Sletten, Arthur C.

Abstract Pancreatic cancer, one of the most lethal forms of human cancer, is largely resistant to many conventional chemotherapeutic agents. Although many therapeutic approaches focus on tumor growth, metastasis is a primary factor contributing to lethality. Therefore, novel therapies to target metastatic invasion could prevent tumor spread and recurrence resulting from local and distant metastasis. The protein Vav1 is aberrantly expressed in more than half of pancreatic cancers. Its expression promotes activation of Rac and Cdc42 and leads to enhanced invasion and migration, as well as increased tumor cell survival and proliferation, suggesting that Vav1 could be a potent therapeutic target for pancreatic cancer. The purine analogue azathioprine, well known for its function as an anti-inflammatory compound, was recently shown to function by inhibiting Vav1 signaling in immune cells. We therefore hypothesized that azathioprine could also inhibit Vav1 in pancreatic tumor cells to reduce its proinvasive functions. Indeed, we have found that treatment of cultured pancreatic tumor cells with azathioprine inhibited Vav1-dependent invasive cell migration and matrix degradation, through inhibition of Rac and Cdc42 signaling. Furthermore, azathioprine treatment decreased metastasis in both xenograft and genetic mouse models of pancreatic cancer. Strikingly, metastasis was dramatically reduced in Vav1-expressing tumors arising from p48Cre/+, KrasG12D/+, p53F/+ mice. These inhibitory effects were mediated through Vav1, as Vav1-negative cell lines and tumors were largely resistant to azathioprine treatment. These findings demonstrate that azathioprine and related compounds could be potent antimetastatic agents for Vav1-positive pancreatic tumors. Cancer Res; 75(14); 2907–15. ©2015 AACR.

Abstract Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal forms of human cancer, due in large part to a high incidence of metastasis and resistance to conventional chemotherapies. As such, new therapies are needed to halt the metastatic process. The proto-oncogene Vav1 is ectopically expressed in over half of human pancreatic cancers, and its expression correlates with a poor prognosis in humans and promotes survival and transformation in isolated tumor cells. We have found that Vav1, through its GEF activity towards Rac1, also potently induces migration by pancreatic tumor cells. In addition, Vav1 also signals through Cdc42 to drive the formation of invadopodia and matrix degradation, which also promotes tumor cell invasion. Importantly, the activation of Vav1 circumvents the requirement for active Src in this process. Because Vav1 is ectopically expressed in pancreatic cancers and is a potent regulator of tumor cell survival, proliferation, and invasive migration, it could provide a specific therapeutic target for these tumors. Recently, the anti-inflammatory agent azathioprine was described as an inhibitor of Vav1/Rac1 activity in hematopoietic cells. Therefore, we hypothesized that azathioprine could also inhibit the Vav1-driven processes of proliferation and invasion in pancreatic tumor cells. Indeed, we have determined that azathioprine treatment of cultured human PDAC cells reduces viability and proliferation. Further, similar to siRNA-mediated depletion of Vav1, azathioprine treatment reduces cell migration, as well as the matrix degradation required for invasion. Vav1-expressing tumor cells were more sensitive to azathioprine than Vav1-negative cells, supporting the premise that the inhibitory effects of azathioprine are mediated through Vav1. Finally, azathioprine also reduces metastasis in a xenograft model of pancreatic cancer. Taken together, these data uncover a role for aberrantly expressed Vav1 in regulation of the cytoskeletal machinery required for migration and invasion, and in the promotion and metastasis of pancreatic cancer. Even with the expression of multiple Rho family GEFs, tumor cells that upregulate Vav1 have become dependent upon this exchange factor, implicating Vav1 as a potent node for therapeutic intervention. Supported by R01 CA104125 and R03 CA155778 to M.A.M. Citation Format: Gina L. Razidlo, Christopher Magnine, Arthur C. Sletten, Rachel M. Hurley, Mark A. McNiven. Attenuation of pancreatic cancer cell migration and invasion through a targeted inhibition of the Rac GEF Vav1. [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 4026. doi:10.1158/1538-7445.AM2014-4026

Autophagy is a central mechanism by which hepatocytes catabolize lipid droplets (LDs). Currently, the regulatory mechanisms that control this important process are poorly defined. The small guanosine triphosphatase (GTPase) Rab7 has been implicated in the late endocytic pathway and is known to associate with LDs, although its role in LD breakdown has not been tested. In this study, we demonstrate that Rab7 is indispensable for LD breakdown (“lipophagy”) in hepatocytes subjected to nutrient deprivation. Importantly, Rab7 is dramatically activated in cells placed under nutrient stress; this activation is required for the trafficking of both multivesicular bodies and lysosomes to the LD surface during lipophagy, resulting in the formation of a lipophagic “synapse.” Depletion of Rab7 leads to gross morphological changes of multivesicular bodies, lysosomes, and autophagosomes, consequently leading to attenuation of hepatocellular lipophagy. Conclusion : These findings provide additional support for the role of autophagy in hepatocellular LD catabolism while implicating the small GTPase Rab7 as a key regulatory component of this essential process. (H epatology 2015;61:1896–1907)