Transforming growth factor-beta (TGF-β) is a secreted cytokine that signals via serine/threonine kinase receptors and SMAD effectors. Although TGF-β acts as a tumor suppressor during the early stages of tumorigenesis, it supports tumor progression in advanced stages. Indeed, TGF-β can modulate the tumor microenvironment by modifying the extracellular matrix and by sustaining a paracrine interaction between neighboring cells. Due to its critical role in cancer development and progression, a wide range of molecules targeting the TGF-β signaling pathway are currently under active clinical development in different diseases. Here, we focused on the role of TGF-β in modulating different pathological processes with a particular emphasis on gastrointestinal tumors.

The TGFβ receptor inhibitor galunisertib demonstrated efficacy in patients with pancreatic ductal adenocarcinoma (PDAC) in the randomized phase II H9H-MC-JBAJ study, which compared galunisertib plus the chemotherapeutic agent gemcitabine with gemcitabine alone. However, additional stromal paracrine signals might confer adaptive resistance that limits the efficacy of this therapeutic strategy. Here, we found that autotaxin, a secreted enzyme that promotes inflammation and fibrosis by generating lysophosphatidic acid (LPA), mediates adaptive resistance to TGFβ receptor inhibition. Blocking TGFβ signaling prompted the skewing of cancer-associated fibroblasts (CAF) toward an inflammatory (iCAF) phenotype. iCAFs were responsible for a significant secretion of autotaxin. Paracrine autotaxin increased LPA-NFκB signaling in tumor cells that triggered treatment resistance. The autotaxin inhibitor IOA-289 suppressed NFκB activation in PDAC cells and overcame resistance to galunisertib and gemcitabine. In immunocompetent orthotopic murine models, IOA-289 synergized with galunisertib in restoring sensitivity to gemcitabine. Most importantly, treatment with galunisertib significantly increased plasma levels of autotaxin in patients enrolled in the H9H-MC-JBAJ study, and median progression-free survival was significantly longer in patients without an increase of autotaxin upon treatment with galunisertib compared with those with increased autotaxin. These results establish that autotaxin secretion by CAFs is increased by TGFβ inhibition and that circulating autotaxin levels predict response to the combination treatment approach of gemcitabine plus galunisertib. Significance: TGFβ inhibition skews cancer-associated fibroblasts toward an inflammatory phenotype that secretes autotaxin to drive adaptive resistance in PDAC, revealing autotaxin as a therapeutic target and biomarker of galunisertib response.

The TGFβ receptor inhibitor galunisertib (GAL) has been demonstrated as an effective strategy for the treatment of pancreatic cancer (PC) patients, in part through the modulation of stromal microenvironment. However, alternative pathways driving stromal paracrine signals might impair its effect. Autotaxin (ATX) is an enzyme that converts lysophosphatidylcholine into its bioactive lipid, lysophosphatidic acid (LPA), which in turn promotes inflammation and fibrosis, and contributes to treatment resistance. IOA-289 is a novel ATX inhibitor in clinical development for the treatment of tumors burdened with a high degree of stromal involvement. Here, we hypothesized that ATX could sustain an adaptive response upon inhibition of TGFβ receptor signaling, impairing GAL antitumor activity.Among ten different murine PC orthotopic models, mice bearing RC416 tumors had the highest plasma levels of TGFβ1 as well as high infiltration of cancer associated fibroblasts (CAFs) and fibrosis as measured by Masson’s staining. In coculture models of RC416 with murine pancreatic stellate cells (mPSC4), we measured a significant overexpression of ATX upon treatment with galunisertib mainly by mPSC4, which showed a skewing toward an inflammatory iCAF phenotype through the upregulation of CXCL1 and LY6C1 and the down-regulation of myCAF markers including ACTA2 and Taglin. In the RC416-mPSC4 coculture model, anti-tumor activity was achieved with a combination of gemcitabine plus GAL plus IOA-289, whereas these treatments were ineffective in single culture conditions. Mice bearing RC416 tumors reached a statistically significantly longer survival following treatment with gemcitabine plus IOA-289 or gemcitabine plus GAL [median overall survival (mOS)= 35 days, p<0.05] compared with any single treatment or no treatment control. Most importantly, mice treated with the triple combination of gemcitabine plus IOA-289 and galunisertib had a significantly longer survival than any double combination treatments [mOS=47 days, p<0.05]. In mice treated with gemcitabine in combination with IOA-289 and/or GAL we measured a significantly reduced infiltration of CAFs, a reduction in plasma levels of CTGF and a diminished degree of fibrosis compared with any single control treatment.In conclusion, we demonstrate that the overexpression of autotaxin by stromal PSC might be an adaptive mechanism in response to TGFβ receptor inhibition. IOA-289 is a novel agent that warrants further clinical development in combination with gemcitabine plus GAL for the treatment of PC patients. Citation Format: Fabio Sabbadini, Monica Bertolini, Domenico Mangiameli, Serena Contarelli, Zoë Johnson, Michael M. Lahn, Silvia Pietrobono, Davide Melisi. Targeting stromal autotaxin synergizes with TGF beta inhibition in pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1064.

The TGFβ receptor inhibitor galunisertib demonstrated efficacy in patients with pancreatic ductal adenocarcinoma (PDAC) in the randomized phase II H9H-MC-JBAJ study, which compared galunisertib plus the chemotherapeutic agent gemcitabine with gemcitabine alone. However, additional stromal paracrine signals might confer adaptive resistance that limits the efficacy of this therapeutic strategy. Here, we found that autotaxin, a secreted enzyme that promotes inflammation and fibrosis by generating lysophosphatidic acid (LPA), mediates adaptive resistance to TGFβ receptor inhibition. Blocking TGFβ signaling prompted the skewing of cancer-associated fibroblasts (CAF) toward an inflammatory (iCAF) phenotype. iCAFs were responsible for a significant secretion of autotaxin. Paracrine autotaxin increased LPA-NFκB signaling in tumor cells that triggered treatment resistance. The autotaxin inhibitor IOA-289 suppressed NFκB activation in PDAC cells and overcame resistance to galunisertib and gemcitabine. In immunocompetent orthotopic murine models, IOA-289 synergized with galunisertib in restoring sensitivity to gemcitabine. Most importantly, treatment with galunisertib significantly increased plasma levels of autotaxin in patients enrolled in the H9H-MC-JBAJ study, and median progression-free survival was significantly longer in patients without an increase of autotaxin upon treatment with galunisertib compared with those with increased autotaxin. These results establish that autotaxin secretion by CAFs is increased by TGFβ inhibition and that circulating autotaxin levels predict response to the combination treatment approach of gemcitabine plus galunisertib.Significance:TGFβ inhibition skews cancer-associated fibroblasts toward an inflammatory phenotype that secretes autotaxin to drive adaptive resistance in PDAC, revealing autotaxin as a therapeutic target and biomarker of galunisertib response.