Pharmacological inhibition of MERTK induces in vivo retinal degeneration: a multimodal imaging ocular safety assessment

Gregory Hamm,Paul Smith,Ruth Macdonald,Gareth Maglennon,William Mccoull,Stewart Jones,Jon Winter-Holt,Lindsay Mcmurray,Alexander R Harmer,James Blades,Julia Johansson,Philip B Rawlins,Muireann Coen,Peter Barton,Paul Fitzpatrick,Ann Doherty,Beth Williamson,Jayne Harris

doi:10.1007/s00204-021-03197-8

Abstract

The receptor tyrosine kinase, MERTK, plays an essential role in homeostasis of the retina via efferocytosis of shed outer nuclear segments of photoreceptors. The Royal College of Surgeons rat model of retinal degeneration has been linked to loss-of-function of MERTK, and together with the MERTK knock-out mouse, phenocopy retinitis pigmentosa in humans with MERTK mutations. Given recent efforts and interest in MERTK as a potential immuno-oncology target, development of a strategy to assess ocular safety at an early pre-clinical stage is critical. We have applied a state-of-the-art, multi-modal imaging platform to assess the in vivo effects of pharmacological inhibition of MERTK in mice. This involved the application of mass spectrometry imaging (MSI) to characterize the ocular spatial distribution of our highly selective MERTK inhibitor; AZ14145845, together with histopathology and transmission electron microscopy to characterize pathological and ultra-structural change in response to MERTK inhibition. In addition, we assessed the utility of a human retinal in vitro cell model to identify perturbation of phagocytosis post MERTK inhibition. We identified high localized total compound concentrations in the retinal pigment epithelium (RPE) and retinal lesions following 28 days of treatment with AZ14145845. These lesions were present in 4 of 8 treated animals, and were characterized by a thinning of the outer nuclear layer, loss of photoreceptors (PR) and accumulation of photoreceptor outer segments at the interface of the RPE and PRs. Furthermore, the lesions were very similar to that shown in the RCS rat and MERTK knock-out mouse, suggesting a MERTK-induced mechanism of PR cell death. This was further supported by the observation of reduced phagocytosis in the human retinal cell model following treatment with AZ14145845. Our study provides a viable, translational strategy to investigate the pre-clinical toxicity of MERTK inhibitors but is equally transferrable to novel chemotypes.

Highlights

MERTK is a receptor tyrosine kinase (RTKs) and part of the TAM family which includes TYRO3 and AXL
We describe a pre-clinical strategy to investigate the potential for ocular toxicity of a novel and selective MERTKi; which involved the application and integration of stateof-the-art technologies including multi-modal mass spectrometry imaging (MSI), histopathology and EM
We have shown changes at the histological, molecular and ultra-structural levels by combining light microscopy, mass spectrometry imaging and electron microscopy to enable a holistic understanding of ocular compound exposure and subsequent pathological consequences

Summary

Introduction

MERTK is a receptor tyrosine kinase (RTKs) and part of the TAM family which includes TYRO3 and AXL. The TAM family is involved in regulation of the innate immune response, for example, through MERTK-mediated efferocytosis of apoptotic cells by macrophages and of AXL-mediated dampening of TLR inflammatory responses (Graham et al 2014). TAM RTKs are overexpressed in a wide variety of human cancers with their induction in tumor cells conferring a survival benefit. A body of evidence has arisen that inhibition of their kinase activity can be used as a strategy to reverse the immunosuppressive tumor microenvironment, and slow cancer cell proliferation There is increased interest in their potential as novel oncolytic therapeutic targets

Methods

Results

Discussion

Conclusion