Citations

Abstract Pancreatic cancer is the fourth leading cause of cancer death in the United States with current 5-year survival rates at 6%. Greater than 90% of all pancreatic cancers harbor an activating mutation in codon 12 of the Kras gene (KrasG12D). Protein Kinase C iota (PKCι) has been shown to mediate KrasG12D downstream signaling and acinar-to-ductal metaplasia in an explant model of pancreatic cancer initiation. Additionally, inhibition of PKCι in human pancreatic cancer cells leads to decreased tumor size, angiogenesis, and metastasis in an orthotopic tumor model, suggesting that PKCι plays a role in maintaining the oncogenic phenotype of pancreatic cancer cells. We sought to test the hypothesis that PKCι plays a role development and maintenance of pancreatic cancer using a mouse model of KrasG12D-induced pancreatic cancer. Purpose: To identify the effect of PKCι loss on pancreas function and development of KrasG12D-induced pancreatic cancer. Procedures: Pancreata from mice with pancreas-specific deletion of PKCι, with and without pancreas-specific KrasG12D expression, were analyzed for histological and protein expression alterations. Acinar cells isolated from these transgenic mouse models were subjected to in vitro analyses to determine the role of PKCι in pancreatic cell homeostasis. Results: Mice lacking pancreatic PKCι survived and developed normally. Pancreatic knockout of PKCι expression resulted in a phenotype consistent with a defect in autophagic flux. In vitro analyses of isolated acinar cells supported the hypothesis that PKCι promotes autophagy in pancreatic cells. Autophagy exhibits a dual functionality in pancreatic cancer, acting as a tumor suppressor in normal cells and a tumor promoter in established tumor cells. We sought to determine the effect of PKCι knock-out on both of these cell types in a mouse model of pancreatic cancer. Using the LSL mouse model of K-ras induced pancreatic cancer, we demonstrated that pancreatic loss of PKCι lead to an increase in the number of Pancreatic Intraepithelial (PanIN) lesions formed in young mice. However, these lesions were protected from progressing past the early PanIN1/2 stage such that no mice lacking PKCι developed cancer, even after 1.5 years of age. By contrast, over 50% of mice with wild-type PKCι developed cancer; of those without cancer, many had progressed to the PanIN-3 stage, a stage representative of carcinoma in-situ. These data are similar to that seen in mouse models of pancreatic cancer with genetic inhibition of autophagy. Analysis of PanINs in these mice revealed that those lacking PKCι expressed significantly more P62 protein, a selective autophagy substrate, compared to PanINs with wild-type PKCι, supporting a block in the autophagic flux. These data suggest that PKCι plays an important role in both the initiation and progression of pancreatic cancer, possibly by regulating autophagic flux. Impact: Inhibition of pancreatic PKCι expression prevented progression of PanIN lesions past the early PanIN1/2 stage, resulting in a complete block in cancer development in mouse models of pancreatic cancer. Our preliminary data suggests that PKCι promotes cancer progression by regulating autophagic flux, a process that has been shown to be essential for KrasG12D-mediated pancreatic cancers. Pancreatic cancer is characterized by resistance to current therapies. Novel targets for new and more effective therapies are critical to improve outcomes for patients with this deadly disease. Inhibitors of PKCι are currently in clinical trials for ovarian and lung cancers. Our data supports the feasibility of using PKCι inhibitors for therapy in pancreatic cancer. Citation Format: Kristin S. Inman, Amanda A. Francis, Nicole R. Murray. PKCι in pancreatic cancer development: A possible role for autophagy. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Innovations in Research and Treatment; May 18-21, 2014; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2015;75(13 Suppl):Abstract nr A02.

Abstract Matrix metalloproteinase 10 (Mmp10; stromelysin 2) is a member of a large family of structurally related matrix metalloproteinases, many members of which have been implicated in tumor progression, invasion and metastasis. However, emerging evidence suggests a possible role for Mmps in tumor initiation. Here we reveal an unexpected role for Mmp10 in the maintenance and tumorigenicity of mouse lung cancer stem-like cells (CSC). Mouse lung cancer cell cultures enriched in stem-like cells grow as undifferentiated tumor “oncospheres” that express elevated levels of the cancer stem cell markers Aldh1, CD133, Nanog, Notch3, Notch 4, Hey 1 and Hey 2. These cells also express elevated levels of Mmp10 mRNA and secrete elevated levels of Mmp10 protein. Functionally, these lung CSCs exhibit self-renewal, the ability to clonally expand, enhanced transforming potential in vitro, and enhanced tumorigenic properties when injected orthotopically into the lungs of syngeneic mice. RNAi-mediated knockdown of Mmp10 in these cells leads to a loss of stem cell marker gene expression and a dramatic inhibition of oncosphere growth, clonal expansion, transformed growth in vitro, and lung tumor formation and metastasis in vivo. In contrast, oncospheres implanted into syngeneic non-transgenic or Mmp10−/− mice show no significant difference in tumor initiation, growth or metastasis, demonstrating the importance of Mmp10 produced by cancer cells rather than the tumor microenvironment in lung tumor initiation and maintenance. Analysis of gene expression data from human tumors demonstrates that Mmp10 is elevated in many human tumor types including lung, head and neck, esophageal, colon, breast, melanoma, bladder, cervical, ovarian, prostate and brain. Furthermore, gene set enhancement analysis (GSEA) demonstrates that elevated Mmp10 expression correlates with tumor metastasis and with cancer stem-like genomic signatures in human lung tumors. Thus, Mmp10 is required for maintenance of a highly tumorigenic, cancer-initiating, metastatic stem-like cell population in lung cancer. Our data demonstrate for the first time that Mmp10 is a critical lung cancer stem cell gene and novel therapeutic target for lung cancer stem cells. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-204. doi:1538-7445.AM2012-LB-204

Abstract Pancreatic cancer is the fourth leading cause of cancer deaths in the United States, with an overall 5-year survival rate of <5%. Pancreatic ductal adenocarcinoma (PDAC), the most common form of pancreatic cancer, is highly resistant to conventional chemotherapies, underscoring the critical need for new molecular targets for pancreatic cancer chemotherapy. The KRAS proto-oncogene is mutated in >90% of PDAC. Protein kinase Cι (PKCι) is required for the oncogenic Ras–mediated transformed growth of lung cancer and intestinal epithelial cells. However, little is known about the role of PKCι in pancreatic cancer. In this study, we evaluated the expression of PKCι in human pancreatic cancer and the requirement for PKCι for the transformed growth and tumorigenicity of PDAC cells. We find that PKCι is significantly overexpressed in human pancreatic cancer, and high PKCι expression correlates with poor patient survival. Inhibition of PKCι expression blocks PDAC cell transformed growth in vitro and tumorigenicity in vivo. Inhibition of PKCι expression in pancreatic tumors also significantly reduces tumor angiogenesis and metastasis. Analysis of downstream PKCι effectors implicates the Rac1-MEK/ERK1/2 signaling axis in PKCι-mediated transformed growth and cellular invasion. Taken together, our data show a required role for PKCι in the transformed growth of pancreatic cancer cells and reveal a novel role for PKCι in pancreatic cancer cell metastasis and angiogenesis in vivo. Our results strongly indicate that PKCι will be an effective target for pancreatic cancer therapy. Cancer Res; 70(5); 2064–74

Abstract Colon cancer develops over a period of 10 to 15 years, providing a window of opportunity for chemoprevention and early intervention. However, few molecular targets for effective colon cancer chemoprevention have been characterized and validated. Protein kinase CβII (PKCβII) plays a requisite role in the initiation of colon carcinogenesis in a preclinical mouse model by promoting proliferation and increased β-catenin accumulation. In this study, we test the hypothesis that PKCβII is an effective target for colon cancer chemoprevention using enzastaurin (LY317615), a PKCβ-selective inhibitor, in a mouse model of colon carcinogenesis. We find that enzastaurin potently reduces azoxymethane-induced colon tumor initiation and progression by inhibiting PKCβII-mediated tumor cell proliferation and β-catenin accumulation. Biochemically, enzastaurin reduces expression of the PKCβII- and β-catenin/T-cell factor–regulated genes PKCβII, cyclooxygenase II, and vascular endothelial growth factor, three genes implicated in colon carcinogenesis. Our results show that enzastaurin is an effective chemopreventive agent in a mouse model of sporadic colon cancer that significantly reduces both tumor initiation and progression by inhibiting expression of proproliferative genes. Thus, PKCβII is an important target for colon cancer chemoprevention and the PKCβ-selective inhibitor enzastaurin may represent an effective chemopreventive agent in patients at high risk for colon cancer. [Cancer Res 2009;69(4):1643–50]

Abstract We previously showed that elevated expression of either protein kinase CβII (PKCβII) or PKCι/λ enhances colon carcinogenesis in mice. Here, we use novel bitransgenic mice to determine the relative importance of PKCβII and PKCι/λ in colon carcinogenesis in two complimentary models of colon cancer in vivo. Bitransgenic mice overexpressing PKCβII and constitutively active PKCι (PKCβII/caPKCι) or kinase-deficient, dominant-negative PKCι (PKCβII/kdPKCι) in the colon exhibit a similar increase in colon tumor incidence, tumor size, and tumor burden in response to azoxymethane (AOM) when compared with nontransgenic littermates. However, PKCβII/kdPKCι mice develop predominantly benign colonic adenomas, whereas PKCβII/caPKCι mice develop malignant carcinomas. In contrast, PKCβ-deficient (PKCβ−/−) mice fail to develop tumors even in the presence of caPKCι. Our previous data indicated that PKCβII drives tumorigenesis and proliferation by activating β-catenin/Apc signaling. Consistent with this conclusion, genetic deletion of PKCβ has no effect on spontaneous tumorigenesis in Apcmin/+ mice. In contrast, tissue-specific knockout of PKCλ significantly suppresses intestinal tumor formation in Apcmin/+ mice. Our data show that PKCβII and PKCι/λ serve distinct, nonoverlapping functions in colon carcinogenesis. PKCβII is required for AOM-induced tumorigenesis but is dispensable for tumor formation in ApcMin/+ mice. PKCι/λ promotes tumor progression in both AOM- and Apcmin/+-induced tumorigenesis. Thus, PKCβII and PKCι, whose expression is elevated in both rodent and human colon tumors, collaborate to drive colon tumor formation and progression, respectively. [Cancer Res 2009;69(2):656–62]

Suppression of colon carcinogenesis by peroxisome proliferator-activated receptor (PPAR)-γ is likely due to some effect of PPAR-γ on normal colonic epithelial cells. However, our understanding of the effects of PPAR-γ in such cells is limited. We analyzed the abundance, distribution, and function of PPAR-γ in epithelial cells isolated from the murine proximal and distal colon. Marked differences in PPAR-γ abundance and distribution were observed, suggesting tissue-specific responses. Analysis of PPAR-γ effects on DNA synthesis, formation of preneoplastic lesions, and activation of MAPK signaling in proximal and distal colonic epithelial cells in vivo indicates that PPAR-γ regulates both tissue-specific and common responses within the proximal and distal colon. Three major functional cohorts of PPAR-γ target genes were identified by genomic profiling of isolated colonic epithelial cells: genes that are involved in metabolism, in signaling, and in cellular adhesion and motility. Two subsets of PPAR-γ target genes were differentially expressed in the proximal and distal epithelium. Proximal target genes were primarily involved in metabolic activities, whereas signal transduction, adhesion, and motility targets were more pronounced in the distal colon. Remarkably, those target genes that are differentially expressed in the proximal colon were all induced on activation of PPAR-γ, whereas all target genes that are preferentially expressed in the distal colon were repressed. Our data indicate that PPAR-γ exerts both common and tissue-specific effects in the colon and challenge the general conclusions that PPAR-γ is induced on differentiation of colonic epithelial cells and that this receptor stimulates differentiated function in epithelial cells throughout the colon.