CAF hierarchy driven by pancreatic cancer cell p53-status creates a pro-metastatic and chemoresistant environment via perlecan

Claire Vennin,David Herrmann,Marina Pajic,Pauline Mélénec,Yingxiao Wang,Andrew Da Silva,Daniel Roden,Aurélie Cazet ,Zehra Elgundi,Shane T Grey,Australian Pancreatic Genome Initiative ,Anthony J Gill,Romain Rouet,Anaiis Zaratzian,Suresh Mathivanan,Monisha Samuel,Sean Warren ,Mark Pinese,Daniel Christ,Morghan C Lucas,Ashleigh Parkin,Gabriella Kalna,Jennifer P Morton,Daniel A Reed ,Kendelle J Murphy,Benjamin L Parker,Aleš Benda ,Max Nobis,A W Braithwaite ,David R Croucher,Phoebe A Phillips,John M Whitelock,Owen J Sansom,Wilfred Leung,Thomas R Cox

doi:10.1038/s41467-019-10968-6

Abstract

Heterogeneous subtypes of cancer-associated fibroblasts (CAFs) coexist within pancreatic cancer tissues and can both promote and restrain disease progression. Here, we interrogate how cancer cells harboring distinct alterations in p53 manipulate CAFs. We reveal the existence of a p53-driven hierarchy, where cancer cells with a gain-of-function (GOF) mutant p53 educate a dominant population of CAFs that establish a pro-metastatic environment for GOF and null p53 cancer cells alike. We also demonstrate that CAFs educated by null p53 cancer cells may be reprogrammed by either GOF mutant p53 cells or their CAFs. We identify perlecan as a key component of this pro-metastatic environment. Using intravital imaging, we observe that these dominant CAFs delay cancer cell response to chemotherapy. Lastly, we reveal that depleting perlecan in the stroma combined with chemotherapy prolongs mouse survival, supporting it as a potential target for anti-stromal therapies in pancreatic cancer.

Highlights

Heterogeneous subtypes of cancer-associated fibroblasts (CAFs) coexist within pancreatic cancer tissues and can both promote and restrain disease progression
Following FACS isolation, imaging of GFP in cancer cells and matched CAFs confirmed that CAFs isolated from both Genetically engineered mouse models (GEMMs) do not express E-cadherin-GFP (Fig. 1b), while immunohistochemistry (IHC) and immunofluorescence analyses of markers for CAFs29 (Fig. 1c) and pancreatic epithelial cancer cells (Fig. 1d) validated the identity and purity of the isolated CAFs
CAF activation was confirmed using an in vitro collagen contraction assay (Fig. 1e and Supplementary Fig. 1a)[22,39]. While both CAF populations were able to actively remodel collagen, there was a greater degree of contraction in CAF matrices isolated from KPC tumor compared to KPflC tumor (hereafter referred to as mutanteducated CAFs, or mt-e-CAFs and as flox-educated-CAFs, or fl-eCAFs, respectively (Fig. 1f and Supplementary Fig. 1a))

Summary

Introduction

Heterogeneous subtypes of cancer-associated fibroblasts (CAFs) coexist within pancreatic cancer tissues and can both promote and restrain disease progression. We and others have used the poorly-metastatic Pdx1-Cre; LSL-K-rasG12D/+; LSL-p53fl/+ (KPflC) and the highly metastatic Pdx1-Cre; LSL-K-rasG12D/+; LSL-p53R172H/+ (KPC) mouse models These models differ only in the p53 status of pancreatic epithelial cells and previous work demonstrated that mutant p53R172H in the epithelial cells drove the progression of invasive and metastatic PC over and above the effects of loss-ofp[538,16,17,18,19], in agreement with observations in patients[4]. We and others have previously demonstrated that re-shaping the pancreatic stroma can impair tumor progression and improve the efficacy of standard-of-care therapies[2,22,23,24,25,26,27] This feedback is reciprocal, and cancer cells with distinct characteristics have been shown to locally tune the tumor stroma[28,29,30]. Understanding the intricate, cell-specific crosstalk occurring between distinct cancer and stromal cell populations will be critical to develop effective stromal-based therapeutics for PC, which is a highly molecularly heterogeneous disease[5,9,30,37,38]

Methods

Results

Conclusion