PD-L1 Tumor-infiltrating Lymphocytes Research Articles

Characterization of CD66b and its relationship between immune checkpoints and their synergistic impact in the prognosis of surgically resected lung adenocarcinoma

ObjectivesCD66b positive tumor-infiltrating neutrophils (TINs) are key immunity cells in the tumor microenvironment (TME). However, their relationship with clinicopathological features, immune checkpoints (ICs), and prognostic value remains undetermined in lung adenocarcinoma (LUAD). In this study, we aimed to characterize the infiltration by TINs and the prognostic significance in patients with surgically resected LUAD. Materials and methodsExpression of CD66b and ICs, including PD-L1, PD-1, CTLA4, LAG3, TIM3, TIGIT, VISTA, and BTLA, in both cancer cell and tumor-infiltrating lymphocytes (TILs) were estimated by immunohistochemistry in resected LUAD. The associations between CD66b expression and clinicopathological characteristics in patient prognoses were analyzed. We also verified results in another cohort from 85 patients with untreated LUAD and further analyzed the correlation between CD66b expression and EGFR and KRAS mutation status in addition to the rearrangement of the anaplastic lymphoma receptor tyrosine kinase gene (ALK). ResultsA total of 240 patients were included in this study. CD66b expression was observed in 87 (36.2%) samples. ICs including PD-L1, PD-1, CTLA4, LAG3, TIM3, TIGIT, VISTA, and BTLA were observed in percentages that ranged from 23.8% to 59.4%. Positive CD66b expression significantly correlated with smoking history (p = 0.029), pathological stage (p = 0.040), and the positive expression of LAG-3 (p < 0.001), PD-1 (p = 0.008), CTLA-4 (p = 0.013), TIM-3 (p = 0.025), TIGIT (p = 0.002), PD-L1 in TILs (p = 0.015), and PD-L1 in tumor cells (p = 0.010). CD66b positivity was significantly associated with worse recurrence-free survival (RFS) (hazard ratio, HR, 1.687; 95% confidence interval, CI, 1.058–2.690, p = 0.028) and overall survival (OS) (HR, 1.667; 95% CI, 1.097–2.534, p = 0.017). Subgroup analysis revealed that the CD66b+/LAG-3 + group had the worst RFS (5-year rate: 39.5%,) and OS (5-year rate: 53.7%,), while the CD66b−/LAG-3 − group had the best RFS (5-year rate: 65.6%) and OS (5-year rate: 78.8%). The p value in analysis of RFS and OS was 0.005 and 0.008, respectively. In the verification set, high expression of CD66b was also significantly correlated with the positive expression of LAG-3 (p < 0.001), PD-1 (p = 0.002), CTLA-4 (p = 0.034), TIM-3 (p = 0.049), PD-L1 in TILs (p = 0.003), and PD-L1 in tumor cells (p = 0.045). There was no correlation between CD66b expression and positive TIGIT expression (p = 0.077), EGFR mutation (p = 0.223), KRAS mutation (p = 0.151), and ALK fusion (p = 0.310). ConclusionCD66b had a relatively high positive expression rate and special clinicopathological features in patients with LUAD. CD66b + TINs were related to the expression of ICs and associated with poor prognoses in LUAD. A combination of CD66b and ICs, especially LAG-3 could further stratify patients into different groups with distinct prognoses.

Expression and Prognostic Value of Tumor-Infiltrating Lymphocytes and PD-L1 in Hepatocellular Carcinoma.

AimTo explore the difference in tumor-infiltrating lymphocytes (TILs) and programmed death-ligand (PD-L1) in primary hepatocellular carcinoma (HCC) and its adjacent tissues, and to evaluate their effect on HCC prognosis.MethodsLiver cancer and paracancerous tissue samples were collected from 72 patients who underwent radical hepatectomy between December 15, 2017 and January 9, 2019. Flow cytometry was used to detect the distribution of TILs and PD-L1, analyze the correlation between the expression of CD8/CD3 and PD-L1 and clinical-pathological parameters, and evaluate their effect on the prognosis of HCC patients.ResultsThe distribution proportion of CD3+T cells, CD4+T cells, and PD-L1 in liver cancer were significantly higher than in paracancerous tissues, while the distribution proportion of CD8+T cells was significantly lower (all P<0.05). In HCC, the distribution proportion of CD8+T cells was related to tumor size and stage, while the PD-L1 expression was related to the tumor stage only (all P < 0.05). Univariate analysis showed that tumor differentiation, TNM stage, expression of CD8/CD3, and PD-L1 in tumor tissue were related to disease-free survival (DFS)(P < 0.05); multivariate Cox regression analysis showed that tumor differentiation, TNM stage, CD8/CD3, and PD-L1 expression were independent influencing factors of postoperative DFS (P < 0.05). Kaplan–Meier survival curve analysis showed that the DFS of CD8/CD3 high expression group was significantly higher than that of the low expression group, and the DFS of PD-L1 low expression group was significantly higher than that of the high expression group (all P < 0.05).ConclusionThere are significant differences in the distribution of TILs and PD-L1 in HCC and paracancerous tissues. The expression of CD8/CD3 and PD-L1 in tumor-infiltrating lymphocytes in HCC may help evaluate the immunological indexes of prognosis after radical resection of HCC and to further the study of immunotherapy in patients with HCC.

BDNF expression in GISTs predicts poor prognosis when associated with PD-L1 positive tumor-infiltrating lymphocytes

ABSTRACT Substantial evidence indicates that brain-derived neurotrophic factor (BDNF) plays an important role in tumorigenesis, in addition to its primary role in neuronal activity. Gastrointestinal stromal tumors (GISTs), which are the most common mesenchymal neoplasms of the gastrointestinal tract, contain multiple types of tumor-infiltrating lymphocytes (TILs) that express relevant immune checkpoint proteins. However, no data have been reported on the role of BDNF in GISTs. This study aimed to investigate the expression pattern and prognostic value of BDNF in GIST patients with different degrees of risk, as well as the relationship between BDNF expression and immune checkpoints. Immunohistochemistry (IHC) demonstrated that higher BDNF expression was more likely to be present in high-risk patients and suggested a poor prognosis. A similar phenomenon was demonstrated in plasma. Even more interesting was that a positive correlation was present between BDNF and PD-L1+ expression on TILs. Moreover, high BDNF expression levels in combination with a high PD-L1+ TIL count predict extremely poor survival. The combination of BDNF expression and TIL PD-L1+ expression as a single biomarker was a powerful significant independent predictor of prognosis. Taken together, BDNF expression may serve as a significant prognostic factor, as the combination of BDNF expression and the PD-L1+ TIL subset led to superior prediction of GIST prognosis. Furthermore, our research coupled a neurotrophin with immunity, which provides novel evidence of neural and immune regulation in a clinical study of GIST.

Clinicopathologic features, tumor immune microenvironment and genomic landscape of Epstein-Barr virus-associated intrahepatic cholangiocarcinoma

Little is known about Epstein-Barr virus (EBV)-associated intrahepatic cholangiocarcinoma (EBVaICC) because of its rarity. We aimed to comprehensively investigate the clinicopathology, tumor immune microenvironment (TIME) and genomic landscape of this entity in southern China. We evaluated 303 intrahepatic cholangiocarcinomas (ICCs) using in situ hybridization for EBV. We compared clinicopathological parameters between EBVaICC and nonEBVaICC, and we analyzed EBV infection status, tumor-infiltrating lymphocytes (TILs) and genomic features of EBVaICC by immunohistochemistry, double staining, nested PCR, multiplex immunofluorescence staining, fluorescence in situ hybridization and whole-exome sequencing. EBVaICC accounted for 6.6% of ICCs and was associated with EBV latency type I infection and clonal EBV isolates. Patients with EBVaICC were more often female and younger, with solitary tumors, higher HBV infection rates and less frequent cirrhosis; the lymphoepithelioma-like (LEL) subtype was more common in EBVaICC. EBVaICC was associated with a significantly larger TIME component than nonEBVaICC. The LEL subtype of EBVaICC - associated with a significantly increased density and proportion of CD20+ B cells and CD8+ T cells - was associated with significantly higher 2-year survival rates than conventional EBVaICC and nonEBVaICC. Both PD-1 and PD-L1 in TILs, and PD-L1 in tumor cells, were overexpressed in EBVaICC. High PD-L1 expression in tumor cells and high CD8+ TIL densities were significantly more common in EBVaICC than in nonEBVaICC. Seven genes (MUC4, DNAH1, GLI2, LIPE, MYH7, RP11-766F14.2 and WDR36) were mutated in at least 3 patients. EBVaICC had a different mutational pattern to liver fluke-associated cholangiocarcinoma and HBV-associated ICC. EBVaICC, as a subset of ICC, has unique etiological, clinicopathological and genetic characteristics, with a significantly larger TIME component. Paradoxically, patients with EBVaICC could be candidates for immune checkpoint therapy. Epstein-Barr virus (EBV) is associated with a subtype of intrahepatic cholangiocarcinoma, with unique clinicopathological and genetic characteristics. The tumor immune microenvironment is also different in this tumor subtype and patients with EBV-associated intrahepatic cholangiocarcinoma may respond well to immune checkpoint inhibitors.

Abstract P4-10-30: PD-L1 expression in tumor infiltrating lymphocytes (TILs) inversely correlates with androgen receptor (AR) expression in HER2-positive breast cancer

Abstract Background: Human epidermal growth factor receptor 2 (HER2)-positive breast cancer accounts for 20-30% of breast carcinomas and is associated with a poor prognosis. While anti-HER2 targeted therapies have improved outcomes within this group, a subset of these tumors eventually develop resistance urgently requiring novel therapeutic options. In addition to classic HER2+ subtype, estrogen receptor (ER)-positive and HER2-positive tumors (luminal B) are another subtype of HER2 expressing breast carcinomas which could also benefit from additional therapies. Targeting the androgen receptor (AR) pathway and the PD-L1 axis may represent potential new therapeutic strategies in breast cancer. Our group previously reported that AR expression is seen in ¾ of HER2+ carcinomas and is more common in luminal B compared to classic HER2+ tumors. In this report, we evaluated expression of PD-L1 in tumor infiltrating lymphocytes (TILs) in both classic HER2 + and Luminal B subtypes of breast carcinomas and correlated the expression of PD-L1 with AR expression in these tumors. Methods: The study population consisted of 114 patients with HER2-positive invasive breast carcinoma diagnosed from 2009-2014 at Northwestern Memorial Hospital. Electronic medical records were reviewed for patient demographics (mean age 54, range 23-80). Pathologic tumor characteristics (histologic type, size, and grade) and tumor marker profile (ER and HER2) were evaluated. Tissue microarrays were constructed (3 cores from each case to account for tumor heterogeneity) for immunohistochemical evaluation of AR and PD-L1. AR was graded as positive when at least 1% of tumor cells showed nuclear immunoreactivity. For PD-L1, the extent of staining (0%=0, 1-10%=1, 11-50%=2, and 51-100%=3) and staining intensity (0, 1+, 2+, 3+) were assessed in TILs. Then a composite score (CS) was calculated by multiplying extent and intensity results (range 0-9; 0-3=negative, 4-9=positive). Results: Of the 114 studied breast carcinoma cases, 54 were classic HER2+ tumors (47%) and 60 were luminal B (53%) tumors. Approximately 69% of classic HER2+ and 87% of luminal B cases were positive for AR. Of interest, in classic HER2+ cases, the expression of PD-L1 in TILs was inversely correlated with the expression of AR, with 82% (14/17) of AR- tumors being positive for PD-L1 in TILs compared to only 46% (17/37) of AR+ tumors with PD-L1 expression in TILs (p&amp;lt;0.01). No association of expression of PD-L1 in TILs with AR expression was observed in luminal B cases (62% in AR+ versus 50% in AR- tumors; p=0.70). Moreover, in classic HER2+/AR- group, grade 3 tumors were seen almost exclusively in the PD-L1 expressing group (13 versus 3), in contrast to the equal number of grade 3 tumors in classic HER2+/AR+ group regardless of PD-L1 expression in TILs (14 versus 14). No association of the PD-L1 expression with age, race, and tumor histologic types was observed in classic HER2+/AR+ or classic HER2+/AR- cases. Conclusions: Our results demonstrated that the expression of PD-L1 in TILs was inversely correlated with the expression of AR in classic HER2 positive tumors but not in luminal B tumors. We also found that in classic HER2+/AR- group, grade 3 tumors were seen almost exclusively in the PD-L1 expression group. Our findings add to the body of knowledge of the molecular mechanisms that drive the different molecular subtypes of breast carcinomas and raise the interesting possibility that new treatment strategies such as targeting the PD-L1 axis may show more benefit against the AR negative subtype of the classic HER2 positive carcinomas. Citation Format: Tiansheng Shen, Jennifer L. Pincus, Kalliopi P. Siziopikou. PD-L1 expression in tumor infiltrating lymphocytes (TILs) inversely correlates with androgen receptor (AR) expression in HER2-positive breast cancer [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P4-10-30.

Immunogenomic landscape of hepatocellular carcinoma with immune cell stroma and EBV-positive tumor-infiltrating lymphocytes

The immunogenomic characteristics of hepatocellular carcinomas (HCCs) with immune cell stroma (HCC-IS), defined histologically, have not been clarified. We investigated the clinical and molecular features of HCC-IS and the prognostic impact of Epstein-Barr virus (EBV) infection. We evaluated 219 patients with conventional HCC (C-HCC) and 47 with HCC-IS using in situ hybridization for EBV, immunohistochemistry, multiplex immunofluorescence staining, and whole exome and transcriptome sequencing. Human leukocyte antigen types were also extracted from the sequencing data. Genomic and prognostic parameters were compared between HCC-IS and C-HCC. CD8 T cell infiltration was more frequent in HCC-IS than C-HCC (mean fraction/sample, 22.6% vs. 8.9%, false discovery rate q <0.001), as was EBV positivity in CD20-positive tumor-infiltrating lymphocytes (TILs) (74.5% vs. 4.6%, p <0.001). CTNNB1 mutations were not identified in any HCC-IS, while they were present in 24.1% of C-HCC (p = 0.016). Inhibitory and stimulatory immune modulators were expressed at similar levels in HCC-IS and EBV-positive C-HCC. Global hypermethylation, and expression of PD-1 and PD-L1 in TILs, and PD-L1 in tumors, were also associated with HCC-IS (p <0.001), whereas human leukocyte antigen type did not differ according to HCC type or EBV positivity. HCC-IS was an independent factor for favorable recurrence-free survival (adjusted hazard ratio [aHR] 0.23; p = 0.002). However, a subgroup of tumors with a high density of EBV-positive TILs had poorer recurrence-free (aHR 25.48; p <0.001) and overall (aHR 9.6; p = 0.003) survival, and significant enrichment of CD8 T cell exhaustion signatures (q = 0.0296). HCC-IS is a distinct HCC subtype associated with a good prognosis and frequent EBV-positive TILs. However, paradoxically, a high density of EBV-positive TILs in tumors is associated with inferior prognostic outcomes. Patients with HCC-IS could be candidates for immunotherapy. Hepatocellular carcinomas with histologic evidence of abundant immune cell infiltration are characterized by frequent activation of Epstein-Barr virus in tumor-infiltrating lymphocytes and less aggressive clinical behavior. However, a high density of Epstein-Barr virus-positive tumor-infiltrating lymphocytes is associated with inferior prognostic outcomes, possibly as a result of immune escape due to significant CD8 T cell exhaustion.

Abstract P5-12-09: The ≥5% cut-off reveals tumor PD-L1 positivity as potential selection biomarker for patient enrollment into the trials evaluating anti-PD-1 or anti-PD-L1 agents in neoadjuvant treatment of triple negative breast cancer

Abstract Background: Durable responses of triple negative breast cancer (TNBC) to pembrolizumab (anti-PD-1) or atezolizumab (anti-PD-L1) have been reported in the metastatic setting. Moreover, it is currently being hypothesized that immune checkpoint inhibitors might be more effective in the neoadjuvant setting, due to better preserved anti-tumor immunity in early TN disease. However, biomarkers predictive of response to anti-PD-1 or anti-PD-L1 agents, as well as biomarker-based strategies for testing those drugs in the neoadjuvant setting are still lacking. We evaluated PD-L1 protein expression and the composition of tumor-infiltrating lymphocytes (TILs) in untreated TNBC, in order to get a better insight into the TNBC sub-population(s) which would be suitable for neoadjuvant anti-PD-1 or anti-PD-L1 therapy evaluation. Methods: TNBC patients consecutively treated at the Jean Perrin Comprehensive Cancer Centre (Clermont-Ferrand, France), from 01/01/2010 to 12/31/2014, were included. FFPE tumor tissues were assessed for PD-L1 expression by immunohistochemical (IHC) laboratory-developed test (clone 28-8, Abcam), in tumor cells (tPD-L1) and in TILs. Positivity cut-offs evaluated were ≥1%, ≥5% and ≥10%. The amount CD8+, CD4+, FoxP3+ or PD-1+ TILs was determined by counting those cells, detected by IHC methods, within 5 consecutive HPFs (x400), from tumor invasive front toward tumor center. Clinical disease stage was determined using the TNM system. Results: One hundred and two TNBCs were assessed. There were 28.4%, 23.5% or 16.7% tPD-L1-positive cases (cs), for cut-offs ≥1%, ≥5% or ≥10%, respectively. Similarly, 32.4%, 15.7% or 5.9% of cs were positive for PD-L1 in TILs, using the same cut-offs. With ≥5% as cut-off, positivity for tPD-L1 significantly correlated with the amount of CD8+ (p=0.023), FoxP3+ (p=0.0036) or PD-1+ TILs (p=0.043). The same cut-off, applied to TILs, revealed significant correlations between PD-L1 positivity and the amount of each CD8+, CD4+ or PD-1+ TILs (p=0.025, 0.039 and 0.0042, respectively). Interestingly, when the ≥5% cut-off was used, tumors of T2 size were most frequently tPD-L1+ (11/41 cs, 26.8%), compared with the T1 (3/35 cs, 8.6%) and T3+T4 (3/18 cases, 16.7%) (p=0.04). With regards to TILs, with the ≥5% cut-off, the PD-L1+ cases belonged exclusively to the T1+T2 group (15/76), whereas the T3+T4 group was PD-L1-negative (0/18 cs). Other two cut-offs revealed only occasional correlations. Conclusion: This single-center, real-world TNBC cohort contained a high number of smaller tumors (T1-T2). The IHC-based PD-L1 assessment, with ≥5% as the positivity cut-off, revealed that approximately 1/4 of TNBC could be candidates for neoadjuvant anti-PD-1/anti-PD-L1 approaches. Combined with the amount of CD8+ and PD-1+ TILs, tumor PD-L1 positivity might make an easy-to-use composite biomarker for the 1st-level patient selection for PD-1 or PD-L1 inhibitors in neoadjuvant TNBC treatment. The 2nd level could exploit, for example, the assessment of mutation burden in tumors with low tPD-L1 or amount of CD8+ or PD-1+ TILs. Such tumors might be more frequent among larger TNBC (T3-T4). Citation Format: Finck W, Passildas J, Poirier C, Kwiatkowski F, Abrial C, Durando X, Penault-Llorca F, Radosevic-Robin N. The ≥5% cut-off reveals tumor PD-L1 positivity as potential selection biomarker for patient enrollment into the trials evaluating anti-PD-1 or anti-PD-L1 agents in neoadjuvant treatment of triple negative breast cancer [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P5-12-09.

Circulating Lymphocytes, PD-L1 Expression on Tumor-infiltrating Lymphocytes, and Survival of Colorectal Cancer Patients with Different Mismatch Repair Gene Status.

Clinical outcomes of checkpoint blockade immunotherapy on colorectal cancer (CRC) are influenced by mismatch repair (MMR) gene status, which is associated with distinct tumor immune infiltrates and systemic inflammatory response status. However, the prognostic value of PD-L1 expression and the systemic inflammatory response for patients with MMR deficiency has not been fully investigated. In this study, we examined the association of systemic inflammatory markers, PD-1/PD-L1 pathway expression, microsatellite instability (MSI) status, and clinicopathological characteristics of CRC with patient survival between MMR-deficient (dMMR) group (N=168) and MMR-proficient (pMMR) group (N=169). We found a large proportion of dMMR CRC patients displayed increased level of systemic inflammatory markers such as C-reactive protein, Neutrophil/Lymphocyte Ratio (NLR), Glasgow Prognostic Score (GPS), and low expression of PD-L1 in tumor stroma. Several systemic inflammatory markers were associated with AJCC stage only in dMMR patients. Similarly, Tumor infiltrating lymphocyte (TIL) PD-L1 or stroma PD-L1 expression was associated with AJCC stage only in dMMR patients. Circulating serum lymphocytes and TIL PD-L1 expression are both independent prognosis predictors for CRC patients. Overall, we found that dMMR CRC displayed a comprehensively distinct tumor immune microenvironment and systemic inflammatory response makers. PD-L1 expression at different location has different impacts on CRC patient survival, and the TIL PD-L1 expression might be a potential predictor for dMMR CRC patient response to anti-PD-1 therapy.

Case-control study of PD-1, PD-L1, and B7-H3 expression in human immunodeficiency virus infected (HIV+) and uninfected patients with head and neck cancers.

e23194 Background: HIV+patients are typically excluded from immunotherapy trials but there are limited data on co-signaling molecule expression in these patients, including in head and neck squamous cell carcinoma (HNSCC). Methods: A case control study was conducted to evaluate tumor tissue sections for PD-L1, PD-1, and B7-H3 expression in HIV+HNSCC patients (n = 12) and HNSCC controls (n = 12). Cases and controls were matched for age at diagnosis, race, gender, TNM stage, and primary site. Smoking status, HIV RNA viral load (VL), and CD4 count were also recorded. Immunostained tumor sections were analyzed for percent of tumor cells expressing PD-L1 (PD-L1%) and B7-H3(B7-H3%) (Abcam), and percent of tumor infiltrating lymphocytes (TIL) expressing PD-1 (TIL-PD-1%) and PD-L1 (TIL-PD-L1%) (Abcam). Statistical analysis used the non-parametric Mann-Whitney test, chi-square test and Spearman’s rank correlation, Rs. Results: The 12 HIV+ HNSCC cases were predominantly male (67%), black race (67%) and cigarette smokers (100%), with a median age of 50.4 years, median viral load (VL) of 52399 copies/mL, median CD4 count (CD4) of 236cells/uL, predominantly locally advanced (75% Stage III/IVa/IVb), and oropharyngeal primary site (42%). Defining positive expression as &gt; 5%, 42% of HIV+ HNSCC patients tumors were positive for PD-L1, 100% for B7-H3, and 92% had TILs that expressed PD-1 and PD-L1. HIV+ patients had significantly higher B7-H3% (Median 60% vs. 20%, p = 0.03) and TIL-PD-L1% (Median 15% vs. 10%, p = 0.045) compared to controls. There were no differences in PD-L1% or TIL-PD-1%. In HIV+ cases, increased VL correlated with increased TIL-PD-1% (Rs = 0.73, p = 0.011) and increased CD4 count correlated with increased tumor PD-L1% (Rs = 0.62, p = 0.04). There were no other significant correlations between CD4 count or VL and co-signaling molecule expression. Conclusions: HIV+ HNSCC patients had significantly higher tumor B7-H3 and TIL PD-L1 expression, with similar tumor PD-L1 and TIL PD-1 expression, compared to HIV negative HNSCC control patients. These findings support inclusion of HIV+ HNSCC patients in immunotherapy trials with checkpoint inhibitors.

Case-control study of PD-1, PD-L1 and B7-H3 expression in lung cancer (CA) patients (pts) with and without human immunodeficiency virus (HIV) infection.

11606 Background: PD-1, PD-L1 and B7-H3 are co-signaling molecules involved in CA immunology. There are limited data on expression of these molecules in HIV-infected (HIV+) lung CA pts and these pts are routinely excluded from immunotherapy trials. Methods: We reviewed archived lung CA tissue samples from HIV+ cases (n = 13) and HIV-uninfected controls (n = 13) from 2001-2015. Cases and controls were matched by histology and stage. Baseline demographics were collected for all pts. CD4 count and HIV RNA viral load (VL) were collected for HIV+ pts. Immunostained tumor sections were analyzed for percent of tumor cells expressing PD-L1 and B7-H3 (Abcam), and percent of tumor-infiltrating lymphocytes (TIL) expressing PD-1 and PD-L1 (Abcam). Positive expression was defined as &gt; 5%. Statistical analysis was performed using the non-parametric Mann-Whitney test and the chi-square test. Proportions are specified as percentage with 95% confidence limits in parentheses. Results: Lung CA HIV+ case pts were predominantly male (62%), black race (100%), adenocarcinoma histology (77%), stage 4 disease (62%), and had a median age of 48 years. Of case pts with available data, mean CD4 count was 307 (range 37-617) and mean HIV VL was 29,400 (range 0-100,000). PD-L1 expression on tumor cells was positive in 23% (8%, 50%) of cases and 46% (23%, 71%) of controls. B7-H3 expression on tumor cells was positive in 92% (67%, 99%) of cases and 69% (42%, 87%) of controls. PD-1 expression on TIL was positive in 69% (42%, 87%) of cases and 54% (29%, 77%) of controls. PD-L1 expression on TIL was positive in 31% (13%, 58%) of cases and 69% (42%, 87%) of controls (p = 0.05). B7-H3 percent expression on tumor cells was significantly higher in cases vs controls (median 90% vs 20%, p = 0.005), but there were no significant differences in percent expression of PD-L1 on tumor cells, PD-1 on TIL or PD-L1 on TIL. Conclusions: HIV+ lung CA pts had significantly higher B7-H3 tumor percent expression compared to HIV-uninfected controls, with similar rates of PD-L1 tumor percent expression, PD-1 TIL percent expression and PD-L1 TIL percent expression. These results support inclusion of HIV+ lung CA pts in future immunotherapy trials.

Temporal and spatial discordance of programmed cell death-ligand 1 expression and lymphocyte tumor infiltration between paired primary lesions and brain metastases in lung cancer

The dynamics of PD-L1 expression may limit its use as a tissue-based predictive biomarker. We sought to expand our understanding of the dynamics of PD-L1 expression and tumor-infiltrating lymphocytes (TILs) in patients with lung cancer-related brain metastases. Paired primary lung cancers and brain metastases were identified and assessed for PD-L1 and CD3 expression by immunohistochemistry. Lesions with 5% or greater PD-L1 expression were considered positive. Agreement statistics and the χ(2) or Fisher's exact test were used for analysis. We analyzed 146 paired lesions from 73 cases. There was disagreement of tumor cell PD-L1 expression in 10 cases (14%, κ = 0.71), and disagreement of TIL PD-L1 expression in 19 cases (26%, κ = 0.38). Most paired lesions with discordant tumor cell expression of PD-L1 were obtained 6 or more months apart. When specimens were categorized using a proposed tumor microenvironment categorization scheme based on PD-L1 expression and TILs, there were significant changes in the classifications because many of the brain metastases lacked either PD-L1 expression, tumor lymphocyte infiltration or both even when they were present in the primary lung cancer specimens (P = 0.009). We identified that there are significant differences between the tumor microenvironment of paired primary lung cancers and brain metastases. When physicians decide to treat patients with lung cancer with a PD-1 or PD-L1 inhibitor, they must do so in the context of the spatial and temporal heterogeneity of the tumor microenvironment.

PD-L1 expression in the Merkel cell carcinoma microenvironment: association with inflammation, Merkel cell polyomavirus and overall survival.

Merkel cell carcinoma (MCC) is a lethal, virus-associated cancer that lacks effective therapies for advanced disease. Agents blocking the PD-1/PD-L1 pathway have demonstrated objective, durable tumor regressions in patients with advanced solid malignancies and efficacy has been linked to PD-L1 expression in the tumor microenvironment. To investigate whether MCC might be a target for PD-1/PD-L1 blockade, we examined MCC PD-L1 expression, its association with tumor-infiltrating lymphocytes (TILs), Merkel cell polyomavirus (MCPyV), and overall survival. Sixty-seven MCC specimens from 49 patients were assessed with immunohistochemistry for PD-L1 expression by tumor cells and TILs, and immune infiltrates were characterized phenotypically. Tumor cell and TIL PD-L1 expression were observed in 49% and 55% of patients, respectively. In specimens with PD-L1(+) tumor cells, 97% (28/29) demonstrated a geographic association with immune infiltrates. Among specimens with moderate-severe TIL intensities, 100% (29/29) demonstrated PD-L1 expression by tumor cells. Significant associations were also observed between the presence of MCPyV DNA, a brisk inflammatory response, and tumor cell PD-L1 expression: MCPyV(-) tumor cells were uniformly PD-L1(-). Taken together, these findings suggest that a local tumor-specific and potentially MCPyV-specific immune response drives tumor PD-L1 expression, similar to previous observations in melanoma and head and neck squamous cell carcinomas. In multivariate analyses, PD-L1(-) MCCs were independently associated with worse overall survival (hazard ratio 3.12; 95% CI, 1.28-7.61; p=0.012). These findings suggest that an endogenous immune response promotes PD-L1 expression in the MCC microenvironment when MCPyV is present, and provide a rationale for investigating therapies blocking PD-1/PD-L1 for patients with MCC.