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Zhiyuan Yu, Ghasem Yazdanpanah, Tye S Thompson, Kevin Burgess, Cintia S De Paiva; Small molecule agonists of Tyrosine Kinase receptors improve dry eye disease. Invest. Ophthalmol. Vis. Sci. 2022;63(7):1979 – A0309.
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Nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin3 (NT-3) bind to tyrosine kinase (Trk) receptors, TrkA, TrkB, and TrkC, respectively. This study investigated the efficacy of novel molecule agonists of Trk receptors in an in vivo model of dry eye disease.
Small molecule agonists for TrkC (C1) or pan-Trk (pan) were synthesized based on structural formula and β-turns. C57BL/6J mice were subjected to desiccating stress (DS) and received bilateral eye drops of C1, pan or vehicle (2x/day). The corneal barrier function was assessed by uptake of a fluorescent dye (n=11-18/ group). The conjunctival goblet cell (GC) density was measured in paraffin-embedded, PAS-stained sections (n=10-14/group). Corneal epithelial lysates were collected for either western blot or RNA extraction. Extracted total RNAs were used for NanoString analysis. Immunofluorescent staining was performed on wholemount corneas using anti-A20 and anti-PTGER4 antibodies.
Compared to naïve mice, desiccated mice treated with vehicle showed corneal barrier disruption (55±11 vs 97.8±38.4 gray levels, P<0.01) and goblet cell loss (50±11 vs 37±11 goblet cells/mm, P<0.01). Compared to treatment with vehicle, pan and C1- treated mice showed an improvement in corneal barrier function (98±38 vs 59±17 and 72±18 gray levels, respectively, P<0.01), and higher conjunctival GC density (37±11 vs 46±10 and 52±13 GC/mm, respectively, P<0.05). NanoString results revealed upregulation of specific mRNA transcripts (Ptger4, Tnfaip3 [encoding A20], Il1a and Ptger4, Tlr3, Osal1, respectively) in Pan and C1-treated corneas compared to vehicle-treated corneas (all P-adj<0.05). Western blot results showed that treatment with pan and C1 decreased vehicle-induced NFkB nuclear translocation after DS for one day and increased PTGER4 and A20 protein levels after 5 days of DS in corneal epithelium lysates. These results were confirmed by immunostaining using antibodies for A20 and PTGER4 in wholemount corneas.
Small molecule agonists of Trk receptors improve dry eye disease by decreasing NFkB activation and increasing protein expression of anti-inflammatory molecules A20 and PTGER4. Further studies are needed to investigate whether pan and C1 analogs modulate other pathways involved in dry eye.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.
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