Abstract 1681: Simultaneous measurement and significance of PD-1, LAG-3 and TIM-3 expression in human solid tumors
Authors: Morgado Micaela , Datar Ila , Wang Jun , Sanmamed Miguel F. , McEachern Kristen , Jenkins David , Chen Lieping , Carvajal-Hausdorf Daniel , Rimm David L. , Herbst Roy S. , Schalper Kurt A. 
Published: August 2018

Introduction: T-cells recognizing tumor antigens can express diverse immune inhibitory receptors such as PD-1, LAG-3 and TIM-3. Blockade of PD-1 induces prominent clinical benefit in patients with melanoma, lung, bladder and gastric carcinomas. However, the relative expression and significance of immune inhibitory receptors across different tumors is unknown.

Methods: Using multiplexed quantitative immunofluorescence (QIF) we measured the levels of CD3 (rabbit polyclonal, Dako), PD-1 (EH33, CST), LAG-3 (17B4, Abcam) and TIM-3 (D5D5R, CST) in &gt;1,300 human malignancies including 119 triple negative breast carcinomas (TNBC), 225 ovarian carcinomas, 382 melanomas, 259 colorectal tumors, 229 bladder urothelial carcinomas and 130 gastric malignancies. The cohorts included FFPE tumor tissues and were studied using tissue microarrays (TMAs). The targets were measured in all cells of the preparation using fluorescence co-localization with DAPI and specifically in CD3-positive T-lymphocytes. Associations between the markers across tumors and with survival were studied.

Results: PD-1, LAG-3, and TIM-3 were expressed predominantly in T-cells and showed a positive association with each other. The highest levels were detected in gastric adenocarcinomas followed by urothelial bladder tumors, ovarian malignancies, melanoma, TNBC and colorectal carcinomas. Gastric and bladder tumors showed comparably high levels of all three markers and were significantly higher than the other tumor types (P&lt;0.001). TNBC and colorectal carcinomas showed comparably lower expression. The levels of PD-1, LAG-3 and TIM-3 were positively associated with CD3+ T-cell infiltration in all tumors except in gastric cancer. Melanoma showed the highest CD3 levels and gastric tumors had the lowest. Elevated T-cell PD-1 was significantly associated with longer overall survival in melanoma and colorectal cancer; and elevated LAG-3 was associated with better survival only in melanoma. The levels of TIM-3 were not associated with survival.

Conclusion: PD-1, LAG-3 and TIM-3 are differentially expressed in human solid tumors and show limited prognostic value. Gastric and bladder carcinomas show the highest levels of the targets, while TNBC and colorectal cancer show the lowest. Despite having the highest T-cell infiltration, melanomas show intermediate levels of the immune inhibitory receptors. These data demonstrate the differences in the immune composition of human solid tumors and could be used for optimal design and interpretation of clinical trials.

Citation Format: Micaela Morgado, Ila Datar, Jun Wang, Miguel F. Sanmamed, Kristen McEachern, David Jenkins, Lieping Chen, Daniel Carvajal-Hausdorf, David L. Rimm, Roy S. Herbst, Kurt A. Schalper. Simultaneous measurement and significance of PD-1, LAG-3 and TIM-3 expression in human solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1681.