High-accuracy Detection of PD-L1 3'-UTR Disruption by Immunohistochemistry and Fluorescence in Situ Hybridization on Formalin-fixed Paraffin-embedded Sections

Programmed death-ligand 1 (PD-L1/CD274) structural variation (SV) disrupting the 3'-untranslated region has been highlighted as being associated with PD-L1 overexpression. In the present study, we evaluated lymphoma tissue samples to investigate the applicability of immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) for detecting the PD-L1 SV involving the 3'-untranslated region. In total, 1052 lymphoma samples were screened using IHC, and 99 IHC screening-positive samples were evaluated with FISH (non-Hodgkin lymphoma [NHL, n=58] and Hodgkin lymphoma [HL, n=41]). Of these, 92 samples showed strong PD-L1 expression with 2 PD-L1 antibodies (E1J2J and SP142) (concordant PD-L1 IHC), whereas 7 samples showed strong PD-L1 expression only with E1J2J (discordant PD-L1 IHC). Abnormal FISH findings for PD-L1 were detected in all evaluated samples (51 NHLs and 41 HLs). A structural abnormality pattern was observed in 17 of the 51 evaluated NHL samples (33%). In contrast, all 41 HL samples showed a copy number abnormality pattern, with 1 exhibiting a structural abnormality pattern. Target-capture sequencing of the PD-L1 gene was performed on 73 of the 99 IHC screening-positive samples, comprising 41 NHLs and 32 HLs. PD-L1 SVs were detected in 16 (39%) of the 41 NHL samples and in only one of the 32 HL samples (3%). Samples exhibiting discordant PD-L1 IHC and/or FISH structural abnormality patterns were shown to harbor PD-L1 SV by target-capture sequencing, with positive and negative predictive values of 94% and 96%, respectively. Our approach is an alternative to target-capture sequencing for evaluating PD-L1 gene abnormalities.