PD-1 Expression In Tumor Infiltrating Lymphocytes Research Articles

Immune biomarker expression in the tumor microenvironment in Chinese patients with esophageal squamous cell carcinoma was explored.

e15542 Background: Esophageal squamous cell carcinoma (ESCC) is one of the most common malignancies in China, of which the standard treatment of advanced esophageal cancer is chemotherapy followed by surgery. Recently, immunotherapy has shown great potential in ESCC treatment. The aim of this study was to explore the landscape of immune-related gene expression in the tumor microenvironment (TIME) and further explore the relationship between immune biomarkers and therapeutic effect. Methods: We retrospectively analyzed PDL1(clone sp142)/PD1 expression in a cohort of Chinese patients (male = 92, female = 18; median age = 64, range from 36 to 84) with ESCC. Patient samples were detected in Genecast. Co. from February 2017 to November 2018. Another 6 patients (male = 4, female = 2) who accepted anti-PD1/PD-L1 antibody treatment were analyzed. PD-L1/PD-1 expression and the incidence of CD8, CD57, CD68, CD163 TILs were immunohistochemically. Results: In tumor cells, more than 32% (36/110) samples of ESCC patients was stained with PDL1 (≥1%), while52.8% (21/36) of these patients highly expressed PDL1 (≥10%, 10%-98%). In tumor infiltrating lymphocytes (TILs), 31.82% (35/110) patients expressed PDL1 (≥1%) and 1.82% (2/110) highly expressed PDL1 (≥10%, 10%-50%). For PD1 expression in TILs, 17.27% (19/110) samples were positively stained with PD1 antibody (≥1%). In a subset of ESCC patients who received immune checkpoint blockade therapy (including 2 responders and 4 non-responders), the median percentage of CD163, CD68, PDL1, CD8, PD1 and CD57 were 1.49% (0.44%-9.98%), 0.40% (0.16%-19.08%), 3.74% (0%-45%), 9.52% (4.20%-13.79%), 4.51% (1.37%-33.43%) and 0.68% (0.08%-1.74%), respectively in tumor area and 4.14% (1.85%-15.99%), 1.62% (0.21%-11.47%), 4.87% (0.33%-18.97%), 7.74% (5.88%-14.35%), 3.93% (1.92%-20.54%) and 0.92% (0.27%-2.18%), respectively in stroma area. When compared with non-responders, samples from the responders showed higher CD8 expression in tumor area. Conclusions: Percentage of PDL1/PD1 expression is relatively high in Chinese patients with ESCC. High levels of CD8 positive TILs might be beneficial to patients with ESCC. Clinical investigations with large sample sizes are warranted to prove our findings.

Evaluation of programmed cell death protein 1 (PD-1) expression as a prognostic biomarker in patients with clear cell renal cell carcinoma

ABSTRACT Programmed cell death protein 1 (PD-1) immune checkpoint inhibitors have shown activity in patients with advanced renal cell carcinoma (RCC). However, the role of PD-1 expression in tumor-infiltrating lymphocytes (TILs) as a biomarker for poor outcome is not clear. In this study, we evaluated the prognostic value of TIL PD-1 expression in patients with clear cell RCC (ccRCC). 82 patients who underwent nephrectomy for localized or metastatic ccRCC and followed up for at least four years were searched from our database and retrospectively enrolled. Their fixed primary tumor specimens were stained with anti-PD-1 (NAT105). The specimens were classified as negative or positive for PD-1 expression, and the positive specimens were further scored in 10% increments. 37 (45.12%) patients were negative (<1% stained), 26 (31.71%) patients were low (<10 and 10%), and 19 (23.17%) patients were high (20–50%) for PD-1 expression. The prognostic value of TIL PD-1 expression was evaluated by univariate Cox proportional hazards regression on overall and recurrence-free survivals. Higher TIL PD-1 expression was not associated with increased risk of death (P = 0.336) or with increased risk of recurrence (P = 0.572). Higher primary tumor stage was associated with increased risk of recurrence (P = 0.003), and higher Fuhrman nuclear grade was associated with increased risk of death (P <0.001) and with increased risk of recurrence (P <0.001). Our study shows that TIL PD-1 expression by immunohistochemistry (IHC) does not correlate with poor clinical outcome in patients with ccRCC and is inferior to established prognosticating tools.

Abstract 3849: HuPBL humanized models as a tool to evaluate the efficacy and immune regulation of immune checkpoint inhibitors

Abstract After the success of PD-1, PD-L1 and CTLA-4 antibodies, a number of immune-checkpoint inhibitors are already in advanced stages. Meanwhile, tumor infiltrating lymphocytes (TIL) are well recognized as features of tumor progression and keys to effective cancer immunotherapy. Therefore, there is an urgent need to establish an effective pre-clinical platform for the evaluation of therapeutical outcomes in the human immune-system and the understanding of TILs. Either traditional syngeneic models or human target knockin models have their drawbacks: syngeneic models disable the development of antibodies that do not recognize mouse receptors; neither of them can achieve a good clinical prediction due to their inherent property as a murine system. To get a reconstitution of the human immune system in murine models, we have established several customized Human peripheral blood lymphocytes (hu-PBL) humanized models in NOG (NOD/Shi-scid/IL-2Rγnull) mice either systemically or focalized in tumors. Those models were validated by immune-checkpoint inhibitors for efficacy and by conducting TIL analysis with the support of sophisticated Flow Cytometry-Based Approach. Here we report that the engrafted human cells can expand and infiltrate in tumors in response to xenogenic stimulation with functional Th and Tc detected. The cell lineages are mainly CD3+ T lymphocytes (90%) and to lesser extent NK/NKT cells; the redistribution in tumors is unique not only compared to splenic and peripheral blood T cells, but also varies among different cancer cell lines when the same donor was applied. In addition, cell surface expression of immune checkpoint targets showed unique redistributions. We further investigated the correlation between the efficacy and the immune-regulation in these models: Models with more dense infiltration at the early phase usually grew more slowly than its NON-PBMC control, which also are prone to be better responders to immunotherapy due to the higher Effector cell: Target cell ratio (E:T ratio). When compared the efficacy of 2 different hPD-1 test antibodies in A375 systemic humanized model, the one with better efficacy also achieved stronger inhibition on PD-1 expression in TILs. Treatment with the clinically used Opdivo showed not only different responses in HCC827, A375 and A431 xenograft models but also unique profiles in TILs subsets and related immune-checkpoint expression. These models can be used for the purpose of pre-clinical immune-checkpoint inhibitor and bi-specific antibody screening under human background, as well as for further investigations regarding distinct functions of TILs subsets and target expression. Citation Format: Xuzhen Tang, Xianzhi Zhai, Hui Qi, Qin Hong, Qiyao Zhang, Lei Jin, Shaoyu Yan, Zhuozhi Wang, Yong Zheng, Qingyang Gu, Norman Zhang, Jing Li, Qunsheng Ji. HuPBL humanized models as a tool to evaluate the efficacy and immune regulation of immune checkpoint inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3849. doi:10.1158/1538-7445.AM2017-3849

Abstract PD6-07: PD-L1 is highly expressed in tumor infiltrating lymphocytes in pregnancy associated breast cancer

Abstract Background: Pregnancy associated breast cancer (PABC), diagnosed during or after gestation, is typically triple negative, and is associated with a poor prognosis. Those diagnosed within two years have an even worse outcome. We previously assessed the immune microenvironment of invasive breast carcinomas in young women and reported that tumor infiltrating lymphocytes (TILs) were more prominent in PABC. Programmed cell death protein 1 (PD-1) is upregulated following activation of lymphocytes, while programmed death ligand 1 (PD-L1) is one of the primary ligands that it interacts with to inhibit T-cell activation and proliferation. PD-L1 may also be constitutively expressed on tumor cells as a result of oncogenic signaling or epithelial-mesenchymal transition. Emerging evidence suggests that the effect of the local immune system, particularly the interactions between PD-1 and PD-L1, is also key in breast cancer progression and in breast tumor responses to chemotherapy and targeted therapy. In this study, we assessed expression of PD-1 and PD-L1 in both TILs and tumor cells in PABC and in age-/stage-/grade-matched nulliparous women, and correlated their expression with clinicopathologic characteristics in this aggressive type of breast carcinomas. Design: 21 patients diagnosed with PABC within two years of pregnancy (mean age=35.7, range=26-48) and 15 matched controls (mean age=37.5, range=29-51) were evaluated. Slides were reviewed and pathologic tumor characteristics, including TILs, were noted. Immunohistochemical stains for PD-1 and PD-L1 were performed. Extent (1=1-25% positive tumor cells, 2=26-50%, 3=51-75%, 4=76-100%) and intensity (1=weak, 2=moderate or 3=strong) of staining were assessed. A composite score (CS) was calculated by multiplying the extent by intensity (range=0-12; weak=1-3; moderate=4-8 and strong=9-12). Results: The mean CS for PD-L1 in TILs was significantly higher in PABC (5.86) compared to controls (3.07), p=0.03. Further, strong expression of PD-L1 in TILs was only observed in PABC (9/21, 42.9%); none of the controls had strong PD-L1, p=0.01. The high expression of PD-L1 in PABC TILs was independent of tumor grade, hormone receptor and HER2 status, and other histologic features including lymph node metastasis. Expression of PD-1 in TILs was similar in both PABC and controls (mean CS 6.81 and 5.36, respectively). Immunoreactivity in the tumor cells themselves was rare with only two PABC and four control cases expressing PD-1 and PD-L1. Conclusion: 1. TILs in PABC have significantly higher PD-L1 expression. 2. Strong expression of PD-L1 in TILs was only observed in PABC. 3. High PD-L1 expression in TILs was independent of the tumor characteristics in this series. 4. PD-1 is expressed similarly in TILs in both PABC and controls. 5. Rare cases may have PD-1 and PD-L1 expression in the tumor cells themselves. The results of our study showing significant expression of PD-L1 in PABC TILs add to the understanding of the role of the microenvironment in breast cancer progression. These complex interactions between tumors cells and the local immune system may predict response to therapy and investigation into the role of immune based therapies is under way in these aggressive breast carcinomas that affect young women. Citation Format: Blanco Jr LZ, Pincus JL, Siziopikou KP. PD-L1 is highly expressed in tumor infiltrating lymphocytes in pregnancy associated breast cancer [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr PD6-07.

PUB115 Clinical Significance of PD-L1 Status and the Expression of PD-1 in Tumor-Infiltrating Lymphocytes (TILs) in Lung Cancer

PD-L1 status in lung cancer may be a predictive biomarker for the use of PD1/PD-L1 inhibitors. Nonetheless, it is still unclear which immunohistochemistry platform for PD-L1 assessment should be implemented for optimal prognostic/predictive information. We investigated whether the PD-L1 status in early-stage lung cancer is associated with specific clinical features or survival, as well as the PD1 expression in TILs. IHC was performed on FFPE surgical tumors, arrayed on tissue microarrays with duplicate cores. PD-L1 status was assessed with two ABs: Ventana (SP124) (PD-L1V) and Cell Signaling (E1L3N) (PD-L1CS). PD1 expression in TILs was determined with the AB Ventana (NAT105). All patients (n=482) had early stage T1/T2-N0 and underwent curative surgery between 1987 and 2002. PD-L1 membranous staining was scored as positive if >5% of tumor cells. PD1 was considered as positive if >10% in TILs. Tabled 1PD-L1(V)PD-L1(CS)PD1All cases (including carcinoids)7%18%50%Histology Sq Non-Sqp=0.0043 12% 4%p<0.0001 30% 12%56% 49%pStage IA IB5% 10%p=0.017 15% 24%51% 53%Grading G1 G2 G3p=0.004 1% 6% 12%p=0.0002 7% 18% 27%p=0.001 41% 45% 61% Open table in a new tab Patient’s median age (IQR) 68 years (59-73), gender male/female 53%/47%, smoking status never/former/current/missing 14%/26%/49%/11%, histology squamous-cell carcinoma (Sq)/Non-squamous (Non-Sq)-NSCLC/carcinoid 37%/55%/7%, p-stage IA/IB 52%/48%. A table shows the association between biomarker status (PD-L1(V), PD-L1(CS) and PD1 )and tumor features (including Pearson’s Chi-Square p-values <0.05). All carcinoids were negative for PD-L1, except for one. The degree of agreement between the two PD-L1 Abs was moderate (Kappa=0.47). Median OS was 76 months. No statistically significant association between PD-L1 status and OS was observed. PD1 expression in TILs was associated with worse survival in Sq tumors. Median OS for cases with PD1 positive vs negative TILs was 54 vs 79 months, respectively (log-rank p=0.08; Wilcoxon p=0.02). In this study we observed a higher expression of PD-L1 using the CS antibody with a moderate agreement between the two antibodies. Sq tumors showed a significantly higher expression of PD-L1 compared with non-Sq tumors. PD-1 expression in TILs was associated with worse survival in Sq tumors.

PD1 and PDL1 expression in primary central nervous system diffuse large B-cell lymphoma are frequent and expression of PD1 predicts poor survival.

Primary central nervous system diffuse large B-cell lymphoma (PCNS-DLBCL) is a rare and aggressive type of diffuse large B-cell lymphoma (DLBCL) whit poorly understood pathogenesis. Finding biomarkers associated with patient survival may be important for understanding its physiopathology and to develop new therapeutic approaches. We investigated 32 PCNS-DLBCL from immunocompetent patients for BCL2, CMYC, LMO2, and P53 expression and for cytogenetic aberrations of BCL2, BCL6, and MYC genes, all known for their prognostic value in systemic DLBCL (s-DLBCL). We analyzed PD1 and PDL1 protein expression in both tumor infiltrating lymphocytes (TILs) and tumor cells. Finally, we searched for correlation between biological data and clinical course. The PCNS-DLBCL expressed BCL2, CMYC, LMO2, and P53 at similar frequency than s-DLBCL but without significant prognostic on survival. None cases harbored aberrations involving BCL2 and MYC gene whereas BCL6 abnormalities were present in 20.7% of cases but without value on survival. Expression of PD1 in TILs and PDL1 in tumor cells was observed at higher rates than in s-DLBCL (58% and 37%, respectively). The PD1 expression in TILs correlated with PDL1 expression in tumor cells (P=.001). Presence of PD1 positive TILs was associated with poorer overall survival (P=.011). Patients with PDL1 overexpression tended to better response to chemotherapy (P=.23). In conclusion PCNS-DLBCL pathogenesis differs from s-DLBCL without prognostic value of the phenotypic and cytogenetic parameters known for their pejorative impact in the latter. The PD1/PDL1 pathway plays a strong role in PCNS-DLBCL and represents an attractive target for this aggressive lymphoma.

A novel iNKT-mediated vaccine maneuver overcomes PD-1-mediated T cell exhaustion in cancer (165.13)

Abstract Our immune system plays an important role in preventing cancer. However, tumor-derived immuno-suppression hinders effective anti-tumor immunity. T cell dysfunction or exhaustion in tumor-bearing hosts is one such mechanism. Numerous studies have identified PD-1 and Tim-3 as markers of exhausted T cells, and antibodies blocking PD-1 and Tim-3 have been shown to partially restore T cell function. We have found that both Tim-3 and PD-1 are expressed on CD8+ tumor-infiltrating lymphocytes (TILs) in an animal model of B cell lymphoma and that these cells represent the predominant fraction of T cells infiltrating tumors. We find that iNKT stimulation using the ligand, alpha-galactosylceramide, combined with TLR9 stimulation using the ligand, CpG, can down-regulate Tim-3 and PD-1 expression in TILs, restore TIL function as measured by proliferation and production of TNF and IFNg in response to tumor, and induce long-lasting therapeutic immune responses in mice with large advanced (&amp;gt;1cm2) tumors.