High-throughput screen reveals that atypical antipsychotics promote continual efferocytosis by antagonizing dopamine signaling and promoting vitamin A-dependent Arginase 1 upregulation

Programmed cell removal, or efferocytosis, has been proposed as a method to dispose of unwanted cells that accumulate and promote diseases such as cancer, infection, and atherosclerosis. Several pro-efferocytic therapies are currently in development, but toxicity related to the off-target clearance of healthy tissue has led to the premature termination of multiple first-generation clinical programs. To identify new pro-efferocytic therapies with established risk profiles, we conducted a high-throughput screen of about 3000 FDA-approved drugs and compounds. This allowed us to identify the atypical antipsychotic drug, thiothixene, as a promising candidate for clinical translation. Leveraging thiothixene's known inhibitory effects on a wide range of neurotransmitters and catecholamines, we found that dopamine has a potent inhibitory effect on efferocytosis, which thiothixene could partially offset. Mechanistic studies revealed that thiothixene's pro-phagocytic effects were not mediated by alterations in so-called 'eat me' molecules or efferocytic receptors, but instead occurred due to the induction of a process known as continual efferocytosis. RNA-sequencing revealed that these effects occurred via alterations in the retinol binding receptor, Stra6L, which enhanced vitamin A-dependent signaling and drove upregulation of the continual efferocytosis mediator, Arginase 1. Taken together, this unbiased screen identified the unanticipated anti-efferocytic properties of dopamine, while highlighting the translational potential of a generic, FDA-approved anti-psychotic drug which has been in use for more than 50 years. This therapy may represent a novel approach for promoting continual efferocytosis and the removal of diseased tissue that otherwise could promote a range of clinical disorders. Overall design: BMDMs were prepared from three mice. At day4 of differentiation, 7x10^5 macrophages were seeded into 6-well plates, and cells were treated with 2µM thiothixene or DMSO for 24 hours at day 6. Drug-treated macrophages were washed with PBS once and cells were lysed with 0.5 ml TRIzol™ Reagent. Total RNA was extracted according to the protocol from manufacturer and qualified on a NanoDrop™ OneC Microvolume UV-Vis Spectrophotometer (Thermo Scientific™ 840274200). Three biological replicates per treatment group with a total of 200 ng RNA per sample were sent to Novogene Co. (Sacramento, CA, USA) for sample quality control, library preparation, and sequencing.