Analysis of disease-free survival in CheckMate 274 by PD-L1 combined positive score and tumor proportion score.

How low can you go? PD-L1 expression as a biomarker in trials of cancer immunotherapy

High Expression of Programmed Death Ligand 1 and Programmed Death Ligand 2 in Ophthalmic Sebaceous Carcinoma: The Case for a Clinical Trial of Checkpoint Inhibitors

2656 Background: Immune checkpoint inhibitors (ICIs) benefit patients with multiple cancer types, and the expression of programmed death ligand 1 (PD-L1) protein assessed by immunohistochemistry (IHC) has been correlated with response and survival benefit from anti-PD-1/PD-L1 ICI therapies in several cancer types. IHC assay with E1L3N clone is currently approved by the NMPA for PD-L1 detection in non-small cell lung cancer (NSCLC), but its performance in other cancer types needs to be further validated. Thus, we performed this study to examine the prevalence of PD-L1 expression in a wide variety of tumor types in Chinese pan-cancer patients using the E1L3N antibody clone. Methods: From 2019 to 2022, a total of 13,647 patient across multiple tumor types were analyzed. Tumor tissue samples were subjected to IHC. PD-L1 expression was tested using the E1L3N PD-L1 IHC assay (Amoy Diagnostics, Xiamen, China), and scored with the tumor proportion score (TPS) and combined positive score (CPS). PD-L1 expression status was identified as positive and negative according to TPS or CPS. For NSCLC: negative (TPS < 1%), low expression (TPS 1-49%), and high expression (TPS ≥ 50%). For gastric/gastroesophageal junction carcinoma (GC/GJC): negative (CPS < 1), low expression (CPS 1-5), and high expression (CPS ≥ 5). For other cancer: negative (CPS < 1), low expression (CPS 1-10), and high expression (CPS ≥ 10). Results: In total, 14 cancer types were enrolled in the present study, including 68.2% NSCLC, 6.9% colorectal carcinoma (CRC), and 3.8% GC/GJC, and the remaining 21.1% include esophageal cancer (EC), breast cancer, cervical cancer (CC), biliary tract cancer (BTC), pancreatic cancer (PC), ovarian cancer, renal cancer, small cell lung cancer, urothelial carcinoma (UC), bladder cancer, head and neck squamous cell carcinoma (HNSCC), etc. PD-L1 positivity by TPS was observed in 52.5% NSCLC patients, similar to the frequency of PD-L1 expression in Chinese NSCLC patients detected using 22C3 clone. 76.0% and 36.7% of GC/GJC patients with PD-L1-positive tumors at a CPS cutoff of ≥ 1 and ≥ 5, respectively. PD-L1 positive expression frequency was higher than that in western GC/GJC patients detected using 22C3 and 28-8 clone (as reported from previous research: PD-L1 positivity with CPS ≥ 1 was 45.5% using the 22C3 assay and 49.1% using the 28-8 assay). 73.6% and 17.5% of CRC patients with PD-L1-positive tumors at a CPS cutoff of ≥ 1 and ≥ 10, respectively. In addition, 8 tumors (EC, CC, breast cancer, PC, BTC, UC, HNSCC, and bladder cancer) had the prevalence of PD-L1 CPS ≥ 1 greater than 50%. Conclusions: E1L3N can be used as a reliable alternative antibody clone to evaluate PD-L1 expression status not only in NSCLC patients but also in other cancer types.

Oxidative stress and the immune microenvironment both contribute to the pathogenesis of esophageal squamous cell carcinoma (ESCC). However, their interrelationships remain poorly understood. We aimed to examine the status of key molecules involved in oxidative stress and the immune microenvironment, as well as their relationships with each other and with clinicopathological features and prognosis in ESCC. The expression of programmed death-ligand 1 (PD-L1), CD8, nuclear factor erythroid-2 related factor-2 (NRF2), and NAD(P)H quinone oxidoreductase 1 (NQO1) was detected using immunohistochemistry in tissue samples from 176 patients with ESCC. We employed both combined positive score (CPS) and tumor proportion score (TPS) to evaluate PD-L1 expression and found a positive correlation between CPS and TPS. Notably, PD-L1 expression, as assessed by either CPS or TPS, was positively correlated with both NRF2 nuclear score and NQO1 score in stage II-IV ESCC. We also observed a positive correlation between the density of CD8+ T cells and PD-L1 expression. Furthermore, high levels of PD-L1 CPS, but not TPS, were associated with advanced TNM stage and lymph node metastases. Moreover, both PD-L1 CPS and the nuclear expression of NRF2 were found to be predictive of shorter overall survival in stage II-IV ESCC. By using the Mandard-tumor regression grading (TRG) system to evaluate the pathological response of tumors to neoadjuvant chemotherapy (NACT), we found that the TRG-5 group had higher NRF2 nuclear score, PD-L1 CPS, and TPS in pre-NACT biopsy samples compared with the TRG-3 + 4 group. The NQO1 scores of post-NACT surgical specimens were significantly higher in the TRG-5 group than in the TRG 3 + 4 group. In conclusion, the expression of PD-L1 is associated with aberrant NRF2 signaling pathway, advanced TNM stage, lymph node metastases, and unfavorable prognosis. The dysregulation of PD-L1 and aberrant activation of the NRF2 signaling pathway are implicated in resistance to NACT. Our findings shed light on the complex interrelationships between oxidative stress and the immune microenvironment in ESCC, which may have implications for personalized therapies and improved patient outcomes.

The tumor proportion score (TPS) and combined positive score (CPS) have been developed to assess programmed death ligand 1 (PD-L1) expression in gastric cancer (GC). This study aimed to elucidate the role of TPS and CPS as prognostic biomarkers. A total of 191 patients with GC who received curative gastrectomy were retrospectively enrolled. Double immunohistochemistry of PD-L1 and ionized calcium binding adaptor molecule 1 was performed to clearly distinguish PD-L1 expression between tumor cells and macrophages. Survival analysis for PD-L1 expression by TPS and CPS was performed. PD-L1 positivity was detected in 39 patients (20.4%) by TPS and in 137 patients (71.7%) by CPS. Patients with PD-L1 positivity by CPS experienced significantly shorter overall survival (OS) (P < 0.01) and relapse-free survival (RFS) (P = 0.01) than the other patients. In contrast, patients with either PD-L1 status by the TPS showed similar OS and RFS times. Multivariate Cox regression analysis for OS and RFS demonstrated that PD-L1 positivity by CPS was a significant independent factor for poor OS (hazard ratio [HR] 2.27, 95% confidence interval [CI] 1.27-4.37, P < 0.01) and RFS (HR 1.81, 95% CI 1.07-3.22, P = 0.03). CPS was shown to be a more useful assessment method of determining PD-L1 expression than TPS as a prognostic biomarker. This suggests that the total number of PD-L1-expressing cells, including tumor and immune cells, is a more sensitive prognostic biomarker than the number of PD-L1-expressing tumor cells in GC.

Programmed death-ligand 1 (PD-L1) expression in some tumors has prognostic implications. This work aims at investigating PD-L1 expression in diffuse large B-cell lymphoma (DLBCL) and to study its association with clinicopathological variables. The study consisted of 75 DLBCL patients who were cared for at the King Hussein Cancer Center during the period 2015-2018. The expression of PD-L1 in tumor tissue was assessed by immunohistochemistry using the anti-human PD-L1 (Clone 22C3) monoclonal antibody. The correlation between gender, age, clinical stage, pre-treatment-LDH level, tumor location, response to therapy, overall and event-free survival with PD-L1 expression was studied. Six patients were excluded from further analysis as they were in relapse at the time of tissue sampling. The tumor proportion score (TPS) was ≥1% in 16/69 (23.2%) of DLBCL cases while the combined positive score (CPS) at a cut-off of ≥20 was observed in 23/69 (33.3%) cases. No significant difference in PD-L1 expression was found between germinal center B-cell-like (GCB) and non-GCB subtypes. Similarly, no differences in PD-L1 expression (at CPS ≥20 and TPS ≥1) were found between different genders, age groups, clinical stages, tumor location, and patient response to therapy. However, base-line lactate dehydrogenase was significantly elevated in patients with PD-L1 CPS ≥20. The overall survival was not significantly different between PD-L1-positive and -negative groups. On the other hand, the median event-free survival was higher in either of the PD-L1 TPS or CPS negative groups at 107months each versus 54 months in the PD-L1 positive group of either category. PD-L1 expression can predict event-free survival in DLBCL cases and therefore poor prognosis.

Background: Programmed cell death-ligand 1 (PD-L1) is of great interest in breast cancer (BC). PD-L1 IHC 22C3 pharmDx (SK006) is an approved companion diagnostic assay for patients with triple-negative breast cancer (TNBC). Multiple sub-types of BC are identified by the binary expression status (positive/negative) of human epidermal growth factor 2 (HER2) and the hormone receptors (HRs) estrogen receptor (ER) and progesterone receptor (PR). The ability of the assay to reliably identify PD-L1 expression in TNBC (ER-/PR-/HER2-) tissues was previously demonstrated. However, the sensitivity of the assay on HR+/HER2- specimens (including ER+/PR+, ER+/PR-, and ER-/PR+; non-TNBC) is not as well-known. The data presented here demonstrates the ability of the assay to reliably identify the dynamic range of PD-L1 expression on non-TNBC specimens when assessed using the Combined Positive Score (CPS). Methods: Primary and metastatic BC specimens, sourced from multiple anatomical sites, were prepared and stained according to the assay’s Instructions for Use (IFU). The PD-L1 IHC 22C3 pharmDx immunohistochemical assay uses Monoclonal Mouse Anti-PD-L1, Clone 22C3, for detection of PD-L1 protein in formalin-fixed, paraffin-embedded tissues. Stained sections were scored using CPS, which is the number of PD-L1 staining cells (tumor cells, lymphocytes, macrophages) divided by the total number of viable tumor cells, multiplied by 100. CPS totals are displayed as integers with a maximum CPS of 100. Additional cut sections from the same specimens were also prepared and stained using Agilent ER, PR and HER2 reagents, where positivity or status was based upon the IFU for each reagent. Each specimen was then grouped into non-TNBC or TNBC sub-types. Prevalence was assessed to demonstrate that the assay will detect the target molecule (PD-L1 protein) in BC specimens across the dynamic range of PD-L1 expression (CPS 0–100). For the purposes of this abstract, scores were reported according to the CPS ≥ 1 and CPS ≥ 10 cutoffs. Results: Prevalence data of Agilent’s internal BC tissue bank was evaluated. At the time of the assessment, two categories of BC specimens and their totals were grouped: 149 non-TNBC and 184 TNBC. When examining the CPS ranges for the non-TNBC specimens, 55/149 (36.9%) were found to have a CPS ≥ 1 and 17/149 (11.4%) had a CPS ≥ 10. When examining the CPS for the TNBC specimens, 115/184 (62.5%) were found to have a CPS ≥ 1 and 57/184 (31.0%) had a CPS ≥ 10. For all specimens of each cohort (non-TNBC and TNBC), the assay identified PD-L1 expression across the dynamic range. This analysis indicates that the dynamic range of PD-L1 expression is present across all BC tissues regardless of HR status, though a higher prevalence is noted in TNBC specimens at the CPS ≥ 1 cutoff. Conclusions: Overall, the assay detected PD-L1 protein across the dynamic expression range (CPS 0–100) for both non-TNBC and TNBC specimens. In this small specimen cohort, retrospective analysis demonstrated that HR status may correlate to a difference in PD-L1 prevalence in HER2- breast cancer specimens. This analysis further demonstrated that PD-L1 IHC 22C3 pharmDx can identify PD-L1 expression in the dynamic range within non-TNBC specimens, specifically. Further studies should be conducted within different combinations of HR status and PD-L1 prevalence to expand on this data, to address gaps in knowledge, and to learn if HR status may inform PD-L1 immune checkpoint inhibitor therapy benefit for BC patients. Citation Format: Joseph Barreto, Jaret Quiroz, Emily Olander, Donna Kell, Siena Tabuena-Frolli, Camilla Recke, Kelly Martyniuk. Hormone receptor status and PD-L1 expression prevalence in breast cancer when assessed with PD-L1 IHC 22C3 pharmDx and Combined Positive Score [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P3-03-18.

IntroductionIn advanced-stage NSCLC, tumor proportion score (TPS) is typically used to predict the efficacy of immune checkpoint inhibitors (ICIs). Nevertheless, in other cancer types, the combined positive score (CPS), which covers programmed death-ligand 1 (PD-L1) expression on both tumor and surrounding immune cells, is used. We investigated the predictive value of CPS in comparison to TPS in advanced NSCLC. MethodsA monocenter, retrospective study was performed in patients with advanced NSCLC treated with ICI monotherapy between 2015 and 2021. Hematoxylin and eosin and PD-L1 were stained on baseline tumor biopsy samples to score PD-L1 by both TPS and CPS. Positivity for TPS and CPS was defined as a score of 1% or above. Progression-free survival and overall survival (OS) were assessed for TPS and CPS scores. ResultsAmong the 187 included patients, PD-L1 positivity was found in 112 patients (59.9%) by TPS and 135 patients (72.2%) by CPS. There was no significant difference in OS between TPS− and TPS+ patients (p = 0.20). Nevertheless, CPS+ patients did have a longer OS than CPS− patients (p = 0.006). OS was superior in both TPS−/CPS+ and TPS+/CPS+ as compared with TPS−/CPS− cases (p = 0.018 and p = 0.015, respectively), whereas no considerable differences in OS were found between TPS−/CPS+ and TPS+/CPS+ cases. ConclusionsThis retrospective real-world population study revealed that CPS differentiated OS better than TPS in patients with advanced NSCLC with ICI monotherapy. Remarkably, this was driven by the performance of the TPS−/CPS+ subgroup, indicating that CPS may be a better predictive biomarker for ICI efficacy.

Predictive biomarkers for nivolumab in recurrent or metastatic head and neck squamous cell carcinoma (RMHNSCC) have not yet been established. The tumor proportion score (TPS), combined positive score (CPS), and soluble forms of programmed cell death ligand-1 (PD-L1) and programmed cell death ligand-2 (PD-L2) were retrospectively analyzed in patients with RMHNSCC treated with nivolumab. The positivity rates for TPS (PD-L1), CPS (PD-L1), TPS (PD-L2), and CPS (PD-L2) were 73.8%, 78.2%, 56.4%, and 78.2%, respectively. Patients with high TPS (PD-L1), CPS (PD-L1), or CPS (PD-L1 and PD-L2) showed significantly prolonged progression-free survival. Favorable overall survival was associated with high CPS (PD-L1 and PD-L2) and low soluble PD-L1 and PD-L2 levels. The expressions of tissue and soluble PD-L1/2 were not correlated. Our study revealed that compared to PD-L1 expression alone, dual expression of PD-L1 and PD-L2 in tissue or soluble form could be feasible biomarkers in patients with RMHNSCC who received nivolumab.

BackgroundThe COVID-19 pandemic brought a host of new challenges, including the immediate need for digital solutions addressing the lack of remote options available to pathologists in the field of immunohistochemistry...

Genetic alternations and immune characteristics in patients with small cell lung cancer

BackgroundFor non-small cell lung cancer (NSCLC) the most used method for analysing programmed cell death ligand 1 (PD-L1) expression is the Tumor Proportion Score (TPS). Nevertheless, for other tumour types,...

Background: Immune checkpoint inhibitors (ICIs), which improve overall survival (OS) among patients with advanced solid tumors, have been approved by the US Food and Drug Administration (FDA) as standard first-line treatment in several cancers, regardless of programmed death ligand 1 (PD-L1) status. However, the benefits of ICIs for patients with low or negative PD-L1 expression remain unclear. Methods: This analysis encompassed cancers eligible for first-line ICI treatment without PD-L1-based restrictions. We included randomized, parallel-group phase III clinical trials that compared immunotherapy with conventional care for advanced solid malignancies until December 2023; all studies reported OS and performed PD-L1-based stratification. Summary effects were estimated using random-effects models. Reconstructed individual patient data (IPD) from reported Kaplan-Meier (KM) curves of OS and PFS using the "IPDfromKM" method, while also inferring unreported data for the PD-L1 low-expression group through the application of "KMSubtraction". Pooled analysis was conducted using the KM method with the log-rank test and restricted mean survival time difference (RMST-D) to assess ICI benefit under various cutoff values and scoring methods. This study is registered with PROSPERO (CRD42023441048). Results: In the study-level analysis, 39 randomized controlled trials involving 25,536 patients revealed that both high and low PD-L1-expressing subgroups derived a significant clinical benefit from ICIs (hazard ratios [HRs] were 0·69 and 0·81, respectively). There was no survival benefit for patients with low PD-L1 expression in the following types of cancers: esophageal squamous cell carcinoma (ESCC), biliary tract cancer (BTC), hepatocellular carcinoma (HCC), and melanoma. Conversely, patients with human epidermal growth factor receptor 2 (HER2)-negative gastroesophageal adenocarcinoma (GEA) exhibited a marginal benefit (HR=0·89, 95% confidence interval [CI]=0·80-0·99). In patient-level analysis, the ESCC combined positive score (CPS) &amp;lt;10 subgroup derived a significant OS benefit (HR 0·78, 95% CI=0·63-0·96, P=0·02; RMST-D=2·34 months); the ESCC tumor proportion score (TPS) &amp;lt;1% subgroup showed no significant improvement in OS benefit (HR=0·87, 95% CI=0·72-1·06, P=0·16). Among patients with HER2-negative GEA, no significant OS benefit was observed in the CPS &amp;lt;1, CPS &amp;lt;5, and CPS 1-4 subgroups (HRs=0·95, 0·89, and 0·97, respectively). There were significant OS benefits in the subgroups with CPS &amp;lt;10 (HR=0·867, 95% CI=0·754-0·996, P=0·0482; RMST-D=1·78 months, P=0·038) and CPS 1-9 (HR=0·83, 95% CI=0·73-0·95, P=0·0085; RMST-D=2·21 months, P=0·007). Interpretation: Study-level data suggest that first-line ICI benefits for low or negative PD-L1 expression subgroups with BTC, HCC, and melanoma require further evidence. Patient-level data indicate that, contrary to FDA guidelines, ESCCs with TPS &amp;lt;1% and HER2-negative GEAs with CPS &amp;lt;5 do not benefit from the addition of ICIs to conventional chemotherapy. More nuanced clinical trials and predictive biomarkers are warranted. Citation Format: Peng Wu, Chaoqi Zhang, Dongyu Li, Xuanyu Gu, Dexin Shang, Nan Sun, Jie He. Individual Patient Analysis of Clinical Efficacy for PD-1/PD-L1 Inhibitors as First-Line Standard Treatment in Advanced Solid Tumors Exhibiting Low PD-L1 Expression [abstract]. In: Proceedings of the AACR IO Conference: Discovery and Innovation in Cancer Immunology: Revolutionizing Treatment through Immunotherapy; 2025 Feb 23-26; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Immunol Res 2025;13(2 Suppl):Abstract nr B033.

Programmed death ligand 1 (PD-L1) is a prognostic marker in several malignancies, but the prognostic value of PD-L1 expression in colorectal cancer (CRC) is inconclusive. Lack of standardized scoring systems for PD-L1 evaluation in CRC has led to inconsistent findings. This exploratory study aimed to evaluate PD-L1 expression using manual and digital methods and correlate the results to overall survival (OS) in a cohort of patients with pT3 and pT4 colon cancer (CC). From a previously published study, 162 patients with pT3 and pT4 CC were included. One tumor slide representing the invasive margin was selected and stained for PD-L1 (22C3). PD-L1 was evaluated according to the tumor proportion score (TPS), the immune cell score (ICS), and the combined positive score (CPS). Additionally, a digital algorithm for detecting PD-L1 positive cells was developed. The manual and digital PD-L1 expression scores were correlated to OS in the entire cohort, and in subgroups according to mismatch repair (MMR) status. Only 1 case was classified with a TPS of ≥ 1%. The ICS was classified as < 1%, 1-9%, and ≥ 10% in 66%, 30.2% and 3.7% of the tumors, respectively. The CPS was classified as < 1, 1-9, and ≥ 10 in 82.7%, 15.4%, and 1.9% of the tumors, respectively. A high digital PD-L1 expression score was an independent prognostic factor for longer OS, adjusted for age, pT-category and adjuvant chemotherapy (aHR = 0.40, 95% CI = 0.19-0.86, p = 0.018). In the pMMR subgroup, both a CPS ≥ 1 and a high digital score were significantly associated with longer OS in the multivariate analyses (aHR = 0.24, 95% CI = 0.06-0.99, p = 0.049, and aHR = 0.34, 95% CI = 0.12-0.97, p = 0.043, respectively). Our results suggest that high PD-L1 expression is associated with longer OS. In future studies examining PD-L1 expression as a prognostic biomarker in CC, assessing PD-L1 expression using a digital approach or the CPS can be recommended.

Comparison of three PD-L1 immunohistochemical assays in head and neck squamous cell carcinoma (HNSCC).