No abstract

Several innate sensing pathways contribute to the control of early cytomegalovirus (CMV) infection, leading to a multiphasic type I interferon (IFN-IC ytomegalovirus (CMV) (human herpesvirus 5 [HHV-5], a betaherpesvirus) is one of the most common etiological agents of chronic viral infection in humans, with primary infection developing in the majority of people at relatively young ages and lasting for life (1). Although acute CMV infection is mostly asymptomatic in healthy individuals, debilitating and even fatal complications arise in those with weakened immune systems, including pregnant women, infants, transplant recipients, and HIV patients. Remarkably, a recent analysis of monozygotic twin pairs revealed that CMV infection was the primary factor driving nonheritable diversity in the immune system (2). Consistent with the dramatic impact of CMV on the host immune system, infection has been associated with multiple chronic inflammatory conditions, including high blood pressure, heart disease/atherosclerosis, cancer, and aging-related immunodeficiencies (3, 4). Despite the clinical importance of human CMV (HCMV), there is currently no vaccine, and antiviral drugs are relatively toxic and cannot restrict viral persistence/latency.Both mouse CMV (MCMV) and HCMV contain a linear double-stranded DNA (dsDNA) genome of ϳ230 kb encoding hundreds of viral open reading frames (ORFs) (5, 6). CMV harbors an arsenal of immunoregulatory genes that allow the virus to evade or dampen host immune defenses. However, despite these numerous strategies, the virus still induces significant levels of type I interferon (IFN-I) from infected cells both in vitro and in vivo. We have previously shown that MCMV induces a biphasic IFN-I response in vivo, with the first wave of IFN-I being completely independent of Toll-like receptor (TLR), Nalp3, and MAVS and emanating from virally infected stromal cells during the first 12 h (7,8). This "first burst" of IFN-I is followed by a later wave produced at 36 to 44 h by plasmacytoid dendritic cells (DCs) (pDCs) and conventional DCs (cDCs) in response to viral particles released from the infected stroma (7,(9)(10)(11). It is the first wave of IFN-I that is critical to limit viral spread in the peripheral organs (7), while the second pDC-derived wave is mostly dispensable for restricting MCMV replication levels (12). Despite our previous work showing that lymphotoxin (LT) signaling is required to promote an optimal first-phase IFN-I response to MCMV in vivo (7,8,11,13,14), the specific innate sensing pathway required for this initial production has remained uncharacterized, although

Endoplasmic reticulum (ER) stress and the unfolded protein response (UPR) signaling have been shown to be dysregulated in multiple cancer types. Glucose regulatory protein 78 (GRP78), the master regulator of the UPR, plays a role in proliferation, invasion, and metastasis in cancer. Cancer stem cells (CSCs) make up a crucial component of the tumor heterogeneity in pancreatic cancer, as well as other cancers. “Stemness” in pancreatic cancer defines a population of cells within the tumor that have increased therapeutic resistance as well as survival advantage. In the current study, we investigated how GRP78 was responsible for maintaining “stemness” in pancreatic cancer thereby contributing to its aggressive biology. We determined that GRP78 downregulation decreased clonogenicity and self-renewal properties in pancreatic cancer cell lines in vitro. In vivo studies resulted in delayed tumor initiation frequency, as well as smaller tumor volume in the shGRP78 groups. Additionally, downregulation of GRP78 resulted in dysregulated fatty acid metabolism in pancreatic tumors as well as the cells. Further, our results showed that shGRP78 dysregulates multiple transcriptomic and proteomic pathways that involve DNA damage, oxidative stress, and cell death, that were reversed upon treatment with a ROS inhibitor, N-acetylcysteine. This study thus demonstrates for the first time that the heightened UPR in pancreatic cancer may be responsible for maintenance of the “stemness” properties in these cells that are attributed to aggressive properties like chemoresistance and metastasis.

Resident fibroblasts that contact tumor epithelial cells (TEC) can become irreversibly activated as cancer-associated-fibroblasts (CAF) which stimulate oncogenic signaling in TEC. In this study, we evaluated the crosstalk between CAF and TEC isolated from tumors generated in a mouse model of KRAS/mutp53-induced pancreatic cancer (KPC mice). Transcriptomic profiling conducted after treatment with the anticancer compound Minnelide revealed deregulation of the TGF-β signaling pathway in CAF, resulting in an apparent reversal of their activated state to a quiescent, non-proliferative state. TEC exposed to media conditioned by drug-treated CAF exhibited a decrease in oncogenic signaling as manifested by downregulation of the transcription factor Sp1. This inhibition was rescued by treating TEC with TGF-β. Given promising early clinical studies with Minnelide, our findings suggest that approaches to inactivate CAF and prevent tumor-stroma crosstalk may offer a viable strategy to treat pancreatic cancer.

Pancreatic tumors are renowned for their extremely hypoxic centers, resulting in upregulation of a number of hypoxia mediated signaling pathways including cell proliferation, metabolism and cell survival. Previous studies from our laboratory have shown that Minnelide, a water-soluble pro-drug of triptolide (anti-cancer compound), decreases viability of cancer cells in vitro as well as in vivo. However, its mechanism of action remain elusive. In the current study we evaluated the effect of Minnelide, on hypoxia mediated oncogenic signaling as well as stemness in pancreatic cancer. Minnelide has just completed Phase 1 trial against GI cancers and is currently awaiting Phase 2 trials. Our results showed that upon treatment with triptolide, HIF-1α protein accumulated in pancreatic cancer cells even though hypoxic response was decreased in them. Our studies showed even though HIF-1α is accumulated in the treated cells, there was no decrease in HIF-1 binding to hypoxia response elements. However, the HIF-1 transcriptional activity was significantly reduced owing to depletion of co-activator p300 upon treatment with triptolide. Further, treatment with triptolide resulted in a decreased activity of Sp1 and NF-kB the two major oncogenic signaling pathway in pancreatic cancer along with a decreased tumor initiating cell (TIC) population in pancreatic tumor.