Introduction
Redx Pharma is developing RXC008, a gastrointestinal (GI)-restricted highly potent and selective ‘pan-ROCK’ inhibitor (inhibits rho-associated coiled-coil kinase ROCK1 and ROCK2) to treat fibrostenosis due to Crohn’s disease. ROCK1/2 are key signalling kinases in fibrosis and suppression of ROCK activity is highly efficacious in multiple rodent models of fibrotic disease. Systemic inhibition of both ROCK1/2 leads to hypotension limiting its therapeutic use; RXC008 is therefore designed to be restricted to the GI tract with minimal systemic exposure.
Aims & Methods
RXC008 potency was determined against ROCK1 and ROCK2 isolated enzymes in a biochemistry assay, by surface plasmon resonance (SPR) against ROCK2 and in a T853-pMYPT1 (Thr853 phospho-Myosin Phosphatase Target Subunit 1) cell assay. Anti-fibrotic efficacy was determined in vivo using the 9-week chronic dextran sulphate sodium (DSS) and the 45-day adoptive T-cell transfer (AT) murine models of colitis, by staining with Masson’s Trichrome and by Magnetic Resonance Imaging (MRI). Gene expression changes were assessed by RNAseq. T853-pMYPT1 was measured in tissue and cells by ELISA. RXC008 concentrations were measured by Liquid Chromatography with Tandem Mass Spectrometry in colon tissue and plasma.
Results
RXC008 inhibited ROCK1 and ROCK2 with 1.19/1.3nM potencies respectively in biochemical assays, displayed picomolar affinity for ROCK2 binding in SPR, and suppressed T853-pMYPT1 levels in HAP1 cells expressing ROCK1 and ROCK2 with a potency of 4.6nM. Blood pressure provides a sensitive biomarker for peripheral pan-ROCK inhibition. Minimal plasma exposure of RXC008 and no clinical signs associated with hypotension were detected in rats administered oral doses up to 1000 mg/kg/day for 28-days. In a dog telemetry study, minimal plasma exposure of RXC008 and no effect on blood pressure were detected at oral doses up to 1000mg/kg, while an intra-venous dose of 1mg/kg gave plasma compound concentrations ten-fold above the HAP1 cell IC50 together with a significant reduction in blood pressure.
In the DSS model, oral dosing (po) of 10mg/kg RXC008 once daily (QD) attenuated the disease activity index (diarrhea, body weight, hematochezia) and significantly (p<0.0001) suppressed colon fibrosis measured by Trichrome staining and MRI. In this model, RXC008 modulated the expression of >2000 genes (p≤0.01, log2 fold change) in the colon, including fibrotic genes Col1a1 and TGFβ, the extra-cellular matrix gene Wisp2 and tissue repair genes AhR and IL-22Rα. Monotherapy RXC008 (10, 30 or 100 mg/kg po QD) reduced the fibrosis score in the AT model (P<0.05), in contrast the anti-inflammatory agents anti-p40 or anti-TNFα alone had no effect on fibrosis. RXC008 (3, 10 and 30 mg/kg po QD) also significantly (p<0.05) reduced fibrosis in combination with anti-TNFα. Minimal to no plasma exposure of RXC008 was detected in the AT model despite efficacious GI tissue exposure. RXC008 significantly reduced T853-pMYPT1 levels in colon tissue in healthy and AT mice. Ex-vivo, RXC008 suppressed T853-pMYPT1 in fibroblasts (pIC50s: 7.83, 9.55, 9.02) isolated from fibrostenotic resection tissue from three Crohn’s Disease patients.
Conclusion
RXC008 suppresses fibrosis in animal models of colitis and is being developed as a novel first-in-class therapy to inhibit fibrotic stricture formation in patients with Crohn’s Disease. A first-in-human clinical trial (ISRCTN82038982) commenced in February 2024 and will include assessment of T853-pMYPT1 in tissue biopsy samples for target engagement.
Disclosure
Kirsty Houslay, Amy Marshall, Manuela La Montagna, Katie Anderson, Kay Eckersley, Clifford D. Jones, Andrew Belfield, Helen Mckeever, Helen Timmis, Richard Armer, Caroline Phillips, Elaine Kilgour are employees of Redx Pharma. Debby Laukens has received research funding from Redx Pharma.
