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bioRxiv · 10.1101/2022.10.31.513798

Automated analysis of PD1 and PDL1 in lymph nodes and the microenvironment of transmissible tumors in Tasmanian devils

Russell, G. G.·Palmieri, C.·Darby, J.·Morris, G. P.·Fountain-Jones, N. M.·Pye, R.·Flies, A. S.
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Abstract

The wild Tasmanian devil (Sarchophilus harrisii) population has suffered a devastating decline due to two clonal transmissible cancers. Devil facial tumor 1 (DFT1) was first observed in 1996, followed by a second genetically distinct transmissible tumor, devil facial tumor 2 (DFT2), in 2014. DFT1/2 frequently metastasize, with lymph nodes being common metastatic sites. Downregulation of MHC-I by DFT1 cells is a primary means of evading allograft immunity aimed at polymorphic MHC-I proteins. DFT2 cells constitutively express MHC-I, and MHC-I is upregulated on DFT1/2 cells by interferon gamma, suggesting other immune evasion mechanisms may contribute to overcoming allograft and anti-tumor immunity. Human clinical trials have demonstrated PD1/PDL1 blockade effectively treats patients showing increased expression of PD1 in tumor draining lymph nodes, and PDL1 on peritumoral immune cells and tumor cells. The effects of DFT1/2 on systemic immunity remain largely uncharacterized. This study applied the open-access software QuPath to develop a semiautomated pipeline for whole slide analysis of stained tissue sections to quantify PD1/PDL1 expression in devil lymph nodes. The QuPath protocol provided strong correlations to manual counting. PD-1 expression was approximately 10-fold higher than PD-L1 expression in lymph nodes and was primarily expressed in germinal centers, whereas PD-L1 expression was more widely distributed throughout the lymph nodes. The density of PD1 positive cells was increased in lymph nodes containing DFT2 metastases, compared to DFT1. This suggests PD1/PDL1 exploitation may contribute to the poorly immunogenic nature of transmissible tumors in some devils and could be targeted in therapeutic or prophylactic treatments.

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Russell, G. G., Palmieri, C., Darby, J., Morris, G. P., Fountain-Jones, N. M., Pye, R., Flies, A. S.. 2022-11-01. Automated analysis of PD1 and PDL1 in lymph nodes and the microenvironment of transmissible tumors in Tasmanian devils. https://doi.org/10.1101/2022.10.31.513798

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