PD-L1 Score Research Articles

Abstract 4955: Impact of PD-1, PD-L1 and EBI3 on prognosis in a cohort of localized high grade undifferentiated pleomorphic sarcoma patients

Abstract Introduction: Studies of soft-tissue sarcomas (STS) and therapeutic outcomes are limited by their rarity and heterogeneity. Despite intensive treatment, including chemotherapy, surgery and radiation therapy, 30% of patients develop recurrent disease, and the outcome of the patients with recurrent or metastatic sarcomas remains poor. The SARC028 phase II, multicenter trial of pembrolizumab reported promising activity in selected histologic subtypes of advanced STS, including undifferentiated pleomorphic sarcoma (UPS). On the other hand, multicentre phase II trial (PEMBROSARC) found that PD-1 inhibition has limited activity in selected STS (UPS and LMS and GIST). The application of immune checkpoint blockade in sarcoma is in its infancy and we do not yet understand which patients will benefit from these therapies. Methods: We explored the degree of PD-L1, PD-1, and CD8 expression in tumor tissue and microenvironment and their clinical impact in a retrospective analysis of 50 UPS sarcoma samples, at diagnosis. PD-L1 status was assessed on whole sections of fixed tissue. We also studied expression by tumors cells of two heterodimeric cytokines IL-27 (EBI3 / p28) and IL-35 (EBI3 / p35), known for inducing the expression of immune checkpoints. Their expression was investigated by analyzing the expression of their shared subunit EBI3 by immunohistochemistry. Patients presented with grade 3 FNCLCC, localized, sarcoma of the extremities, with a high risk of local and distant recurrence and were treated in a relatively homogeneous way (surgery followed by adjuvant radiation therapy and chemotherapy). Results: The median overall survival was 63.4 months after a median follow-up of 42.0 months (range, 3.7- 108.3 months). The PD-L1 score on tumor cells (E1L3N, Cell Signaling) was negative in 94% of cases, with only 1 that had more than 10% PD-L1-positive tumor cells. Three semi-quantitative scores were performed on the immune cells: PD-L1 (same antibody): score 0 (62%), 1 (34%), 2 (4%); PD-1 (PD-1 NAT, Abcam): score 0 (52%), 1 (32%), 2 (16%) and CD8 + (C8/144B, Dako): score 0 (4%), 1 (36%), 2 (58%), NE (2%). PD-1 expression was associated with CD 8 infiltration (chi2 p=0.047). The EBI3 score on tumor cells was positive (>30% positive cells) in 6% of cases and correlated with worse overall survival (p=0.04) and progression-free survival (p<0.01), whereas PD1 (expressed by T cells), PD-L1 (expressed by infiltrating immune cells or tumor cells) and CD8 infiltration lacked prognostic significance. We are now integrating data from Affymetrix microarray gene expression and whole exome to investigate the molecular differences on tumors. Conclusion: This study suggests that targeting immune checkpoints downstream of IL-27/IL-35, different from PD1 may benefit to some UPS patients. Note: This abstract was not presented at the meeting. Citation Format: Pascaline Boudou-Rouquette, Anne Jouinot, Virginie Audard, Franck Letourneur, Béatrice Parfait, François Goldwasser, Benoit Terris, Pierre Laurent-Puig, Jérôme Alexandre, Karen Leroy, Eric Pasmant, Odile Devergne, Frederique Larousserie. Impact of PD-1, PD-L1 and EBI3 on prognosis in a cohort of localized high grade undifferentiated pleomorphic sarcoma patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4955.

A Minimum Of 100 Tumor Cells in a Single Biopsy Sample Is Required to Assess Programmed Cell Death Ligand 1 Expression in Predicting Patient Response to Nivolumab Treatment in Nonsquamous Non–Small Cell Lung Carcinoma

IntroductionProgrammed death ligand 1 (PD-L1) expression is a predictive biomarker for patient response to nivolumab in nonsquamous NSCLC. However, the number of biopsy samples and tumor cells (TCs) required to assess PD-L1 expression remains unclear. MethodsA total of 222 biopsy samples from 80 patients with nonsquamous NSCLC treated with nivolumab were collected. Number of TCs and PD-L1 score were compared among the sample containing the largest number of TCs (Max-TC), the sample containing the smallest number of TCs (Min-TC), and the total samples from each patient. The impact of the number of samples and TCs on the prediction of patient response to nivolumab with use of PD-L1 scores was evaluated. ResultsThere was a mismatch in PD-L1 scores less than 1% and those of at least 1% between Max-TC and the total samples in one patient (1%) and between Max-TC and Min-TC in six patients (8%). The optimal number of TCs to match PD-L1 expression less than 1% versus at least 1% between Max-TC and Min-TC was 100 (sensitivity = 0.676 and 1 – specificity = 0.333). PD-L1 expression of at least 1% in Min-TCs containing at least 100 TCs was associated with longer progression-free survival (median 7.6 versus 1.8 months [p < 0.01]) and overall survival (median not reached versus 9.9 months [p = 0.04]) compared with PD-L1 expression less than 1%. However, there were no differences in progression-free survival (median 3.9 versus 2.3 months [p = 0.37]) or overall survival (median 9.7 versus 7.6 months [p = 0.60]) between PD-L1 expression of at least 1% and PD-L1expression less than 1% in Min-TCs containing fewer than 100 TCs. ConclusionSingle biopsy samples containing at least 100 TCs are required to evaluate PD-L1 expression for predicting patient response to nivolumab.

Phase I study of durvalumab and tremelimumab together with radiotherapy for the adjuvant treatment of intermediate-risk head and neck squamous cell carcinoma.

TPS73 Background: When surgery is used as the primary therapeutic modality for locally-advanced HNSCC, radiation therapy (XRT) is frequently recommended post-operatively due to high rates of recurrence. In high-risk patients; i.e., those with positive surgical margins or extra-nodal extension (ENE) on surgical pathology, the addition of chemotherapy to post-operative XRT (cCRT) provides a survival benefit. In contrast, there is no standard indication for adjuvant systemic therapy in intermediate risk HNSCC—defined in this study as having a T3/T4 primary tumor, perineural or lymphovascular space invasion, and/or lymph node metastasis without ENE. The anti-programmed cell death-ligand 1 (PD-L1) antibody durvalumab (D) has a manageable safety profile and encouraging clinical activity studies across multiple tumor types. Combining D with the anti-cytotoxic T-lymphocyte-associated antigen-4 (anti-CTLA-4) antibody tremelimumab (T) may amplify antitumor T-cell responses and provide synergistic activity. D+T showed antitumor activity and manageable tolerability in advanced NSCLC. Furthermore in preclinical studies, D+T combined with XRT decreased regulatory T-cells and myeloid derived suppressor cells and increased CD8 T-cell recruitment to the tumor microenvironment. Methods: This two-part phase I study will include a safety run-in period using a 3+3 dose de-escalation design based on dose limiting toxicities (DLT). In Cohort 1, D (q3 weeks) +T (q3 weeks) will be given on cycle 1 followed by concurrent XRT for Cycles 2-4. Cycle 5 and 6 will include D only. If DLT’s are met, Cohort -1 will include D (q3 weeks) and T (q6 weeks). If DLT's are found in Cohort -1, Cohort -2 would include only D. A dose expansion cohort (N=24) will follow based on the safest combination in the safety run-in. Translational studies include correlation of pre-treatment immune characteristics including PD-L1 score, TMB, and IFN-gamma signature with disease free survival. DNA and RNA sequencing of pre-treatment tissue and at treatment failure/recurrence will attempt to identify genetic and molecular determinants of IO treatment resistance. (NCT02000947). Clinical trial information: NCT03529422.

Upregulation of PD-L1 and PD-L2 in neck node metastases of head and neck squamous cell carcinoma.

This study aimed on evaluating the expression levels of the immune checkpoint proteins, PD-L1 and PD-L2, in tissue specimens of 175 oral squamous cell carcinomas (OSCC) and 33 corresponding lymph node metastases (LNM). Tissue microarrays of primary OSCC and matched LNM were investigated via immunohistochemistry regarding PD-L1/PD-L2 expression. Results were compared for primary tumors and corresponding metastases and correlated with clinicopathological data. PD-L1 expression in cancer cells correlated significantly with tumor size, and the presence of regional metastases (P < 0.01). PD-L1 expression was significantly higher in metastases as compared to primary OSCC (P < 0.05). Overall survival was significantly worse in patients with higher levels of PD-L1 and PD-L2 score (P < 0.05). This is the first publication to compare PD-L1 and PD-L2 expression in primary OSCC and matched LNM. The results indicate that anti-PD-1 therapy may be of therapeutic use even in early stage OSCC to prevent further progression.

MHC Class 1 and PDL-1 Status of Primary Tumor and Lymph Node Metastatic Tumor Tissue in Gastric Cancers.

The prognosis of metastatic gastric cancer is poor. Despite the use of VEGF-, EGFR-, and HER2-targeting agents, prognosis is still poor in advanced gastric cancer. Although cancer immunotherapy responds well in some patients, clinical use is limited due to unanswered patients. For this reason, it is necessary to know the characteristics of primary and metastatic cancer cells for patient selection for immunotherapy and additional criteria are required. MHC-1 downregulation is most frequently observed in the tumor escape mechanism of cancer cells from the immune system. MHC-1 downregulation with increased PDL-1 expression of cancer cells has an important role in immune escape. MHC-1 downregulation and PDL-1 expression have been shown in many types of cancers. However, there is no study on the status of MHC-1 and PDL-1 in primary and metastatic tumor tissue. In this study, MHC-1 and PDL-1 score in primary and metastatic tumor cells was evaluated in 43 gastric cancer patients with lymph node metastasis. According to our results, the primary tumor PDL-1 score was correlated with the number of metastatic lymph nodes (r = 0.258; p = 0.024) and primary tumor size (r = 0.341; p = 0.045). A similar correlation was found between the primary tumor PDL-1 score and the metastatic tumor PDL-1 score (r = 0.213; p = 0.015). In our study, MHC-1 was found to be higher in primary tumors than metastatic tumors, although not statistically significant (p = 0.054). The results of our study showed high MHC-1 and low PDL-1 expression in primary tumors and low MHC-1 and high PDL-1 expression in metastatic tumors. These results reveal different biological characteristics of primary and metastatic tumor cells.

New 6-factor prognostic model for patients (pts) with advanced urothelial carcinoma (UC) receiving post-platinum atezolizumab.

413 Background: Prognostic factors for overall survival (OS) have been identified in pts receiving post-platinum chemotherapy for advanced UC, but it is unknown whether these factors and/or others optimally predict OS for pts treated with PD1/PD-L1 inhibitor therapy. Methods: Pt level data from two UC salvage trials evaluating atezolizumab were used: IMvigor210 (n = 310) for training and PCD4989g (n = 95) for validation. Univariable and multivariable Cox regression analyses were performed to evaluate the association of the prognostic factors recognized in the chemotherapy setting (ECOG performance status [ECOG-PS], liver metastasis (LM), anemia, treatment-free interval, albumin), neutrophil-lymphocyte ratio (NLR), eosinophil count, platelet count (PLT), site of primary/metastases, stage at diagnosis, smoking, LDH, prior therapies and immune cell PD-L1 status by IHC with OS. Clinical factors were dichotomous and lab values normalized by logarithmic transformation as needed. Stepwise selection was employed to propose an optimal model using the training dataset; pts were then categorized by number of risk factors. Concordance, discrimination (c-statistic) and calibration were assessed in the validation dataset using bootstrap analyses. Results: The factors included in the optimal prognostic model for OS were: ECOG-PS 1 vs. 0 (HR 1.64 [95% CI: 1.20, 2.24], p = 0.002), LM (1.45 [1.08, 1.94], p = 0.014), PLT (1.73 [1.14, 2.61], p = 0.010), NLR (1.84 [1.45, 2.34], p &lt; 0.001), LDH (1.54 [1.19, 1.99], p = &lt; 0.001) and anemia (HR = 1.60 [1.17, 2.21] p = 0.004). The c-statistic was 0.690 (95% CI = 0.649-0.715) and 0.759 (0.694-0.795) in the training and validation datasets, respectively. 1-year OS of pts in the training and the validation cohorts were similar. PD-L1 score was statistically significant when adjusted for the optimal model, but did not improve clinical interpretability (c-statistic = 0.698). Conclusions: A new validated 6-factor prognostic model for OS including ECOG-PS, LM, PLT, NLR, LDH and anemia is proposed in the setting of post-platinum atezolizumab for advanced UC. Applicability of the model to other PD1/PD-L1 inhibitors and PD-L1 IHC assays warrant investigation.

Abstract GS2-06: Phase Ib/II study evaluating safety and efficacy of pembrolizumab and trastuzumab in patients with trastuzumab-resistant HER2-positive metastatic breast cancer: Results from the PANACEA (IBCSG 45-13/BIG 4-13/KEYNOTE-014) study

Abstract Background Preclinical and clinical data suggest that HER2-positive (HER2+) breast cancer (BC) will be amendable to immunotherapeutic approaches. We evaluated pembrolizumab with trastuzumab in patients (pts) with trastuzumab-resistant HER2+, PD-L1 positive (PD-L1pos), unresectable loco-regional or metastatic BC and a parallel cohort of pts with HER2+, PD-L1 negative (PD-L1neg) BC during the phase II study. Methods: Pts with advanced BC and progression on prior trastuzumab-based therapies, ECOG 0-1, and a metastatic tumor biopsy in the last year were eligible. HER2 positivity and quantity of tumor-infiltrating lymphocytes (TILs) on H&amp;E slide were centrally evaluated. PD-L1 score was assessed by Merck central lab. Tumor imaging was performed at weeks 12, 18, 24 and every 12 weeks, thereafter. Primary endpoints were safety of the combination (phase Ib) and objective response rate (ORR) per RECIST 1.1 (phase II). Secondary endpoints were PFS, duration of response, and OS. Phase Ib was a 3+3 dose-escalation of 2 pembrolizumab doses (2mg/kg, 10mg/kg) Q3W. In phase II, pts received pembrolizumab 200mg Q3W for 24 months or until disease progression. Clinically stable pts with progression were allowed to continue pembrolizumab until confirmation on subsequent assessment. Pts with isolated CNS progression were also allowed to continue pembrolizumab after local treatment. Planned total enrollment was 61 pts. For the phase II PD-L1pos cohort, a Simon two-stage design (N=40; proceed if ≥2/17 respond) was used which had 85% power to compare ORR of 7% vs. 22% (1-sided α=0.05). For the PD-L1neg cohort, a single-stage design with 15 pts had &amp;gt;95% power to compare ORR of 1% vs. 20% (1-sided α=0.14). Clinicaltrials.gov: NCT02129556. Results: 6 pts enrolled in phase Ib between April and July 2015; no DLTs were observed. The PD-L1pos cohort enrolled 40 pts between August 2015 and September 2016. The PD-L1neg cohort enrolled May 2016 to April 2017, stopping after 12 pts due to low rate of PD-L1 negativity, maintaining &amp;gt;90% power to detect the target difference in ORR. PD-L1 testing labs changed in April 2016. Prior to this time, QualTek PD-L1 positive was defined as ≥1% on tumor or TILs. Using the Dako 22C3 antibody, positive was defined as tumor PD-L1 combined positive score (CPS)≥1%. 146 pts were screened to enroll 58 pts. Of screened pts, median stromal TILs was 1% (mean: 4.8%, SD: 9.1%, range: 0 to 60%; n=127); 52% of pts were PD-L1pos, with higher positivity rates while using the Dako assay compared with Qualtek (65% vs. 43%, p=0.009). Median TILs of pts in the PD-L1pos cohort was 2% (mean: 8.1%, SD: 11.2%, range: 0 to 40%) and 0% (mean: 1.2%, SD: 2.2%, range: 0 to 5%) in the PD-L1neg cohort. Of enrolled pts, median age was 51yrs (range: 28-72), 69% had visceral metastases. 29% of pts received prior pertuzumab, 72% had prior T-DM1, 40% prior lapatinib. 38% of pts were ER-positive, 62% were ER-negative. Median TILs in enrolled ER pos and ER neg pts were 1.5% and 2.0%, respectively. PD-L1 positivity rates were also not significantly different by ER status (p=0.5). Final safety data and efficacy results will be presented at the meeting. Citation Format: Loi S, Giobbe-Hurder A, Gombos A, Bachelot T, Hui R, Curigliano G, Campone M, Biganzoli L, Bonnefoi H, Jerusalem G, Bartsch R, Rabaglio-Poretti M, Kammler R, Maibach R, Smyth MJ, Di Leo A, Colleoni M, Viale G, Regan MM, Andre F. Phase Ib/II study evaluating safety and efficacy of pembrolizumab and trastuzumab in patients with trastuzumab-resistant HER2-positive metastatic breast cancer: Results from the PANACEA (IBCSG 45-13/BIG 4-13/KEYNOTE-014) study [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr GS2-06.

Genomic and immune landscape of metastatic melanoma (MM) treated with pembrolizumab (PEM).

184 Background: Predictive biomarkers for PEM in MM is an active area of research. Methods: INSPIRE (NCT02644369) is a biomarker-driven Princess Margaret Cancer Centre initiative to evaluate genomic and immunologic changes in tumor and blood of patient (pts) treated with PEM at 200 mg IV Q3W. Baseline (BL) and on treatment (post 2-3 doses of PEM) fresh tumor biopsies were collected for DNA/RNA sequencing, immune-profiling, and tumor PD-L1 expression by immunohistochemistry (QualTek), with longitudinal blood collection for immunophenotyping. Results: As of 9/2017, 11 MM pts (9 cutaneous, 1 choroidal, 1 mucosal) were enrolled. Median treatment duration was 6 months (0.75 – 14.25); 7 pts remain on study. 7/11 (64%) pts had partial response (PR), 3/11 (27%) progressive disease (PD) and 1 was not evaluable (NE) per RECIST 1.1. Based on data for 8 pts (6 PR, 2 PD), higher percentage (%) PD-1 (mean 70 vs 12%), TIGIT (82 vs 43%) and 4-1BB (23 vs 4%) positivity on CD8 T cells in tumor vs blood was seen (all p &lt; 0.05). In BL blood, lower % TIGIT positive CD8 T cells was detected in pts with PR vs PD (38 vs 58%, p &lt; 0.05). T cell profile of 4 pts with paired BL and post PEM samples is shown in Table 1. Mean tumor PD-L1 score was 24% (0 - 90%), with no correlation to PEM response. Genomic analysis on 8 pts (5 PR, 2 PD, 1 NE) found mean tumor mutation burden (TMB) of 538. B2M deletion was seen in 2/2 PD and 0/6 PR pts. Among pts with PRs, 4/6 (67%) had TAP1 and TAP2 amplification post PEM therapy. HLA-A and HLA-B amplifications were each found in 3/6 (50%) PR pts, with only 1 present at BL. TMB was not associated with response to PEM. Conclusions: In this exploratory analysis of MM pts treated with PEM, there was significantly discordant T cell phenotype in tumor vs blood at BL. Our findings suggest B2M deletion to be associated with PEM resistance. The observed development of HLA-A/B and TAP1/2 amplification in 50-67% pts with PR post PEM suggests potential predictive significance, although further validation is warranted. Clinical trial information: NCT02644369. [Table: see text]

35P - A retrospective analysis to evaluate prevalence and correlation between PD-L1 score and tumor mutational burden (TMB) levels in patients with solid tumor malignancies

35P - A retrospective analysis to evaluate prevalence and correlation between PD-L1 score and tumor mutational burden (TMB) levels in patients with solid tumor malignancies

34P - A retrospective analysis of response to checkpoint inhibitors and its correlation to both PD-L1 score and tumor mutation burden in patients with solid tumor malignancies

34P - A retrospective analysis of response to checkpoint inhibitors and its correlation to both PD-L1 score and tumor mutation burden in patients with solid tumor malignancies

PD-L1 expression is associated with epithelial-to-mesenchymal transition in adenocarcinoma of the lung

PD-1/PD-L1-targeted immunotherapy has emerged as a promising therapeutic strategy for pulmonary adenocarcinoma (pADC). Epithelial-to-mesenchymal transition (EMT) is involved in the progression and immune evasion of cancers. Therefore, we investigated the association between PD-L1 expression and EMT phenotype in pADC. Immunohistochemistry for E-cadherin (epithelial marker), ZEB1, SNAIL, SLUG, vimentin (mesenchymal markers), PD-L1, CD8, and PD-1 was performed on 477 cases of pADC. Cases were classified into epithelial, mesenchymal, epithelial-mesenchymal, and unspecified types based on immunohistochemical results. PD-L1 expression was scored as 0 in 14.0% (n=67), 1 in 26.4% (n=126), 2 in 51.2% (n=244), and 3 in 8.4% (n=40). PD-L1 score was positively correlated with SNAIL and vimentin H scores (P<.001, both). After dichotomizing patients into PD-L1-negative and PD-L1-positive groups, PD-L1 positivity was significantly higher in patients with mesenchymal (71.2%; 84/118) and epithelial-mesenchymal (62.7%; 84/134) phenotypes compared with those with epithelial (50.6%; 44/87) and unspecified (50.0%; 35/70) phenotypes (P=.005). The significant association between PD-L1 expression and EMT phenotype was maintained in EGFR-mutated pADCs. Moreover, cases with EMT phenotype (ie, mesenchymal and epithelial-mesenchymal) were infiltrated by higher numbers of CD8+ and PD-1+ cells than those with epithelial and unspecified phenotypes in EGFR-mutated pADCs (P=.043 for CD8+ cells and P<.001 for PD-l+ cells). Particularly, cases with EMT phenotype and PD-L1 expression showed the greatest amount of CD8+ and PD-1+ cells in EGFR-mutated cases (P=.043 for CD8+ cells and P=.005 for PD-1+ cells). This study demonstrates that EMT phenotype is related to PD-L1 overexpression in pADC cells and patients with EMT-phenotype pADC may benefit from PD-1/PD-L1-blocking immunotherapy.

Expression of programmed death 1 (PD-1) and its ligand (PD-L1) in thymic epithelial tumors: Impact on treatment efficacy and alteration in expression after chemotherapy

BackgroundsTo understand the clinical impact of PD-1/L1 expression in thymoma (TM) and thymic carcinoma (TC), we evaluated the frequency of PD-1/L1 expression in pre/post chemotherapy specimens and the correlation with the treatment efficacy. MethodsThe expression of PD-1/L1 was evaluated using immunohistochemistry in patients with TM or TC treated with chemotherapy between 2000 and 2014. Using formalin-fixed, paraffin-embedded tissue samples and a PD-L1 antibody, the expression of PD-L1 in the TM and TC specimens was reported in terms of the H-score (0–300), with a score ≥1 being defined as positive. The PD-1 expression in the tumor-infiltrating immune cells was evaluated based on the intensity (0–3) of staining using a PD-1 antibody. The objective response rate, progression-free survival, and the difference in PD-1/L1 expression between the pre/post chemotherapy were evaluated. ResultsThirty patients (TM/TC 12/18) were evaluated. PD-L1 positivity were TM/TC 67%/41%. Within the PD-L1 positive/negative populations, the objective response rates were 50%/0% for TM and 14%/20% for TC. No significant differences in progression-free survival were seen according to the PD-L1 expression status. Increases in both the PD-L1 and PD-1 scores were observed after chemotherapy in six serial pre/post chemotherapy TM specimens, with a mean PD-L1 score and a median PD-1 intensity of 42/93, and 0/2.5, respectively. ConclusionsThe substantially high expression of PD-L1 and the increase in PD-L1 and PD-1 expression after chemotherapy supports anti-PD-1/L1 drugs therapy for TM and TC as well as the development of a strategy for its sequential use after chemotherapy.

Abstract A90: Tumor associated PDL1 expression pattern in microsopically tumor positive sentinel lymph nodes (SLN) in patients with melanoma

Abstract Background: Anti-PD1/PDL1 immunotherapy has demonstrated unprecedented clinical antitumor activity in patients with metastatic melanoma supporting planned testing as adjuvant therapy in patients with operable disease at high risk of recurrence and death. Characterization of PDL1 expression within clinically/radiologically negative but microscopically tumor positive sentinel lymph nodes is critical to our understanding of the relevance of this immune checkpoint pathway for adjuvant therapy. Methods: Cut sections (5 µm) of sentinel lymph node from 25 patients reported positive for melanoma micrometastasis were included. H&amp;E staining was performed, followed by PD-L1 immunohistochemical (IHC) staining using a preliminary IHC assay with anti-PD-L1 antibody clone 22C3. Slides from 2 patients were also stained using an anti-HMB45/MelA protocol to better ascertain the presence and/or localization of melanoma lesions in the tissue in order to facilitate interpretation of the PD-L1 staining in those samples. The slides were separately evaluated by 2 pathologists. Samples containing metastatic melanoma lesions were scored separately for PD-L1 expression in intratumoral (including along tumor periphery but with clear tumor cell labeling) and peritumoral (expression external to tumor nodule in immediately surrounding tissue) locations. For intratumoral signal, attempts were made to classify the expression as tumor cell associated (T), non-tumor cell associated (NT), or both (T/NT). Scores were assigned using a 0-5 semiquantitative scale assessing prevalence of positive cells where 0 = negative, 1 = minimal or rare, 2 = low, 3 = moderate, 4 = high, and 5 = very high. Samples where melanin content was too high to confidently assign a PD-L1 score were specifically noted. Results: Twenty patients where metastatic melanoma presence in regional sentinel nodes was confirmed by H&amp;E review of the cut sections were included in the final analysis of PDL1 expression in the target population. Table 1 summarizes the study results. Except for samples where melanin content was too high to confidently assign a PD-L1 score, all SLNs examined demonstrated evidence of intratumoral and/or peritumoral PD-L1 expression. Conclusions: PD-L1 expression is readily detectable intratumorally and/or peritumorally within melanoma micrometastases in the SLN. These results support the testing of a therapeutic role for PD1/PDL1 inhibition in the adjuvant therapy setting, targeting melanoma micrometastases. Patient with SLN+ Tumor Associated Expression PD-L1 Score / Tumor ---- PD-L1 Score / Peritumoral 1. 1 T, very weak ---- 1 2. 4, predominantly T, peripheral ---- 1 3. melanin confounds ---- 2 4. 0 ---- 2 (adjacent subcapsular sinus) 5. 4 T/NT---- 2 6. 2, predominantly NT ---- 3 7. 0 ---- 3 (adjacent subcapsular sinus) 8. 2.5, predominantly NT ---- 3 9. 4.5, predominantly T, weak ---- 2 (adjacent sinus) 10. melanin confounds ---- 0 11. 1 T ---- 2.5 12. 0* ---- 2 13. 1 T ---- 1 14. 2.5 NT ---- 2 15. 2 predominantly NT ---- 3 16. 1 T ---- 2 17. melanin confounds ---- 0 18. 1 ---- 1 19. 1 NT ---- 2.5 20. 2 T, melanin confounds ---- 2 *Very few tumor cells. In one region where a few tumor cells were present, a dense area of PD-L1 positivity was present directly adjacent but was impossible to say if all were non-tumor; this is interpreted as most likely. Citation Format: Ahmad A. Tarhini, Jennifer A. Yearley, Christopher Gibson, Uma N. M. Rao, Cindy Sander, John M. Kirkwood. Tumor associated PDL1 expression pattern in microsopically tumor positive sentinel lymph nodes (SLN) in patients with melanoma. [abstract]. In: Proceedings of the AACR Special Conference: Tumor Immunology and Immunotherapy: A New Chapter; December 1-4, 2014; Orlando, FL. Philadelphia (PA): AACR; Cancer Immunol Res 2015;3(10 Suppl):Abstract nr A90.

Tumor associated PD-L1 expression pattern in microscopically tumor positive sentinel lymph nodes in patients with melanoma.

BackgroundCharacterization of PD-L1 expression within clinically/radiologically negative but microscopically tumor positive sentinel lymph nodes (SLN) is important to our understanding of the relevance of this immune checkpoint pathway for adjuvant therapy.MethodsPatients included had primary cutaneous melanoma, Breslow thickness of 2.01–4.0 or >4 mm with or without tumor ulceration (T3a, T3b, T4a, T4b). All patients had microscopically tumor positive SLN. Hematoxylin and eosin (H&E) staining was performed, followed by PD-L1 immunohistochemical (IHC) staining using a preliminary IHC assay with anti-PD-L1 antibody clone 22C3. The slides were separately evaluated by two pathologists (JY and CG). Samples containing metastatic melanoma lesions were scored separately for PD-L1 expression in intratumoral and peritumoral locations, by utilizing two scoring methods.ResultsTwenty-four patients where metastatic melanoma presence in the SLN was confirmed by H&E review of the cut sections were included in the final analysis of PD-L1 expression. SLN tumor size ranged from 1 to 2 mm. For three patients, the melanin content was too high to confidently assign a PD-L1 score. For the remaining 21 patients, all had some evidence of either intratumoral or peritumoral PD-L1 expression. The frequency of intratumoral tumor-associated PD-L1 expression was: 0 % of tumor cells (3 pts, 14 %); <1 % (5 pts, 24 %); 1–10 % (6 pts, 29 %) and >10 % (7 pts, 33 %).ConclusionsTumor-associated PD-L1 expression is readily detectable within melanoma micrometastases in the SLN of the majority of patients. These results support the testing of a therapeutic role for PD1/PD-L1 inhibition in the adjuvant setting, targeting melanoma micrometastases.

Association of PD-1 and PD-L1 protein expression in matched primary and metastatic renal cell carcinoma tumors.

418 Background: Therapy targeting programmed death (PD)-1 and PD-L1 proteins has activity in metastatic renal cell carcinoma (mRCC). Expression of these proteins on tumor and infiltrating immune cells is associated with a higher response to drugs inhibiting this pathway. However, these associations have been made based on primary tumor expression, while therapy is directed against metastatic deposits. Methods: Patients with mRCC and metastases (all clear cell; 3 with sarcomatoid changes) who had undergone resection of both the primary and at least one metastatic tumor were included. Samples were evaluated for PD-1 and PD-L1 expression by immunohistochemistry using anti-PD-1 clone NAT105 and Merck proprietary anti-PD-L1 clone 22C3. Stained sections were scored for PD-1 and PD-L1 using a semi-quantitative 0 to 5 scale (0 = negative, 1 = rare, 2 = low, 3 = moderate, 4= high, 5 = very high). PD-1 expression was limited to cells with lymphoid morphology, PD-L1 expression was evaluated for both tumor and non-tumor (inflammatory, endothelial) cells. Linear regression analysis was performed to assess significance of correlations. Results: Fifty matched primary and metastatic RCC tissue sets were analyzed with 48 evaluable matched pairs. PD-1 score greater than 3 (considered positive) was detected in 9 primary tumors (18%) and 9 metastatic sites (18%). PD-L1 score greater than 3 was detected in 9 primary tumors (18%) and 12 metastatic sites (24%). There was a correlation between PD-1 scores for primary and metastatic pairs (R2=0.194; p&lt;0.002) and between PD-L1 scores for primary and metastatic pairs (R2=0.319; p&lt;0.0001). There was an association between PD-1 score and PD-L1 score for primary tumors (R2=0.513; p&lt;0.0001) and for metastatic sites (R2=0.449; p&lt;0.0001). For PD-1 score, there were 10 pairs (21%) in which either the primary or metastatic site score was ≥3, but the matched pair was &lt; 3. For PD-L1 score, there were 9 pairs (19%) in which either the primary or metastatic site score was ≥3, but the matched pair was &lt; 3. Conclusions: The expression of PD1 and PD-L1 in primary clear cell RCC tumors is correlated with metastatic site expression, although there are a substantial percentage of tumors with discordance.