Isoform-selective TGF-ß3 inhibition for systemic sclerosis.

BACKGROUND: Transforming growth factor ß (TGF-ß) is implicated as a key mediator of pathological fibrosis, but its pleiotropic activity in a range of homeostatic functions presents challenges to its safe and effective therapeutic targeting. There are three isoforms of TGF-ß, TGF-ß1, TGF-ß2, and TGF-ß3, which bind to a common receptor complex composed of TGF-ßR1 and TGF-ßR2 to induce similar intracellular signals in vitro. We have recently shown that the cellular expression patterns and activation thresholds of TGF-ß2 and TGF-ß3 are distinct from those of TGF-ß1 and that selective short-term TGF-ß2 and TGF-ß3 inhibition can attenuate fibrosis in vivo without promoting excessive inflammation. Isoform-selective inhibition of TGF-ß may therefore provide a therapeutic opportunity for patients with chronic fibrotic disorders. METHODS: Transcriptomic profiling of skin biopsies from patients with systemic sclerosis (SSc) from multiple clinical trials was performed to evaluate the role of TGF-ß3 in this disease. Antibody humanization, biochemical characterization, crystallization, and pre-clinical experiments were performed to further characterize an anti-TGF-ß3 antibody. FINDINGS: In the skin of patients with SSc, TGF-ß3 expression is uniquely correlated with biomarkers of TGF-ß signaling and disease severity. Crystallographic studies establish a structural basis for selective TGF-ß3 inhibition with a potent and selective monoclonal antibody that attenuates fibrosis effectively in vivo at clinically translatable exposures. Toxicology studies suggest that, as opposed to pan-TGF-ß inhibitors, this anti-TGF-ß3 antibody has a favorable safety profile for chronic administration. CONCLUSION: We establish a rationale for targeting TGF-ß3 in SSc with a favorable therapeutic index. FUNDING: This study was funded by Genentech, Inc.

Inhibition of MRTF activation as a clinically achievable anti-fibrotic mechanism for pirfenidone.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a progressive fibrotic disease characterised by aberrant fibroblast/myofibroblast accumulation and excessive collagen matrix deposition in the alveolar areas of lungs. As the first approved IPF medication, pirfenidone (PFD) significantly decelerates lung function decline while its underlying anti-fibrotic mechanism remains elusive. METHODS: We performed transcriptomic and immunofluorescence analyses of primary human IPF tissues. RESULTS: We showed that myocardin-related transcription factor (MRTF) signalling is activated in myofibroblasts accumulated in IPF lungs. Furthermore, we showed that PFD inhibits MRTF activation in primary human lung fibroblasts at clinically achievable concentrations (half-maximal inhibitory concentration 50-150 µM, maximal inhibition >90%, maximal concentration of PFD in patients <100 µM). Mechanistically, PFD appears to exert its inhibitory effects by promoting the interaction between MRTF and actin indirectly. Finally, PFD-treated IPF lungs exhibit significantly less MRTF activation in fibroblast foci areas than naïve IPF lungs. CONCLUSIONS: Our results suggest MRTF signalling as a direct target for PFD and implicate that some of the anti-fibrotic effects of PFD may be due to MRTF inhibition in lung fibroblasts.

TGF-ß signaling in myeloproliferative neoplasms contributes to myelofibrosis without disrupting the hematopoietic niche.

Myeloproliferative neoplasms (MPNs) are associated with significant alterations in the bone marrow microenvironment that include decreased expression of key niche factors and myelofibrosis. Here, we explored the contribution of TGF-ß to these alterations by abrogating TGF-ß signaling in bone marrow mesenchymal stromal cells. Loss of TGF-ß signaling in Osx-Cre-targeted MSCs prevented the development of myelofibrosis in both MPLW515L and Jak2V617F models of MPNs. In contrast, despite the absence of myelofibrosis, loss of TGF-ß signaling in mesenchymal stromal cells did not rescue the defective hematopoietic niche induced by MPLW515L, as evidenced by decreased bone marrow cellularity, hematopoietic stem/progenitor cell number, and Cxcl12 and Kitlg expression, and the presence of splenic extramedullary hematopoiesis. Induction of myelofibrosis by MPLW515L was intact in Osx-Cre Smad4fl/fl recipients, demonstrating that SMAD4-independent TGF-ß signaling mediates the myelofibrosis phenotype. Indeed, treatment with a c-Jun N-terminal kinase (JNK) inhibitor prevented the development of myelofibrosis induced by MPLW515L. Together, these data show that JNK-dependent TGF-ß signaling in mesenchymal stromal cells is responsible for the development of myelofibrosis but not hematopoietic niche disruption in MPNs, suggesting that the signals that regulate niche gene expression in bone marrow mesenchymal stromal cells are distinct from those that induce a fibrogenic program.