissue. Our results also show that melanocytes are subject to compressive stresses that fluctuate widely in the nevus and depend on the growth stage. This result is important because mechanical stress has an effect that differs for primary and metastatic melanoma. We show experimentally that osmotic stress and collagen inhibit growth in primary melanoma cells while the effect is much weaker in metastatic cells. Knowing that morphological features of nevi might also reflect geometry and mechanics rather than malignancy could be relevant for diagnostic purposes.

Citation Format: Alessandro Taloni, Alexander A. Alemi, Emilio Ciusani, James Sethna, Stefano Zaperi, Caterina Anna Maria Laporta. Mechanical properties of growing melanocytic nevi and the progression to melanoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 364. doi:10.1158/1538-7445.AM2014-364
Abstract 3644: C-Jun N-terminal kinases and autophagy promote adenoviral and ectopic tumor-associated antigen presentation
Authors: Klein Sarah Renee
Published: May 2015

Adenoviruses are innately immunogenic; thus, targeted immunotherapy with recombinant, oncolytic adenoviruses is under investigation for the treatment of cancer. Evidence indicates adenoviruses induce autophagy that is required for cancer cell lysis. Importantly, autophagy is known to promote the presentation of self and foreign antigens for immune cell activation. The molecular regulation of autophagy and antigen presentation in adenovirus- infected cells remains undefined. Our data indicates adenovirus infection triggered phosphorylation of c-Jun N-terminal kinases (JNK) in vitro and in vivo, which was essential for autophagy. JNK-dependent B cell lymphoma-2 (Bcl-2) phosphorylation stimulated the dissociation of Bcl-2/beclin 1 heterodimers, enabling beclin 1 to initiate autophagy. Additionally, our data demonstrated for the first time that adenoviral epitopes were presented at the surface of infected cells through JNK- and autophagy-dependent mechanisms, as infected JNK- and autophagy-deficient cells were unable to activate splenocytes from adenovirus-immunized mice. Rapamycin, a chemical activator of autophagy, was able to enhance adenoviral antigen presentation. Contrary to starvation-induced autophagy, inhibiting adenovirus-mediated autophagy and antigen presentation required genetic knock-out of both JNK1 and JNK2 isoforms. We propose this unique mechanism for the processing of adenoviral antigens and consequent activation of immune cells can be leveraged in developing recombinant adenoviruses for the advancement of anti-tumor immunotherapy. Thus, we developed a novel adenovirus that expresses a chimeric fiber protein containing the cancer-specific, immunogenic epitope from the epidermal growth factor receptor variant III (EGFRvIII). We confirmed that the EGFRvIII epitope was presented at the surface of cells in an autophagy-dependent manner that could be enhanced by the pro-inflammatory cytokine, interferon gamma. Thus, we discovered molecular events regulating adenovirus-mediated autophagy and antigen presentation that differs from the conventional paradigm which could be employed for the advancement of adenovirus-based vaccines.

Citation Format: Sarah Renee Klein. C-Jun N-terminal kinases and autophagy promote adenoviral and ectopic tumor-associated antigen presentation. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3644. doi:10.1158/1538-7445.AM2014-3644
Abstract 3649: Significance of HIV viral load and CD4 count on Kaposi sarcoma in the era of highly active antiretroviral therapy
Authors: King Gentry T. , Garg Shivani
Published: May 2015

Background: Approximately 20% of patients with HIV will have Kaposi Sarcoma (KS) and about half will die within 3 months of diagnosis. Incidence and outcomes of KS have traditionally been linked to CD4 counts <350. More recently, KS has been increasingly described in higher CD4 counts and variable presentations in the era of Highly-Active-Antiretroviral-Therapy (HAART). No studies have established the role of Viral Load (VL) on KS.

Objective: To determine the effect of CD4 count and HIV VL on KS presentation and outcomes.

Methods: Retrospective review of KS patients with HIV admitted over 12 years (2000-2012). KS patients without HIV infection were excluded. Patients were divided into 4 groups based on CD4 count (≥200 or 100,000 or <100,000). Groups were analyzed with regards to KS severity (good vs poor risk), extent (soft tissue vs visceral), receipt of HAART and survival (6months). Statistical analysis was done via Chi-Square test.

Results: 43 patients were included. KS affected predominantly young (30-40yrs), male (93%) HIV patients with VL >100,000 (85%) and CD4 counts 100,000 had a statistically significant increased incidence for visceral involvement (96%, p100,000 ((66.7% with good risk), increased survival (59.4% survived >6months) and less use of HAART (20%). Although, these differences were statistically insignificant.

Conclusion: HIV VL of >100,000 is correlated with increased severity and extent of KS. Though statistically insignificant, CD4 counts <200 showed a trend for less severe disease and increased survival despite less use of HAART. Larger prospective studies are warranted to elucidate on these differences.

Citation Format: Gentry T. King, Shivani Garg. Significance of HIV viral load and CD4 count on Kaposi sarcoma in the era of highly active antiretroviral therapy. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3649. doi:10.1158/1538-7445.AM2014-3649
Abstract 3653: Nanoparticle-delivered T7-synthesized siRNA enhances cell killing in HER-2 (+) breast cancer
Authors: Alonso-Valenteen Felix , Medina-Kauwe Lali K.
Published: May 2015

We have previously developed the recombinant fusion protein, HerPBK10, for ligand-directed targeting and penetration of HER2+ breast cancer. HerPBK10 contains a ten lysine (K10) carboxy [C] -terminal tail that allows it to non-covalently bind anionic payloads, such as nucleic acids, by electrophilic interaction. In this study, a modified siRNA capable of both gene silencing and immunostimulation is the payload. Here, we test the hypothesis that ligand-directed, protein-based nanoparticles delivering siRNA launch a three-pronged attack on tumors by combining missile-like targeting (known as “transductional targeting”) with siRNA-mediated gene silencing, and cytokine-mediated cell death. The latter effect will be accomplished by taking advantage of the normally undesirable cytokine-mediated cytotoxicity induced by epigenetic modifications on certain forms of siRNA, and delivering this to tumors. Here we will deliver siRNA produced by T7 phage polymerase transcription, which introduces a 5′ triphosphate cap that triggers cytokine-mediated toxicity within host cells. So far, in vitro data has shown that this approach is more effective in reducing cell proliferation than synthetic siRNA against the same gene. Furthermore, in vivo xenograft data shows that this strategy can control and ablate the growth of HER2+ tumors. Future directions include IHC investigation of resected tumors and testing this hypothesis in an immunocompetent in vivo setting where we envision this immune-sensitive strategy having even greater efficacy.

Citation Format: Felix Alonso-Valenteen, Medina-Kauwe. Nanoparticle-delivered T7-synthesized siRNA enhances cell killing in HER-2 (+) breast cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3653. doi:10.1158/1538-7445.AM2014-3653
Abstract 3654: The role of structure-specific endonuclease MUS81 in the induction of cytosolic DNA in tumor cells
Authors: Ho Samantha , Gasser Stephan
Published: May 2015

Genomic DNA is constantly damaged by endogenous and exogenous agents. The DNA damage response (DDR) is vital for recognizing DNA damage and instigating DNA repair reactions to ensure genome stability. MUS81 is a structure-specific endonuclease that is required for resolving stalled replication forks at sites of DNA damage to initiate Homologous Recombination repair. Our preliminary data suggests that the potentially detrimental DNA structures cleaved by MUS81 accumulate in the cytosol of many tumor cells. Furthermore, we found that DNA cleaved by MUS81 induces type I interferon expression in a STING-dependent manner, and initiates an innate immune response in vivo. Cytosolic DNA in tumor cells was also abrogated upon inhibition of DNA damage sensors, ATM/ATR and PARP1. In addition, genetic inhibition of Mus81 in different tumor cells lines impaired expression of cytosolic DNA and type I interferon. In summary, our data suggest that the structure-specific endonuclease MUS81 cleaves damaged nuclear DNA leading to accumulation of genomic DNA in the cytosol of tumor cells, activating immunosurveillance mechanisms.

Citation Format: Samantha Ho, Stephan Gasser. The role of structure-specific endonuclease MUS81 in the induction of cytosolic DNA in tumor cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3654. doi:10.1158/1538-7445.AM2014-3654
Abstract 3657: Cross-regulation between the two subsets of NKT cells is dependent on interferon-gamma
Authors: Izhak Liat , Kato Shingo , Parish Stanley T. , Terabe Masaki , Berzofsky Jay A.
Published: May 2015

Overcoming the suppression of tumor immunity has been a major challenge in the fight against cancer. Suppression of tumor immunity is being regulated by different kinds of suppressor cells and depleting or blocking their function has been the target of immunotherapy development. One subset of immune suppressors is type II NKT cells. We have previously shown that, upon simultaneous stimulation, type I NKT cells that can promote tumor immunity and type II NKT cells cross regulate each other in vitro as well as in vivo. In some tumor models it was found that activation of type II NKT cells by sulfatide can completely abrogate the protective affect of type I NKT cell activation by alpha-galactosylceramide, leading to tumor progression and reduced survival. However the mechanism for this suppression has not been understood. By co-stimulating NKT cells in vitro, we found that type II NKT cells can inhibit the proliferation of type I NKT cells. By blocking a variety of immunregulators, we found that this suppression is dependent on interferon-gamma (IFNg). Blocking IFNg or its receptor reversed the suppression of type I NKT cell proliferation in vitro. In addition suppression was not observed in IFNgKO C57BL/6 NKT cells. It was also found that exogenous IFNg reduces the proliferation of type I NKT cells after stimulation, suggesting that IFNg is necessary for the suppression of type I NKT cells by type II NKT cells. This was unexpected as IFNg is also an effector cytokine secreted by type I NKT cells. The mechanism is under investigation. Blocking the ability of type II NKT cells to suppress type I NKT cells could shift the balance toward immune protection leading to a better tumor outcome.

Citation Format: Liat Izhak, Shingo Kato, Stanley T. Parish, Masaki Terabe, Jay A. Berzofsky. Cross-regulation between the two subsets of NKT cells is dependent on interferon-gamma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3657. doi:10.1158/1538-7445.AM2014-3657
Abstract 366: Translational modeling of docetaxel-thalidomide combination treatment in metastatic, castrate-resistant prostate cancer: predicting clinical response using preclinical data
Authors: Fernandez Eric , Mistry Hitesh , Brightman Frances , Orrell David , Dahut William L. , Figg William D. , Stein Wilfried D. , Chassagnole Christophe
Published: May 2015

A major cause of drug failure in the clinic is that preclinical studies do not predict with sufficient certainty what will happen in clinical practice. Accurately translating information from animal studies to the clinic would have a major impact on attrition rate. We have developed a mathematical model of a tumor cell population called the Virtual Tumour, which has been used extensively to predict the efficacy of single drug or drug combination treatment in preclinical studies. We have now extended and adapted our preclinical model to predict efficacy in the clinic, thus creating the “Virtual Tumour Clinical.” Here we show a comparative study of the preclinical Virtual Tumour calibrated model for prostate tumor xenografts in mice, with a Virtual Tumour Clinical version calibrated with a clinical data set comprising 53 prostate cancer patients treated with thalidomide, 25 treated with docetaxel and 50 treated with a docetaxel and thalidomide combination. PSA measurements were used as proxy for tumor size. We analysed the consistency, the capability and the limitations of the models in translating the effect of the drug combination from the preclinical situation to the clinic.

Citation Format: Eric Fernandez, Hitesh Mistry, Frances Brightman, David Orrell, William L. Dahut, William D. Figg, Wilfried D. Stein, Christophe Chassagnole. Translational modeling of docetaxel-thalidomide combination treatment in metastatic, castrate-resistant prostate cancer: predicting clinical response using preclinical data. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 366. doi:10.1158/1538-7445.AM2014-366
Abstract 3663: Immune suppressive myeloid cells expansion in vitro requires a simulated tumor microenvironment
Authors: Markowitz Joseph , Brooks Taylor R. , Carson William E.
Published: May 2015

Elevated levels of immune suppressor cells induced by tumor derived factors are associated with inhibition of immune responses. One such group of immune suppressor cells is Myeloid Derived Suppressor Cells (MDSCs). They are phenotypically defined in humans as HLADR low or negative and CD33, CD11b, and CD15 or CD14 positive and functionally by their ability to suppress the activation of other immune cells such as T cells. We hypothesized that MDSCs generated in vitro could be propagated but would lose their suppressive ability on the action of T cells in the absence of a simulated tumor microenvironment. MDSCs were first differentiated in vitro from normal donor peripheral blood mononuclear cells (PBMCs) using a simulated tumor microenvironment consisting of either cytokines (GMCSF and IL-6) or tumor cell lines (pancreatic adenocarcinoma SW-1990 or head and neck cancer CAL-27). The MDSCs generated in vitro suppressed T cell proliferation of autologous T cells using the carboxyfluorescein succinimidyl ester (CFSE) assay (31%, 46% and 75% suppression for cytokines, CAL-27 and SW-1990, respectively). Given the MDSCs obtained from co-culture of PBMCs with SW-1990 were the most suppressive on the proliferation of T cells, we attempted to proliferate these cells in vitro to expand the number of MDSC available for experimentation rather than having to purify MDSC from clinical samples. In vitro generated MDSC were propagated using a Rho-associated kinase (ROCK) inhibitor. Generated MDSC collected via magnetic bead isolation (CD33) were cultured with irradiated fibroblast feeder cells (± 3000 rad) in F medium [3:1 (v/v) F-12 Nutrient Mixture (Ham)-Dulbecco's modified Eagle's medium, 5% fetal bovine serum, 0.4 µg/mL hydrocortisone, 5 µg/mL insulin, 8.4 ng/mL cholera toxin, 10 ng/mL epidermal growth factor (or 10 ng/mL GMCSF), and 24 µg/mL adenine]. The ROCK inhibitor was added at 10 µM and the media was exchanged every 3 days. The generated MDSC did propagate as measured by CFSE staining of the generated MDSC; however, the generated MDSCs completely lost their ability to inhibit T cells by day 11 in culture. In conclusion, generated MDSCs can be propagated in culture but lose their ability to suppress T cells in the absence of a simulated tumor microenvironment and the presence of a Rho kinase inhibitor.

Citation Format: Joseph Markowitz, Taylor R. Brooks, William E. Carson. Immune suppressive myeloid cells expansion in vitro requires a simulated tumor microenvironment. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3663. doi:10.1158/1538-7445.AM2014-3663
Abstract 3666: Tumor microenvironment modulation enhances macrophage polarization and T-cell activation resulting in synergized anti-cancer effect
Authors: Huang Yuhui , Jiang Wen , Kim Betty Y.S.
Published: May 2015

Solid cancer microenvironment is heterogeneous in nature, comprised of many stromal cell types that work in concert with malignant tumor cells for its maintenance and growth. Among these cell types, tumor-associated macrophages (TAMs) represent a dominant cell population with immunosuppressive and proangiogenic properties. Present in most human cancers, histologic evidence of increased TAMs is often associated with poorer prognosis. Here, we show that the immunosuppressive properties of TAMs can be overcome by modulating tumor microenvironmental parameters. By restoring tumor vascular network and tissue perfusion via anti-vascular endothelial growth factor receptor 2 (VEGFR2) antibody, significant reduction of tumor tissue hypoxia and acidosis can be accomplished. Alteration of these local environmental factors directly results in the polarization of TAMs from an immune inhibitory (M2-like) towards immune stimulatory (M1-like) phenotype. In addition, the restoration of aberrant and dysfunctional tumor vessels leads to changes in local cytokine and protein expression profiles that favor the recruitment of immune effector cells, including increased CXCL9 expression. As a result, both CD4+ helper and CD8+ cytotoxic T cell populations were significantly increased within the tumor. Based on these findings, we tested the utility of combining the optimal dose of anti-VEGFR2 antibody therapy with B7-H4 blockade in breast cancer models. Our preliminary data suggests that such treatment enhanced TAM polarization to an anti-tumor phenotype, increased tumor immune cell infiltration and decreased T-cell inhibition, resulting in synergized anti-cancer effects. Therefore, these results suggest that tumor microenvironment modulating therapy provides a new and interesting strategy to augment active immunotherapy for breast as well as other cancer types.

Citation Format: Yuhui Huang, Wen Jiang, Betty Y.S. Kim. Tumor microenvironment modulation enhances macrophage polarization and T-cell activation resulting in synergized anti-cancer effect. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3666. doi:10.1158/1538-7445.AM2014-3666
Abstract 368: Inferred regulatory interaction network from prostate cancer reveals potential regulators coordinating progression and metastasis
Authors: Dai Xudong , Gong Yixuang , Galsky Matthew D. , Schadt Eric E. , Oh William K. , Zhu Jun
Published: May 2015

Introduction: Significant genomic reprogramming occurs when prostate cancer cells acquire androgen receptor (AR) independent growth and metastasize to remote sites. Recent whole genome sequencing studies have noted recurrent gene deletion, amplification, fusion and widespread chromoplexy associated with prostate cancer progression. Synchronizing such a multistep multicomponent process, involving both cancer cells and tumor microenvironment, to gain growth advantage is challenging without a master coordinator. Genome-wide siRNA screenings have been undertaken to identify regulators driving such a reprogramming process affecting cancer cell themselves, but not the necessary interaction with host tissue. Thereby a novel approach was developed in the current work to uncover potential regulators that coordinate major cancer hallmarks and tumor-host interactions leading to prostate cancer progression. Methods: A network of potential regulatory interactions among genes was derived from matched prostate cancer expression profiles and copy number variation (CNV) datasets. The relevance of the inferred network was examined by experimentally derived AR target networks and literature mining- based cancer gene interaction networks. Subsequently, a network topology-based matrix, critical node index, was established to identify network modules significantly associated with biochemical recurrence—a clinical surrogate marker representing prostate cancer progression. Results: When bench-marked with established prostate cancer gene sets, the critical node downstream modules were more significantly enriched for individual genes with strong prognostic power (P<1E-5). Furthermore, compounds, which are capable of reversing expression patterns of individual top critical node downstream modules, either inhibited cancer cell growth or tumor progression in public datasets and literature. More importantly, overexpression or knock down of individual critical nodes in LNCaP, 22RV1, and PC3 prostate cancer cell lines demonstrated the potential of critical nodes to coordinate AR-independent growth through regulating host tissue immune interactions. Conclusion: Our preliminary results imply the existence of network modules that can synchronize the effects of genomic reprogramming during prostate cancer progression and metastasis.

Note: This abstract was not presented at the meeting.

Citation Format: Xudong Dai, Yixuang Gong, Matthew D. Galsky, Eric E. Schadt, William K. Oh, Jun Zhu. Inferred regulatory interaction network from prostate cancer reveals potential regulators coordinating progression and metastasis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 368. doi:10.1158/1538-7445.AM2014-368
Abstract 3680: (R,R')-4′-methoxy-1-naphtylfenoterol inhibits pro-survival signaling, proliferation and motility of select human melanoma cell lines
Authors: Wnorowski Artur , Paul Rajib K. , Jimenez Lucita , Toll Lawrence , Kotb Abdelmohsen , Bernier Michel , Wainer Irving W.
Published: May 2015

(R,R')-4′-methoxy-1-naphtylfenoterol (MNF) inhibits proliferation of 1321N1 astrocytoma cell line in a β2-adrenergic receptor (β2AR)-dependent manner, while reducing rat C6 glioma growth in a xenograft mouse model. MNF interferes also with pro-oncogenic signaling induced by the orphan G-protein-linked receptor, GPR55, in human HepG2 hepatocarcinoma and PANC-1 pancreatic carcinoma cells. Here, we utilized a number of cellular and biochemical techniques to investigate the anti-tumorigenic potential of MNF in a panel of human melanoma cell lines. Cell migration, which was performed using scratch wound healing assay, demonstrated the ability of MNF to dose-dependently reduce the motility of EGFR-expressing UACC-647 and M93-047 cell lines in a biphasic (IC50 = 0.32 nM and 1.48 µM) and sigmoidal (IC50 = 7.14 nM) fashion, respectively. In contrast, EGFR-negative UACC-903 cells were unresponsive to MNF. The proliferation rate of UACC-647 cells was diminished by 38 ± 3.7% in the presence of MNF, with an IC50 of 136 nM. Pharmacological inhibition of β2AR with the selective antagonist ICI-118,551 (50 nM) did not confer protection against the actions of MNF on cell motility and proliferation. U-shaped dose-response curves, with a nadir at 1 nM, were observed when the levels of phosphorylated MEK and ERK were determined in MNF-treated UACC-647 cells. A similar dose-response relationship was observed with the β2AR agonists, epinephrine and (R,R')-fenoterol, and cell pretreatment with 50 nM ICI-118,551 abolished MNF-evoked drop in phospho-ERK levels. However, MNF markedly increased inhibitory phosphorylation of eukaryotic elongation factor 2 (eEF2) on Thr56, which was accompanied by global defect in protein biosynthesis as revealed by pulse-labeling experiments with [35S]-methionine. We conclude that the anti-tumorigenic functions of MNF most likely reflect dual effects of MNF through interaction with different receptors in melanoma. This work was supported entirely by the Intramural Research Program of the National Institute on Aging/NIH and NIA/NIH contract N01-AG31009, and the Foundation for Polish Science (TEAM 2009-4/5 programme).

Citation Format: Artur Wnorowski, Rajib K. Paul, Lucita Jimenez, Lawrence Toll, Abdelmohsen Kotb, Michel Bernier, Irving W. Wainer. (R,R')-4′-methoxy-1-naphtylfenoterol inhibits pro-survival signaling, proliferation and motility of select human melanoma cell lines. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3680. doi:10.1158/1538-7445.AM2014-3680
Abstract 3687: Analgesic effects of capsazepine on a mouse model of bone cancer pain
Authors: Balenko Matthew D. , Seidlitz Eric , Singh Gurmit
Published: May 2015

Breast cancer has the highest incidence rate in women, accounting for more than 22% of all cancers and possessing a strong disposition to metastasize to bone. Discovered in 70% of advanced breast cancer patients and over 90% prevalence in patients who die from breast cancer, the affinity of this disease to metastasize bone has become overwhelming. These skeletal metastases become a significant cause of morbidity and mortality in patients with the primary symptom being pain. Pain is a major concern in determining a patient's quality of life and there have been many attempts to understand and control bone pain with little success.

Capsazepine was identified during a high throughput screen to be a potent inhibitor of breast cancer cell-mediated glutamate release, a neurotransmitter with known associations in both neural communication and pain. This drug also has antagonistic effects on transient receptor potential vanilloid type 1 (TRPV-1) receptors which act as key players in both heat and vanilloid-induced nociception. Studies have shown that glutamate plays a role in bone cancer pain, with an excess in free glutamate able to cause pain either directly through excitotoxic pathways or indirectly though the dysregulation of osteoclasts and osteoblasts, causing bone dysregulation. TRPV-1 receptors have also has been implicated in the mechanisms of bone cancer pain, as osteoclasts release protons during bone remodeling which can elicit a TRPV-1-related nociceptive response from neurons in the surrounding periosteum. These findings suggest that Capsazepine may provide a powerful multi-pronged approach to treating cancer pain.

A model of breast cancer-induced bone pain has been established using human MDA-MB-231 breast adenocarcinoma cells injected into the femurs of BALB/c nude mice. Behavioural tests are then preformed using the dynamic plantar aesthesiometer (DPA) and the dynamic weight bearing (DWB) devices on the affected limb for both movement-evoked and spontaneous pain-related behaviours.

Using Capsazepine, delivered intraperitoneally via mini-osmotic drug pumps (5kg/mg and 10kg/mg), we demonstrate a dose-dependent attenuation of pain behaviours in vivo. While confirming tumour presence using immunohistochemistry (IHC) we show that TRPV-1 and glutamate play an important role in the onset and severity of bone cancer pain. The use of Capsazepine was able to attenuate these pain-related behaviours without the side effects of common analgesics.

(This work is supported by the Canadian Institutes of Health Research (CIHR))

Citation Format: Matthew D. Balenko, Eric Seidlitz, Gurmit Singh. Analgesic effects of capsazepine on a mouse model of bone cancer pain. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3687. doi:10.1158/1538-7445.AM2014-3687
Abstract 3688: Arctigenin induced immunogenic cell death through a reactive oxygen species/endoplasmic reticulum stress pathway in lung cancer
Authors: Kuo Chih-Hsin , Hsu Ya-Ling , Huang Ming-Shyan
Published: May 2015

Induction of immunogenic cell death (ICD) is characterized by surface exposure of damage-associated molecular pattern molecules, which can induce a tumor-specific immune response. ICD are promising treatments for cancer by restoring efficient anti-tumor immunity. Here we reported that the arctigenin, an anti-inflammation and anti-cancer agent, elicits a potent anticancer immunity effect by inducing the translocation of calreticulin (CRT) to cell membrane and the release of high-mobility group protein 1 (HMGB1) in lung cancer. Treatment of A549 cells with arctigenin increased the production of ROS generation, which in turn triggered an endoplasmic reticulum (ER) stress response, including the phosphorylation of PERK (PKR-like endoplasmic reticulum kinase and IRE1 (inositol-requiring 1). The antioxidant, EUK8, significantly decreased CRT translocation and HMGB release by inhibiting the ER stress (PERK phosphorylation). Taken together, the production of ROS induced CRT translocation and HMGB1 release, mediating arctigenin-induced enhancement of tumor immunity. This indicated that arctigenin is a potential immunetherapeutic agent against lung tumor.

Citation Format: Chih-Hsin Kuo, Ya-Ling Hsu, Ming-Shyan Huang. Arctigenin induced immunogenic cell death through a reactive oxygen species/endoplasmic reticulum stress pathway in lung cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3688. doi:10.1158/1538-7445.AM2014-3688