W. Amjad1, S. Bonini1, K. Montone2, K. Nathanson3, H. Wachtel1,4 1Hospital Of The University Of Pennsylvania, Department Of Surgery, Philadelphia, PA, USA 2Hospital Of The University Of Pennsylvania, Department Of Pathology And Laboratory Medicine, Philadelphia, PA, USA 3Hospital Of The University Of Pennsylvania, Department Of Genetics, Philadelphia, PA, USA 4University Of Pennsylvania, Perelman School Of Medicine, Philadelphia, PA, USA
Introduction: Pheochromocytoma and paraganglioma (PCC/PGL) are rare neuroendocrine tumors that are associated with pathogenic loss-of-function variants of succinate dehydrogenase subunits (SDHx). Therapeutic strategies include surgical resection as the primary therapy. Radionuclide therapies such as MIBG have since emerged as systemic therapy; however, no targeted therapies exist for PCC/PGL. Homologous repair deficiency in SDHx-associated PCC/PGL has been studied, opening avenues to investigate tumors with poly-ADP-ribose (PARP) expression that confer susceptibility to therapeutic drugs that target DNA damage repair, such as PARP inhibitors. This study aims to assess the correlation between biomarkers of DNA damage repair and KX-1 uptake as an in vitro assay for PARP binding.
Methods: Contiguous sections of frozen human PCC/PGL tissue were processed using a cryotome. Frozen sections were processed for immunohistochemical (IHC) stains: PARP-1, Ki-67, SDHB, CD45, pATM, and gamma-H2AX. Digital autoradiography using phosphorimaging was used for quantitative analysis and producing digital images. KX-1 uptake was quantitatively measured using ImageJ. IHC stained slides were scanned and digitized prior to analysis in QuPath for percent positive and H-score. Univariable linear regression was performed to assess the association between KX-1 specific binding and IHC, assessed by H-score or percent positive cells. Subset analysis was performed by SDHx status (Wildtype vs. SDHx), tissue type (PCC vs. PGL), and disease progression (metastatic vs. nonmetastatic).
Results: A total of 45 samples were included in the study, comprising 6 normal adrenals and 39 tumors (12 PGL and 27 PCC). Of the 45 samples, 9 had an SDHx mutation (SDHA: 1, SDHB: 5, SDHC: 1, SDHD: 2). On univariable linear regression, there were no DNA damage repair biomarkers significantly associated with KX-1 expression by both H-score and percent positive cells. When subset by SDHx status, CD45, a marker of immune cells, was significantly associated (p=0.018) with KX-1 in patients with SDHx pathogenic variants, but not in wildtype patients (p=0.316). When stratified for disease progression, SDHB staining by IHC was negatively correlated with KX-1 specific binding in patients with nonmetastatic disease (p=0.013) but not in patients with metastatic disease (p=0.939).
Conclusion: We found that rates of PARP binding measured by KX-1 assay differed based on SDHx germline status and metastatic disease status. Rates of KX-1 expression appear to correlate with CD45, a marker of immune cells, and expression of SDHB. These data suggest that immune cell infiltrates may impact rates of PARP binding and therefore may potentially modulate responses to PARP inhibitor therapies.
