Untangling the tumorigenic role of homotrimeric collagen I

Pancreatic ductal adenocarcinoma is characterized by a complex microenvironment. In this issue of Cancer Cell, Chen and colleagues define an oncogenic role of tumor-cell-produced collagen I homotrimers, wherein tumor development is promoted by integrin α3/β1-dependent activation of tumor cell signaling and modulation of tumor microbiome and immunity. Pancreatic ductal adenocarcinoma is characterized by a complex microenvironment. In this issue of Cancer Cell, Chen and colleagues define an oncogenic role of tumor-cell-produced collagen I homotrimers, wherein tumor development is promoted by integrin α3/β1-dependent activation of tumor cell signaling and modulation of tumor microbiome and immunity. Solid tumors develop as abnormal tissue ecosystems with an irregular extracellular matrix (ECM) that establishes a biophysically and biochemically pro-tumorigenic environment (Cox, 2021Cox T.R. The matrix in cancer.Nat. Rev. Cancer. 2021; 21: 217-238Crossref PubMed Scopus (295) Google Scholar). This is particularly evident in pancreatic ductal adenocarcinoma (PDAC), in which the majority of the tumor is made up of host stromal cells and an abundant ECM, such that malignant cells constitute less than 15% of the tumor on average (Ho et al., 2020Ho W.J. Jaffee E.M. Zheng L. The tumour microenvironment in pancreatic cancer — clinical challenges and opportunities.Nat. Rev. Clin. Oncol. 2020; 17: 527-540Crossref PubMed Scopus (433) Google Scholar). Pleiotropic tumor-promoting effects have been ascribed to the stiffened ECM, and these effects include activation of mechano-signaling pathways to augment oncogenic signaling, retention of growth factors and cytokines to alter local biochemical signaling pathways, supplying nutrients for tumor cells, and driving an increase in the interstitial pressure that collapses vessels and thus impedes blood flow (Ho et al., 2020Ho W.J. Jaffee E.M. Zheng L. The tumour microenvironment in pancreatic cancer — clinical challenges and opportunities.Nat. Rev. Clin. Oncol. 2020; 17: 527-540Crossref PubMed Scopus (433) Google Scholar). Fibrillar collagens, such as collagen I, make up the majority of the ECM in PDAC and can be produced by both cancer-associated fibroblasts (CAFs) and pancreatic cancer cells (PCCs), which contribute approximately 90% and 10%, respectively (Tian et al., 2019Tian C. Clauser K.R. Öhlund D. Rickelt S. Huang Y. Gupta M. Mani D.R. Carr S.A. Tuveson D.A. Hynes R.O. Proteomic analyses of ECM during pancreatic ductal adenocarcinoma progression reveal different contributions by tumor and stromal cells.Proc. Natl. Acad. Sci. USA. 2019; 116: 19609-19618Crossref PubMed Scopus (187) Google Scholar). Collagen I is synthesized as a precursor with a central triple-helix region and globular peptide extensions at both the N and C termini. The pro-collagen is subsequently matured by proteases, such as BMP1, to release the central triple-helical region, and is post-translationally modified for tensile strength and structural integrity (Barker et al., 2012Barker H.E. Cox T.R. Erler J.T. The rationale for targeting the LOX family in cancer.Nat. Rev. Cancer. 2012; 12: 540-552Crossref PubMed Scopus (428) Google Scholar). Most collagen I is produced as heterotrimers (α1, α2, α1). However, expression of collagen homotrimers (α1, α1, α1) has been reported in embryonic tissue as well as in cancer and fibrosis (Makareeva et al., 2010Makareeva E. Han S. Vera J.C. Sackett D.L. Holmbeck K. Phillips C.L. Visse R. Nagase H. Leikin S. Carcinomas contain a matrix metalloproteinase-resistant isoform of type I collagen exerting selective support to invasion.Cancer Res. 2010; 70: 4366-4374Crossref PubMed Scopus (81) Google Scholar). Recent studies have suggested a tumor-restraining effect of CAF-produced collagen I (Chen et al., 2021Chen Y. Kim J. Yang S. Wang H. Wu C.-J. Sugimoto H. LeBleu V.S. Kalluri R. Type I collagen deletion in αSMA+ myofibroblasts augments immune suppression and accelerates progression of pancreatic cancer.Cancer Cell. 2021; 39: 548-565.e6Abstract Full Text Full Text PDF PubMed Scopus (202) Google Scholar), but whether PCC-produced collagen I is functionally different in PDAC is not as well understood. To address this, Chen and colleagues (Chen et al., 2022Chen Y. Yang S. Tavormina J. Tampe D. Zeisberg M. Wang H. Mahadevan K. Wu C.-J. Sugimoto H. Chang C.-C. et al.An oncogenic collagen I momotrimer from cancer cells bind to a3b1 intrgrin and impacts tumor microbiome and immunity to promote pancreatic cancer.Cancer Cell. 2022; 40Abstract Full Text Full Text PDF Scopus (43) Google Scholar) begin by examining the expression of the two genes (COL1A1 and COL1A2) that make up heterotrimeric collagen I, only to find that the COL1A2 promoter is hypermethylated in established human and murine PCCs, effectively silencing its expression and forcing a switch to collagen I homotrimer formation. To further examine the expression pattern of collagen I, the authors make use of various genetically engineered murine models (GEMMs) of PDAC. Expression of activated KRASG12D in the pancreas, using Pdx1-driven Cre recombinase, KrasG12D/wt;Pdx1-Cre (KC), recapitulates early stage disease and progress to overt PDAC at low penetrance. When this is combined with loss of the tumor suppressor p53, KrasG12D/wt;Trp53fl/fl;Pdx1-Cre (KPPC) or mutant p53, KRasG12D/wt;Trp53R172H/wt;Pdx1-Cre (KPR172HC), animals develop PDAC at high penetrance. Importantly, the ensuing tumors faithfully mimic cardinal features of the desmoplastic tumor microenvironment. When the authors examine PCCs isolated from early stage tumors in the KPPC and KC models, they observe a similar switch from heterotrimeric to homotrimeric collagen I, which suggests that this switch may be an early requirement for PDAC progression. To then test whether homotrimeric collagen I plays a functional role in PDAC development, the authors cross a conditional knockout of Col1a1 with the three different PDAC GEMMs: KPPC, KC, and KPR172HC, to specifically ablate Col1a1 expression in epithelial cancer cells. Indeed, loss of Col1a1 significantly delays PDAC development, and there is a concomitant decrease in the abundance of acinar to ductal metaplasia (ADM) and pancreatic intraepithelial neoplasia (PanIN), a reduced tumor size, and a resulting increase in overall survival. As anticipated, loss of epithelial homotrimeric collagen I is associated with reduced collagen I levels at early developmental stages. However, the authors observe no discernible differences in total collagen I, collagen networks, laminin, collagen IV, and thrombospondin one levels between fully developed wild-type and collagen I knockout tumors. Loss of homotrimeric collagen I significantly reduces malignant properties of PCCs isolated from the KPPC model (KPPCΔColI). Specifically, the authors observe that KPPCΔColI cells are less proliferative, are more sensitive to the chemotherapy gemcitabine, and form a reduced number of tumor spheres and organoids. This effect can be rescued by adding back homotrimetric collagen I, which the authors isolate from a mouse model of brittle bone disease that lacks Col1a2 (Col1a2oim/oim) and thus only forms collagen I homotrimers. Only collagen homotrimers, but not heterotrimers, increase the tumor-sphere-forming ability of KPPCΔColI cells, and this finding supports the possibility that cell autonomous signaling by homotrimeric collagen I provides critical oncogenic signals. In a subsequent series of experiments, the authors show that collagen I homotrimers activate discoiden domain receptor (DDR), FAK, AKT, and MAPK signaling. This is in sharp contrast to heterotrimeric collagen I, which only has minor or no effect on these signaling pathways. Notably, this effect appears to be dependent on integrin α3/β1, where knockdown of either integrin β1 or α3 reduces collagen I-homotrimer-induced tumor cell signaling. Analysis of single-cell RNA sequencing data and immunohistochemical analysis of human PDAC reveals that integrin α3 is predominantly expressed in epithelial cancer cells. Moreover, the abundance of integrin α3 is associated with worse overall survival and a reduced level of T cells in human PDAC, suggesting a possible link between collagen I homotrimers, integrin α3, and anti-tumor immunity. Consistently, the authors note a decrease in the expression of the macrophage/myeloid-derived suppressor cells (MDSCs) attractant Cxcl5 and an increase in the expression of the T cell attractant Cxcl16 in KPPCΔColI tumor cells. This is associated with reduced infiltration of CD11bpos/Gr1pos MDSCs and an increased infiltration of CD4pos and CD8pos T cells in tumors from KPPCΔColI animals. These changes in the immune landscape are functionally important, which is illustrated by an improved response to the immune checkpoint inhibitor anti-PD-1, specifically in the KPPCΔColI animals. Finally, loss of collagen I is also correlated with a shift from an anaerobic to a microaerophilic tumor microbiome as well as a normalization of the gut microbiome in the KPPCΔColI animals with a taxonomy pattern resembling that of tumor-free wild-type littermates. Previous efforts to broadly target the tumor microenvironment in PDAC have emphasized dichotomous pro- and anti-tumor roles of the tumor ecosystem, where ablation of the predominant ECM-producing αSMApos CAFs, also known as myofibroblastic CAFs (myCAFs), or loss of sonic hedgehog paracrine signaling, leads to tumor softening but accelerated metastasis and reduced survival (Helms et al., 2020Helms E. Onate M.K. Sherman M.H. Fibroblast Heterogeneity in the Pancreatic Tumor Microenvironment.Cancer Discov. 2020; 10: 648-656Crossref PubMed Scopus (155) Google Scholar). Similarly, ablation of Col1a1 specifically in αSMApos CAFs also accelerates disease progression (Chen et al., 2021Chen Y. Kim J. Yang S. Wang H. Wu C.-J. Sugimoto H. LeBleu V.S. Kalluri R. Type I collagen deletion in αSMA+ myofibroblasts augments immune suppression and accelerates progression of pancreatic cancer.Cancer Cell. 2021; 39: 548-565.e6Abstract Full Text Full Text PDF PubMed Scopus (202) Google Scholar). The resulting tumors are softer, display more prominent ADM and PanIN lesions, and show increased expression of Cxcl5 and decreased expression of Cxcl16 with concomitant increased infiltration of MDSCs and exclusion of CD8pos T cells. Additionally, other studies have reported that the C-terminal processing of collagen I is decreased in PDAC. Elevated expression of BMP1, which acts as a proteinase to remove the C-terminal pro-peptide, reduces PDAC tumor growth and metastasis in orthotopic implantation models of PDAC, suggesting that accurate processing of collagen I also plays a key role in controlling PDAC progression (Tian et al., 2021Tian C. Huang Y. Clauser K.R. Rickelt S. Lau A.N. Carr S.A. Vander Heiden M.G. Hynes R.O. Suppression of pancreatic ductal adenocarcinoma growth and metastasis by fibrillar collagens produced selectively by tumor cells.Nat. Commun. 2021; 12: 2328Crossref PubMed Scopus (30) Google Scholar). Thus, a picture is emerging in which collagen I heterotrimers act in a tumor-restrictive manner, whereas PCC-expressed collagen I homotrimers act in a tumor-promoting manner (Figure 1). It is less clear whether these apparent opposite effects are due to competition for integrin signaling complexes or whether collagen I homotrimers and heterotrimers are subjected to differential post translational modification to thereby differentially tune local stiffening. The results presented here by Chen and colleagues further emphasize the need to functionally characterize the interplay between the tumor and the host. Thus, although the data describe a clear pro-tumorigenic function of homotrimeric collagen I, tumors still develop in KPPCΔColI animals. This could be due to compensatory changes in the ECM to overcome loss of homotrimeric collagen I, as illustrated by the increased level of total collagen I, picrosirious red staining (a readout of collagen networks), and collagen IV during KPPCΔColI tumor development. Moreover, integrin α3 also acts as a receptor for laminins, which are abundant components of the basement membrane, is functionally important for tumor organoid growth and correlates with worse patient outcome (Below et al., 2022Below C.R. Kelly J. Brown A. Humphries J.D. Hutton C. Xu J. Lee B.Y. Cintas C. Zhang X. Hernandez-Gordillo V. et al.A microenvironment-inspired synthetic three-dimensional model for pancreatic ductal adenocarcinoma organoids.Nat. Mater. 2022; 21: 110-119Crossref PubMed Scopus (53) Google Scholar). Nonetheless, what remains clear is that the developing tumor ecosystem in the absence of collagen I homotrimers displays clear changes in both the immune microenvironment and the microbiome and that these changes offer specific therapeutic opportunities such as increased sensitivity to immune checkpoint inhibition. Thus, irrespective of whether future therapies target the remodeled ECM or the ensuing signaling changes in the tumor cells, deep characterization of the broader tumor ecosystem is warranted to ensure that induced vulnerabilities are therapeutically exploited. Work in systems oncology is supported by a Cancer Research UK Institute Award (C5759/A27412), a Cancer Research UK Program Award (C13329/A21671), and a European Research Council Consolidator Award (ERC-2017-COG 772577). The author would like to acknowledge colleagues on the CRUK Manchester Institute Systems Oncology Team and Dr. Santiago Zelenay for valuable input. Figure 1 was created with BioRender.com. The authors declare no competing interests. Oncogenic collagen I homotrimers from cancer cells bind to α3β1 integrin and impact tumor microbiome and immunity to promote pancreatic cancerChen et al.Cancer CellJuly 21, 2022In BriefChen et al. identify that type I collagen (Col1) produced by pancreatic cancer cells is the abnormal homotrimer variant with oncogenic properties. Deletion of Col1 homotrimer in cancer cells inhibits tumor progression and reshapes the tumor microbiome. Cancer-Col1 homotrimer deletion enhances T cell infiltration and enables efficacy of anti-PD-1 immunotherapy. Full-Text PDF Open Archive