PROJECT SUMMARY/ABSTRACT
Pancreatic ductal adenocarcinoma (PDA), is a lethal malignancy, due in part to a dense, desmoplastic (fibrotic)
tumor microenvironment which aids tumor growth and inhibits drug delivery. Poor survival rates, harsh
treatments, and tumor intrinsic aspects of the disease such as pain and altered cytokine pools, promote high
levels of anxiety in PDA patients. Nearly 25% of pancreatic cancer patients are prescribed benzodiazepines
(BZDs) to treat anxiety, insomnia, or to relieve nausea. Epidemiological studies indicate that BZDs may increase
the risk of cancer development but the role of BZDs in modifying the tumor microenvironment is unknown. Recent
studies indicate that n-unsubstituted BZDs promote the signaling of the proton-sensing G protein-coupled
receptor, GPR68, under acidic conditions, such as those present in the PDA tumor microenvironment. GPR68
is overexpressed by PDA cancer-associated fibroblasts (CAFs), and its expression drives desmoplasia, fibrosis,
and inflammation, microenvironmental features associated with impaired drug delivery and chemoresistance.
To establish clinical relevance, covariate adjusted analyses was conducted of pancreatic cancer patients who
received chemotherapy at Roswell Park from 2004 to 2020. Patients receiving lorazepam (LOR), a strong
GPR68 activator, had significantly decreased progression-free survival (PFS) relative to non-users (HR 3.83
(1.53,9.57)), while patients receiving alprazolam (ALP), a GPR68 non-activator, had significantly improved PFS
(HR 0.38 (0.16-0.92)). Preliminary studies using subcutaneous allografts derived from KPC mice, a genetically
engineered mouse model of PDA, indicated that LOR promoted collagen deposition, desmoplasia, and ischemic
necrosis in vivo, supporting that this BZD may promote an unfavorable tumor microenvironment that could
negatively impact PDA patient survival. Additionally, LOR-treated CAFs had increased expression of pro-
inflammatory and pro-fibrotic genes, suggesting that CAFs may be driving the observed phenotype.
These findings led to the central hypothesis that GPR68-activating BZDs modulate CAF signaling, which
will increase desmoplasia, subsequently constricting the tumor vasculature. The long-term goal is to
determine the impact of BZDs on the PDA tumor microenvironment and chemotherapeutic efficacy. This
hypothesis will be tested with two aims. Aim 1 will determine the impact of LOR/ALP on the PDA vasculature
and desmoplasia in tumor-bearing KPC mice, using imaging and histological methods. Aim 2 will evaluate the
role of BZDs in modifying CAF signaling and activity. Changes in the expression/secretion of pro-fibrotic proteins,
inflammatory cytokines, and GPR68 downstream signaling molecules by BZD-treated immortalized CAFs will be
quantified. Alterations in PDA organoid proliferation and collagen contraction by BZD-treated CAFs will also be
quantified. This research is clinically significant because it will indicate if BZDs commonly prescribed to PDA
patients modify the PDA tumor microenvironment in a manner which may decrease chemotherapeutic efficacy.