Kim, In-Ho

Abstract Background: The Tumor Immune microenvironment (TME) plays an important role in tumor development, progression, and treatment response. Recent studies show that spatial heterogeneity of Tumor-Infiltrating Lymphocytes (TILs), consisting of lymphocytic cell populations, is an important histopathologic feature and a powerful prognostic and predictive marker correlating with cancer progression and treatment response. Detecting and quantifying TIL density in Whole Slide Images (WSIs) is, however, labor-intensive, and it is difficult to assess differences in the composition of immune cells. Therefore, it remains a challenge to establish the analysis of TILs in the complex TME. We present an automated spatial image analysis of TILs on gastric cancer to address cancer immunotherapy. Method: 1,031WSIs of hematoxylin and eosin-stained (H&E) formalin-fixed paraffin-embedded (FFPE) sections from four patient cohorts: TCGA-STAD (329), Yonsei-1 (620 patients), Yonsei-2 (45), and St. Mary (37), were analyzed. Two treatment groups in response were classified as CR + PR, while another two treatment groups in non-response were classified as PD + SD. H&E stained WSIs were normalized using the Reinhard normalization method at 20X magnification for reducing the color variability of the stained tissue images. We used U-Net architecture to predict the normalized H&E stained WSIs. Three U-Net models were trained for predicting tumor, TILs, and stroma, using Adam optimizer, 1e-4 learning rate, a binary loss function, L2 regularization, and 50 epochs. The normalized H&E stained WSIs were predicted using the trained U-Net models. Spatial analysis of TILs was conducted on the predicted WSIs. Regional quantification of TILs was performed by two immune phenotype groups: immune-immersed (TILs enriched in tumor cells) and immune-excluded (TILs not enriched in tumor cells but in stromal cells). Kaplan-Meier with a log-rank test was used to evaluate the survival differences between the groups. Results: In TCGA-STAD, the proportion of immune-immersed composition is significantly different from the immune-excluded (p-value 0.05). In Yonsei-1, the immune-excluded composition is significantly different among the three risk groups (p-value 3e-05). In the Yonsei-2 and St. Mary cohorts, including anti-PD1 treated gastric cancer patients, the immune-immersed composition is significantly different between two response groups (p-value 0.007) and represented the diagnostic accuracy (AUROC: 0.78) on CR + PR groups. Conclusion: These findings suggest that the proposed TIL quantification in the complex TME is strongly associated with patient prognosis and anti-PD1 treatment response in gastric cancer. Citation Format: Sanghoon Lee, Sunho Park, Jae-Ho Cheong, Sam C. Wang, Matthew R. Porembka, In-Ho Kim, Sung Hak Lee, Tae Hyun Hwang. Spatial image analysis of tumor-infiltrating lymphocytes on gastric cancer to predict anti-PD1 inhibitor response [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1360.

Abstract Background: While immune checkpoint inhibitors (ICI) extend survival for patients suffering from a variety of cancers, most gastric cancer (GC) patients do not respond to ICI. There is a critical need to identify novel biomarkers that guide the use of ICI or strategies to improve outcomes for GC patients. We aim to gain a deeper understanding of the GC tumor microenvironment (TME) to refine the use of ICI using spatial transcriptomes from ICI-treated gastric tumors. Method: We collected 12 pre-ICI treatment gastric tumor samples from five responders and seven nonresponders and performed spatial profiling to investigate spatial heterogeneity, functions, and phenotypes of tumor and neighboring stromal and immune cells to identify features associated with ICI response. We used pan-cytokeratin (pan-CK) to mark epithelial cells, CD45 for immune cells, and smooth muscle actin (SMA) for fibroblasts. We generated gene expression profiles of each cell type within regions of interest (ROI) with a NanoString GeoMx Cancer Transcriptome Atlas panel, comprised of 1,800 genes. A total of 58 ROIs were analyzed from responders and 62 ROIs from nonresponders. Results: We measured the spatial heterogeneity of each cell type in ROIs within and across tumors. We found the tumor, immune, and fibroblasts from nonresponders are transcriptionally more heterogeneous, compared to those from responders. This might indicate a higher degree of intratumor transcriptional heterogeneity is linked to ICI resistance. We found that nonresponse was associated with higher expression of VEGFA and VEGFB in tumor cells and higher ACTA2 in fibroblasts, not in other cell types. These findings are intriguing when considering our other finding that increased endothelial cell and fibroblast proportions in the TME are associated with ICI nonresponse. We also found that the TME of nonresponders had fewer M1 and more M2 macrophages, and there were more T cells that overexpressed gene programs consistent with dysfunction and exhaustion. These findings indicate that ICI-resistant TMEs are densely surrounded by immune-suppressive cellular components which might reduce ICI efficacy and result in ICI resistance. Finally, we performed cell-cell interaction and identified ligand-receptor interactions that are stronger in nonresponders, including CXCL12-CXCR4, CD74-MIF, and LGALS9-LRP1 ligand-receptor interactions. The predicted ligand-receptor interactions are present in cell-type-specific manners, indicating the functional role of each cell type in facilitating ICI response. Conclusion: We investigated how tumor and neighboring stromal and immune cells interact within GC TME to mediate ICI response, which demonstrate that spatial analysis could be used to identify novel biomarkers that predict response to ICI, and new therapies to enhance ICI efficacy in GC patients. Citation Format: Sunho Park, Changjing Hong, Jae-Ho Cheong, Sam C. Wang, Matthew R. Porembka, In-Ho Kim, Sung Hak Lee, Tae Hyun Hwang. Spatial architecture and cellular interactions of tumor immune microenvironment to discover biomarkers and predict immune checkpoint inhibitor response in gastric cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2262.

292 Background: Despite recent approval of anti-PD-1 antibodies as 1L therapy in HER2(-) advanced GEA, benefit remains modest and limited largely to PD-L1(+) patients (pts), primarily those with combined positive scores (CPS) ≥5. Thus novel therapeutic approaches are needed for this pt population. DKN-01 is a targeted anti-DKK1 mAb which has demonstrated improved clinical outcomes in pts with elevated tumoral DKK1 expression, a subset of pts with more aggressive disease and shorter overall survival. Methods: DisTinGuish (NCT04363801) is a Phase 2a single arm 2-part trial; Part A investigated DKN-01 (D) + tislelizumab (TS) + CAPOX as 1L therapy for pts with advanced HER2(-) GEA regardless of DKK1 status; Part B investigated two dosing cohorts of D (300 mg and 600 mg) + TS as 2L therapy for DKK1-high advanced GEA pts. Primary objective was to examine safety and tolerability and secondary objectives evaluated multiple efficacy endpoints including overall response rate (ORR) in a modified intent to treat (mITT) population (>1 dose D). Results: Forty-nine pts enrolled between 01 Sept 2020 and 15 Sept 2021; 25 pts in Part A and 24 pts in Part B (D-300 mg). Key clinicopathologic features and efficacy outcomes are shown in Table. The most common D-related AEs were low grade (G1/2) fatigue, nausea, and diarrhea. Nine pts had D-related ≥G3 toxicities, elevated AST/ALT, elevated alkaline phosphatase, hypophosphatemia, hyponatremia, lymphopenia, neutropenia, diarrhea, vomiting, fatigue all occurring in 1 pt and pulmonary embolism in 2 pts (one G5 event). No new safety signals were observed in Part A or B1. Duration of response (DoR), median PFS and median OS have not been reached for Part A. Last pt enrolled in Part B1 on 15 Sept 2021. Conclusions: The combination of D/TS + CAPOX represents a well-tolerated, active 1L combination, particularly for DKK1-high patients consistent with the proposed mechanism of action. Activity appears to be independent of PD-L1 status. Part B1 is aligned with biomarker enrichment and efficacy and biomarker data will be presented along with updated Part A efficacy data. Clinical trial information: NCT04363801. [Table: see text]