SP142 Antibody Research Articles

Abstract C149: Relationship between PD-L1 expression and clinicopathological characteristics in Hispanic/ Latino women with breast cancer

Abstract Background: Programmed death ligand 1 (PD-L1) expression in breast cancer serves as a prognostic marker, particularly in aggressive subtypes like triple-negative breast cancer (TNBC), where it helps in predicting response to immune checkpoint inhibitors and guides immunotherapy use. Although several studies have assessed the prevalence of PD-L1 expression in breast cancer, data from the Hispanic population, especially in Latin American countries, is lacking. This study evaluates PD-L1 expression and its correlation with pathological features in Colombian women. Methods: Women with breast cancer tested for PD-L1 were selected from the archives of the Oncologic Pathology Department at the National Cancer Institute of Colombia from September 2021 to May 2024. Information from their pathology reports was extracted, and correlation analyses were carried out using Fisher's exact test Results: A total of 60 reports were screened, comprising 56 cases of ductal invasive breast carcinoma and 1 case of lobular invasive breast carcinoma. 48 individuals were tested using the PD-L1 22C3 antibody, with 41.67% of them being positive for PD-L1 expression. Among the 48 tumors tested with 22C3, 35 cases were classified as TNBC, and 13 cases corresponded to luminal B subtype with a low positive expression (1-10%) of estrogen receptors. The percentage of positive PD-L1 cases was 37.14% for TNBC and 53.85% for the luminal B subtype. Furthermore, 5 HER2-low cases were identified with 2 of them being positive for PD-L1 expression. In the correlation analyses, PD-L1 positive expression was significantly associated with a more aggressive histologic grade compared to PD-L1 negative tumors (p = 0.007). No significant association was found between PD-L1 expression and HR, HER2, or KI67 status, nor with age, necrosis, lymphovascular involvement, perineural invasion, desmoplastic response, or immune cell infiltration. Among the 12 individuals tested for PD-L1 with the SP142 antibody, none showed positive expression, precluding correlation analyses. Conclusion: This study highlights a significant association between PD-L1 expression and aggressive histologic grade in Colombian women with breast cancer. The findings underscore the potential of PD-L1 as a prognostic marker and its relevance in guiding immunotherapy in this population. Further research is needed to explore PD-L1 expression in broader Hispanic groups to better understand its clinical implications. Citation Format: Daniel Mendivelso-González, Patricia López- Correa, Nicolás Prada-Aceros, Alfredo E Romero-Rojas, Rafael Parra-Medina. Relationship between PD-L1 expression and clinicopathological characteristics in Hispanic/ Latino women with breast cancer [abstract]. In: Proceedings of the 17th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2024 Sep 21-24; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2024;33(9 Suppl):Abstract nr C149.

Tumour immune characterisation of primary triple-negative breast cancer using automated image quantification of immunohistochemistry-stained immune cells

The tumour immune microenvironment (TIME) in breast cancer is acknowledged with an increasing role in treatment response and prognosis. With a growing number of immune markers analysed, digital image analysis may facilitate broader TIME understanding, even in single-plex IHC data. To facilitate analyses of the latter an open-source image analysis pipeline, Tissue microarray MArker Quantification (TMArQ), was developed and applied to single-plex stainings for p53, CD3, CD4, CD8, CD20, CD68, FOXP3, and PD-L1 (SP142 antibody) in a 218-patient triple negative breast cancer (TNBC) cohort with complementary pathology scorings, clinicopathological, whole genome sequencing, and RNA-sequencing data. TMArQ’s cell counts for analysed immune markers were on par with results from alternative methods and consistent with both estimates from human pathology review, different quantifications and classifications derived from RNA-sequencing as well as known prognostic patterns of immune response in TNBC. The digital cell counts demonstrated how immune markers are coexpressed in the TIME when considering TNBC molecular subtypes and DNA repair deficiency, and how combination of immune status with DNA repair deficiency status can improve the prognostic stratification in chemotherapy treated patients. These results underscore the value and potential of integrating TIME and specific tumour intrinsic alterations/phenotypes for the molecular understanding of TNBC.

PD-L1 Expression in Neoplastic and Immune Cells of Thymic Epithelial Tumors: Correlations with Disease Characteristics and HDAC Expression.

Programmed death-ligand 1 (PD-L1) expression in neoplastic and immune cells of the tumor microenvironment determines the efficacy of antitumor immunity, while it can be regulated at the epigenetic level by various factors, including HDACs. In this study, we aim to evaluate the expression patterns of PD-L1 in thymic epithelial tumors (TETs), while we attempt the first correlation analysis between PD-L1 and histone deacetylases (HDACs) expression. Immunohistochemistry was used to evaluate the expression of PD-L1 in tumor and immune cells of 91 TETs with SP263 and SP142 antibody clones, as well as the expressions of HDCA1, -2, -3, -4, -5, and -6. The PD-L1 tumor proportion score (TPS) was higher, while the immune cell score (IC-score) was lower in the more aggressive TET subtypes and in more advanced Masaoka-Koga stages. A positive correlation between PD-L1 and HDAC-3, -4, and -5 cytoplasmic expression was identified. Higher PD-L1 expression in neoplastic cells and lower PD-L1 expression in immune cells of TETs characterizes more aggressive and advanced neoplasms. Correlations between PD-L1 and HDAC expression unravel the impact of epigenetic regulation on the expression of immune checkpoint molecules in TETs, with possible future applications in combined therapeutic targeting.

PD-L1 expression and its correlation with clinicopathological and molecular characteristics in Chinese patients with non-small cell lung cancer.

Little is known about the relationship between programmed cell death-ligand 1 (PD-L1) expression and histologic and genetic features in real-world Chinese non-small cell lung cancer patients. From November 2017 to June 2019, tumor tissues were collected from 2674 non-small cell lung cancer patients. PD-L1 expression was detected with immunohistochemistry using the 22C3 and SP263 antibodies, and patients were stratified into subgroups based on a tumor proportion score of 1%, 1% to 49%, and ≥ 50%. Genetic alterations were profiled using targeted next-generation sequencing. In the total population, 50.5% had negative PD-L1 expression (tumor proportion score < 1%), 32.0% had low-positive expression (1%-49%), and 17.5% had high-positive expression (≥50%). The PD-L1 positive rate was 39.0% in squamous cell carcinomas and 53.6% in adenocarcinomas. PD-L1 expression was higher in squamous cell carcinomas (P < .001) and lower in adenocarcinomas (P < .001). Of the overall patient population, 11.2% had Kirsten rat sarcoma viral oncogene (KRAS) mutations, 44.9% had epidermal growth factor receptor (EGFR) mutations, 2.1% had BRAF V600E mutations, 0.3% had MET exon 14 skipping mutations, 5.4% had anaplastic lymphoma kinase translocations, and 0.9% had ROS proto-oncogene 1 translocations. Patients carrying ROS proto-oncogene 1 translocations (P = .006), KRAS (P < .001), and MET (P = .023) mutations had significantly elevated expression of PD-L1, while those harboring EGFR (P < .001) mutations had lower PD-L1 expression. In our study, PD-L1 expression was significantly higher in squamous cell carcinomas and lower in adenocarcinomas, and was positively associated with MET and KRAS mutations, as well as the wild-type EGFR gene state. Nonetheless, additional studies are needed to further validate those associations and determine the clinical significance for immune checkpoint inhibitors of these factors.

PD-L1 immunohistochemical expression in bladder urothelial cancer with SP263, SP142 and 22C3 antibodies: A comparative study

Programmed death ligand 1 (PD-L1) is currently the only biomarker used for the selection of patients with bladder urothelial cancer for immunotherapy. Several platforms, antibodies and scores are currently available for the evaluation of the expression of PD-L1 in immunohistochemistry (IHC). In this study three different antibodies (SP263, SP142 and 22C3) were compared to establish their performances and concordance rates. Twenty-four consecutive cases of surgically resected urothelial cancers of the bladder were enrolled. All cases were revised, and appropriate tumor areas were selected for IHC. Three commercially available PD-L1 antibodies were tested: 22C3 pharmDx with Dako Autostainer Link 48 (Dako, Carpinteria, Ca), and SP263 and SP142 with the Ventana BenchMark (Ventana Medical Systems, Tucson, AZ) platform. All slides were evaluated by an expert pathologist and both the tumor proportion score (TPS) and the combined positive score (CPS) were determined and compared at two different cut-off levels (≥ 1 and ≥ 10). The SP263 and 22C3 clones produced more positive results with the CPS and TPS scores, respectively. The CPS score identified more positive cases than the TPS score, irrespectively of the clone or the cut-off used; the difference was statistically significant in both the SP263 and SP142 clones with the ≥1 cut-off. No statistically significant differences were found between the clones when the ≥1 cut-off was used, irrespectively of the score. At the contrary, a statistically significant difference (p = 0.024) and a trend to significance (p = 0.082) were respectively found for the TPS and CPS scores, when the SP22C3 and the SP142 clones were compared at a cut-off level of ≥10. The ICC test using CPS was 0.676 and 0.578 for the ≥1 and ≥ 10 cut-offs respectively, and 0.729 and 0.467 respectively for the same cut-offs using TPS. This suggests that the three antibodies under investigation cannot be used interchangeably, especially the 22C3 and SP142 clones which showed statistically significant difference when TPS was tested at a ≥ 10 cut-off.

Association Between PD-L1 status Detected by SP142 and SP263 Antibody Clones and Clinical and Morphological Features of Gastric Cancer Factors

The aim is to study the relationship of PD-L1 status of SP142 and SP263 clones in gastric cancer with its clinical and morphological parameters. Material and methods. Surgical material obtained from 131 patients with a confirmed diagnosis of stomach cancer. Antibodies to PD-L1 SP263 were stained 127 cases, antibodies to PD-L1 SP142 – 126 cases. Reactions with PD-L1 were performed using the Ventana BenchMark Ultra device (Roche Ventana, USA) according to the protocols recommended by the manufacturer of primary antibodies. The results of immunohistochemical staining were compared with the clinical and morphological characteristics of gastric cancer using statistical analysis. Results. The positive PD-L1 status detected by clones SP263 and SP142 is significantly associated with type 2 macroscopic form according to the R. Bormann classification (p=0,003/p=0,003), tubular morphological type according to the WHO 5th edition, 2019 classification (p=0,001/p=0,018), intermediate morphological type according to the classification of P. Lauren (p=0,027/p=0,005) and the absence of signet ring cells (p=0,001/p=0,010). Staining with both clones used was not associated with the gender and age of patients, the size and localization of the tumor, the degree of its differentiation, the presence or absence of emboli in the lumen of blood and lymph vessels, the presence of affected lymph nodes and distant metastases, as well as the clinical stage (p&gt;0,05). The largest proportion of PD-L1 positive cases in the study of SP263/SP142 clones was found in tubular type tumors (75,6%/84,6%) according to the WHO 5th edition, 2019 classification, intermediate type (60,0%/68,0%) according to P. Lauren, type 2 (65,4%/68,0%) according to R. Bormann and in the absence of signet ring cells (70,7%/76,9%). Conclusion. Statistically significant clinical and morphological parameters of gastric cancer, where a reliable relationship between them and the expression of PD-L1 clones SP263 and SP142 was revealed, are: type 2 macroscopic form according to R. Bormann (p=0,003/p=0,003), tubular morphological type according to the classification of WHO 5th edition, 2019) (p=0,001/p=0,018), intermediate type according to P. Lauren (p=0,027/p=0,005) and the absence of signet ring cells (p=0,001/p=0,010).

Expression of PD-L1 in breast invasive lobular carcinoma.

The purpose of this study was to investigate the expression of PD-L1 in invasive lobular carcinoma (ILC) and to determine its implications. Tissue microarrays were constructed for 101 cases of ILC, and immunohistochemical staining for PD-L1 (using 22C3, SP142, and SP263 antibodies) was performed to examine the correlation between staining results and clinicopathologic parameters. The positive cut-off values were defined as tumor cell (TC)≥1%, immune cell (IC)>0%, and IC≥1%. The range of PD-L1 TC positivity was 0.0-2.0%, with PD-L1 SP263 TC showing the highest positivity of 2.0%. The range of PD-L1 IC positivity was 0-21.8% for IC ≥ 1%, with PD-L1 22C3 IC showing the highest positivity. When PD-L1 IC was positive (IC≥1%), the highest antibody agreement was observed between SP263 and SP142 (OA = 93.1%), while the lowest agreement was observed between 22C3 and SP263 (OA = 73.3%, κ = 0.040). PD-L1 22C3 IC positivity (≥1%) was associated with high nuclear grade (p = 0.002), HER-2 positivity (p = 0.019), and pleomorphic type (p = 0.002). PD-L1 expression in ILC shows a low TC positivity rate (0-2%) with various antibody clones and a variable IC positivity rate (0-21.8%). Pleomorphic type ILC exhibits higher PD-L1 IC positivity.

Morphological and Immunocytochemical Characterization of Tumor Spheroids in Ascites from High-Grade Serous Carcinoma.

Tumor spheroids in the ascites of high-grade serous carcinoma (HGSC) are poorly described. Our objective was to describe their morphological features, cellular composition, PD-1 and PD-L1 expression, and survival correlation of these parameters. The density and size of spheroids were assessed in Giemsa-stained smears; the cell composition of spheroids, including tumor cells, immune cells, capillaries, and myofibroblasts, as well as PD-1 and PD-L1 expression on tumor and immune cells was assessed in immunocytochemically stained cell block sections. Forty-seven patients with primary HGSC and malignant ascites were included. A cut-off value for a spheroid density of 10% was established, which significantly predicted overall survival. However, spheroid size did not correlate with survival outcomes. Spheroids were primarily composed of tumor cells, but the presence of lymphocytes and macrophages was also confirmed. Moreover, capillaries were present in the spheroids of three patients, but the presence of myofibroblasts was not confirmed. PD-1 was expressed on lymphocytes but not on tumor cells. PD-L1 expression was seen on both tumor and immune cells, assessed by 22C3 and SP263 antibody clones but not by the SP142 clone. Our results highlight the potential of routine cytopathological techniques to analyze spheroids in HGSC ascites as a valuable tool to investigate their potential as prognostic markers.

Real-world prevalence of PD-L1 expression in non-small cell lung cancer: an Australia-wide multi-centre retrospective observational study

An investigator-initiated, Australia-wide multi-centre retrospective observational study was undertaken to investigate the real-world prevalence of programmed death ligand-1 (PD-L1) expression in non-small cell lung carcinoma (NSCLC). Multiple centres around Australia performing PD-L1 immunohistochemistry (IHC) were invited to participate. Histologically confirmed NSCLC of any stage with a PD-L1 IHC test performed for persons aged ≥18 years between 1 January 2018 and 1 January 2020, and eligible for review, were identified at each centre, followed by data extraction and de-identification, after which data were submitted to a central site for collation and analysis. In total data from 6690 eligible PD-L1 IHC tests from histologically (75%) or cytologically (24%) confirmed NSCLC of any stage were reviewed from persons with a median age of 70 years, 43% of which were female. The majority (81%) of tests were performed using the PD-L1 IHC SP263 antibody with the Ventana BenchMark Ultra platform and 19% were performed using Dako PD-L1 IHC 22C3 pharmDx assay. Reported PD-L1 tumour proportion score (TPS) was ≥50% for 30% of all tests, with 62% and 38% scoring PD-L1 ≥1% and <1%, respectively. Relative prevalence of clinicopathological features with PD-L1 scores dichotomised to <50% and ≥50%, or to <1% and ≥1%, were examined. Females scored ≥1% slightly more often than males (64% vs 61%, respectively, p=0.013). However, there was no difference between sexes or age groups (<70 or ≥70 years) where PD-L1 scored ≥50%. Specimens from patients with higher stage (III/IV) scored ≥1% or ≥50% marginally more often compared to specimens from patients with lower stage (I/II) (p≤0.002). Proportions of primary and metastatic specimens did not differ where PD-L1 TPS was ≥1%, however more metastatic samples scored TPS ≥50% than primary samples (metastatic vs primary; 34% vs 27%, p<0.001). Cytology and biopsy specimens were equally reported, at 63% of specimens, to score TPS ≥1%, whereas cytology samples scored TPS ≥50% slightly more often than biopsy samples (34% vs 30%, respectively, p=0.004). Resection specimens (16% of samples tested) were reported to score TPS ≥50% or ≥1% less often than either biopsy or cytology samples (p<0.001). There was no difference in the proportion of tests with TPS ≥1% between PD-L1 IHC assays used, however the proportion of tests scored at TPS ≥50% was marginally higher for 22C3 compared to SP263 (34% vs 29%, respectively, p<0.001). These real-world Australian data are comparable to some previously published global real-world data, with some differences noted.

Impact of Prolonged Ischemia on the Immunohistochemical Expression of Programmed Death Ligand 1 (PD-L1).

Antibodies targeting programmed death receptor 1 or programmed death ligand 1 (PD-L1) have become a standard of care to treat different cancers; for some of these tumors, there is a correlation between tissue expression of PD-L1 and response rates in patients. Although most of the analytical challenges in the evaluation of PD-L1 expression have been standardized, preanalytical issues have been less explored. The objective of this study was to evaluate the impact of time of ischemia on the performance of 2 commonly used antibodies against PD-L1. Sixteen tonsillectomy samples were kept in ischemia for <30 minutes from sample obtention (control) and 1, 3, 6, 12, and 24 hours at room temperature before formalin fixation and paraffin embedding. Selected areas were inserted into TMA paraffin recipient blocks stained with SP142 and SP263 antibodies and evaluated by 2 blind observers. The proportion of suboptimally stained samples was significantly higher for samples with cold ischemia times 6 hours or over ( P <0.0001). False-negative results were 25% in samples exposed to 6 hours of ischemia and raised to 34% for samples remaining in ischemia for 12 or 24 hours. When all observations were pooled, SP142 provided suboptimal results in 24% of observations and SP263 in 12.5%; this is a statistically significant difference ( P =0.042). In conclusion, the quality of staining for PD-L1 in tonsil samples varies with the time of cold ischemia. The SP142 antibody presented a significantly lower tolerance to prolonged cold ischemia than SP263. These results reveal the relevance of controlled preanalytical processing of samples.

Concurrent predictors of an immune responsive tumor microenvironment within tumor mutational burden-high breast cancer.

Data supporting high tumor mutational burden (TMB-H) as a lone biomarker for an immune-responsive tumor microenvironment (TME) in metastatic breast cancer (MBC) are weak, yet tumor agnostic approval in TMB-H advanced tumors provides immune checkpoint inhibition (ICI) as a clinical option. We evaluated concurrent predictors of immune-responsive and non-responsive TME within MBC. Tumor samples from patients with MBC (N=5621) were analyzed by next-generation sequencing of DNA (592-gene panel or whole exome) and RNA (whole transcriptome) at Caris Life Sciences (Phoenix, AZ). TMB-H threshold was set to ≥ 10 muts/Mb. PDL-1 was evaluated using SP142 antibody. Gene expression profiling and RNA deconvolution were used to estimate immune and stromal cell population abundance in the TME, and transcriptomic signature of immunotherapy response (T cell-inflamed score). 461 (8.2%) TMB-H MBC samples were identified. Consistent with prior studies, TMB-H tumors exhibited significant dMMR/MSI-H enrichment (7 vs. 0%, p<0.0001) and PD-L1+ expression (36 vs. 28%, p<0.05) compared to TMB-L. Across all samples, T cell-inflamed scores were weakly correlated with TMB. TMB-H was not associated with significantly increased immune responsive cell types (CD8+ T-cells, NK cells, or B cells) or immune response gene signatures (e.g. antigen presentation), yet positive trends were observed, while immunosuppressive fibroblasts were significantly decreased in TMB-H tumors (0.84-fold change compared to TMB-L, P<0.05). HR+/HER2- breast cancer was the only subtype in which TMB-H tumors exhibited increased T cell-inflamed scores vs. TMB-L. Concurrent PD-L1+ or dMMR/MSI-H with TMB-H was associated with high T cell-inflamed scores in both HR+/HER2- and TNBC. Among several associated biomarkers, B2M mutations and CD274 amplifications were positively associated with T-cell inflamed scores in TMB-H tumors; CDH1 and ERBB2 mutations were negatively associated. High TMB alone does not strongly correlate with immune infiltrate or immune-related gene signatures in MBC. TMB-H predicts T-cell inflamed signature compared to TMB-L in HR+/HER2- tumors only. Along with MSI-H and PD-L1+, several biomarkers, including B2M mutation and CD274 amplification, may help predict ICI benefit amongst TMB-H tumors. Co-occurring biomarkers within TMB-H breast cancer warrant evaluation in larger cohorts for response or resistance to ICI to develop composite predictive biomarkers in MBC.

Programmed death-ligand 1 (PD-L1) expression in cervical intraepithelial neoplasia and cervical squamous cell carcinoma of HIV-infected and non-infected patients

Programmed death-ligand 1 (PD-L1) is overexpressed in cervical carcinoma, hindering tumor destruction. The aim of this study was to assess PD-L1 expression by immunohistochemistry in cervical squamous cell carcinoma (SCC) and squamous intraepithelial lesions (SILs) from human immunodeficiency virus–positive (HIV+) and human immunodeficiency virus-negative (HIV-) patients. A total of 166 SCC and SIL samples of HIV+ and HIV- patients were included and analyzed for PD-L1 expression through tumor proportion score (TPS), and results were stratified in five TPS groups using SP263 antibody and, combined positive score (CPS) using 22C3 antibody. In cohort 1 (SP263 clone), all HIV+ patients were negative for intraepithelial lesion or malignancy (NILM), and low-grade squamous intraepithelial lesions (LSILs) scored < 1; and 87.5% of high-grade squamous intraepithelial lesions (HSILs) adjacent to SCC, 19% of HSILs non-adjacent to SCC, and 69% of SCCs scored ≥ 1 (15.4% scored 5). In HIV- patients, all NILM, LSILs, HSILs adjacent to SCC, and two HSILs non-adjacent to SCC scored < 1. SCC: 88.2% scored ≥ 1 and 5.9% scored 5. In cohort 2 (SP263 and 22C3 clones), 16.7% of HIV+ patients with SCC were positive with both clones, CPS ≥ 1 (22C3) or score 5 (≥ 50%) (SP263), showing no significant differences in positivity between both clones. These results indicate that a relatively low percentage of SCCs (16.7%; both in HIV+ and in HIV- patients) express PD-L1 (TPS ≥ 50% and CPS > 1), which may be due to some samples being archival material, sample characteristics, or use of different methodologies, highlighting the need for standardization of PD-L1 assessment in SCC of the cervix. The fact that PD-L1 is overexpressed in SILs of HIV+ patients suggests potential additional applications for immunotherapy in this disease.

Tumor-Infiltrating Lymphocytes and PD-L1 Expression in Pleomorphic Lobular Breast Carcinoma.

The prognostic and predictive role of stromal tumor-infiltrating lymphocytes (sTILs) is undetermined in pleomorphic invasive lobular cancer (pILC). The same applies for the expression of PD-1/PD-L1 in this rare breast cancer subtype. Here, we aimed to investigate the expression of sTILs and analyze the PD-L1 expression levels in pILC. Archival tissues from sixty-six patients with pILC were collected. The sTIL density was scored as a percentage of tumor area using the following cut-offs: 0%; <5%; 5-9%; and 10-50%. The PD-L1 expression was analyzed using IHC on formalin-fixed, paraffin-embedded tissue sections using SP142 and 22C3 antibodies. A total of 82% of the sixty-six patients were hormone receptor positive and 8% of cases were triple negative (TN), while 10% showed human epidermal growth factor receptor 2 (HER2) amplification. sTILs (≥1%) were present in 64% of the study population. Using the SP142 antibody, 36% of tumors demonstrated a positive PD-L1 score of ≥1%, and using the 22C3 antibody, 28% had a positive PD-L1 score of ≥1. There was no correlation between sTILs or PD-L1 expression and tumor size, tumor grade, nodal status, expression of estrogen receptor (ER), or amplification of HER2. Our data did not show any difference in survival between the three molecular subtypes of pILC with respect to sTILs and PD-L1 expression. This study shows that pILCs show some degree of sTILs and PD-L1 expression; however, this was not associated with a survival improvement. Additional large trials are needed to understand immune infiltration in lobular cancer, especially in the pleomorphic subtype.

Abstract P4-01-02: Results from a dose escalation phase 1b study of palbociclib and avelumab in advanced breast cancer in the PAveMenT Trial

Abstract Background CDK4/6 inhibitors may trigger anti-tumour immunity, both through tumour cell intrinsic and extrinsic mechanisms, which can be enhanced by immune checkpoint blockade in pre-clinical studies. Although most triple negative breast cancers (TNBC) are resistant to CDK4/6 inhibition, palbociclib (palbo) has activity in pre-clinical models of the luminal androgen receptor (LAR) subtype. Here we report the dose finding phase Ib cohort A of the PAveMenT trial combining avelumab and palbociclib Methods The PAveMenT phase 1b trial consists of a dose finding phase (cohort A), to be followed by an expansion in androgen receptor positive (AR+) TNBC (Cohort B). The phase 1B phase investigated palbo dosing schedules (75mg intermittent dosing (ID), 100mg continuous dosing (CD) and 125mg ID) in combination with the anti-PD-L1 antibody, avelumab 10mg/kg 2-weekly (cohort A) to determine the optimal schedule for cohort B. The CD dose-level was pre-specified as the preferred schedule if tolerated, due to the proliferative nature of TNBC and potential for tumour re-growth with ID. Patients with previously treated advanced breast cancer (ABC) of any histology with measurable disease and adequate organ function, who had not received prior immunotherapy or CDK4/6 inhibitors or more than 2 lines of chemotherapy for ABC, were eligible for cohort A. Dose-limiting toxicities (DLTs) were defined as Grade(G) 4 neutropenia, complicated G3 neutropenia or G4 thrombocytopenia, or ≥G3 immune toxicity during cycle 1. AR staining of ≥10% using the SP107 antibody (Ventana) was considered AR+. PD-L1 was evaluated using CC23 Pharma Dx antibody (DAKO) and PD-L1 positive defined as CPS score of ≥10%. Results Fourteen patients were recruited to Part A, all female, 12 with TNBC, two with ER positive/HER2 negative ABC, median age 53.5 years. Three evaluable patients received palbo 75mg/day ID without DLTs, therefore two further dose levels were opened in parallel. Three evaluable patients received palbo 125mg/day ID without DLTs. Due to a DLT of fever with G3 neutropenia in one of 3 patients receiving palbo 100mg CD, 3 further evaluable patients received 100mg CD without DLTs. Two patients were not evaluable for DLTs having not received a full cycle of treatment so were replaced. One case of grade 3 colitis and one case of grade 2 hypothyroidism (both 100mg CD) occurred outside the DLT period. Amongst 12 patients with TNBC, metastatic tissue was available for 11, archival breast tissue for one. Four of 12 patients were AR+. Five of 10 TNBC patients with sufficient tissue were PD-L1 positive, all of whom had previously tested negative with the SP142 antibody (Ventana). One of the 5 PD-L1+ and 2/5 PD-L1 negative TNBC cases were also AR+. TNBC patients received a median of 2 cycles of treatment (range 1-17). Both ER+ patients were AR+ and PD-L1 negative; one progressed after 4 cycles, the other remains on treatment at 12 cycles. The 100mg CD schedule was selected for cohort B. Conclusions All schedules were tolerable, with activity observed in patients with both ER positive and TNBC cancers. Cohort B of the study is open to recruitment for patients with AR+ TNBC. NCT04360941. Citation Format: Alicia F. Okines, Houman Moghadam, Laura Sparks, Kabir Mohammed, Kathryn Dunne, Ashutosh Nerurkar, Peter Osin, Claire Swift, Ruth Sardinha, Nicholas Turner. Results from a dose escalation phase 1b study of palbociclib and avelumab in advanced breast cancer in the PAveMenT Trial [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P4-01-02.

Programmed Death Ligand-1 Combined Positive Score Concordance and Interrater Reliability in Primary Tumors and Synchronous Lymph Node Metastases in Resected Cases of p16+ Oropharyngeal Squamous Cell Carcinoma.

Pembrolizumab is used in patients with metastatic head and neck squamous cell carcinoma contingent upon the programmed death ligand-1 (PD-L1) combined positive score (CPS). To compare PD-L1 CPS scores derived from paired resected primary tumors (PTs) and lymph node metastases (LMs) in patients with p16+ oropharyngeal squamous cell carcinoma (OPSCC). We identified 38 resected p16+ OPSCCs for which paired PTs and LMs were available. PD-L1 immunohistochemistry using the SP263 antibody clone was done on both the PT and the LM. CPS scoring was performed by 4 observers, and data were analyzed at the CPS cut points of greater than or equal to 1 and 20 in regard to interobserver and interspecimen agreement. Overall agreement between consensus CPS scoring of PT and LM was seen in 76% of paired specimens (κ = 0.53). No specimen received a negative consensus score. Interobserver agreement for both PT and LM was fair to substantial (κ = 0.54 and 0.51, respectively) and was inferior to that seen in a prospective series of unselected head and neck squamous carcinoma cases evaluated at our institution (κ = 0.84). Given the high rates of interobserver and interspecimen variability, evaluation of additional material or by additional observers may be of value in performing CPS scoring in cases of p16+ OPSCC. This is particularly the case when a negative or low-positive result is being evaluated in a patient who is otherwise a good candidate for immunotherapy.

P76-1 Comparison of programmed death ligand-1 expression in NSCLC using 22C3 and SP142 antibodies in a single institute study

The programmed death ligand-1 (PD-L1) expression is an important biomarker in the era of immunotherapy using immune checkpoint inhibitors. PD-L1 testing using the 22C3 antibody (Ab) has been the standard in determining the first-line therapy in advanced non-small cell lung cancer (NSCLC). Atezolizumab monotherapy has become a first-line treatment option based on the IMpower 110 trial result. However, there are few data on the effectiveness of PD-L1 staining using SP142 Ab.

Significance of the effects of chemotherapy on programmed death-ligand 1 expression in triple-negative breast cancer.

Atezolizumab has been approved as an antibody against programmed death-ligand 1 (PD-L1)-positive immune cells in patients with advanced or recurrent triple-negative breast cancer. However, the optimal timing to examine PD-L1 expression remains controversial. We retrospectively researched PD-L1 positivity rates in biopsy, surgical and recurrent specimens from patients with triple-negative breast cancer treated with neoadjuvant chemotherapy. We also examined alterations in PD-L1 and their meaning. In total, 35 triple-negative breast cancer biopsy specimens obtained before neoadjuvant chemotherapy, 20 corresponding specimens obtained after neoadjuvant chemotherapy and 5 corresponding recurrent specimens were obtained. We examined PD-L1 immunohistochemistry on tumor cells and tumor-infiltrating immune cells using SP142 antibody. In comparison with specimens obtained before neoadjuvant chemotherapy, PD-L1 expression randomly changed in immune cells after neoadjuvant chemotherapy, but PD-L1 expression was significantly reduced in tumor cells. Pre-neoadjuvant chemotherapy specimens with low PD-L1 expression (PD-L1 scores of ≤1 for both immune cells and tumor cells) were linked to better disease-free survival (P<0.001) and overall survival (P<0.001) than the other specimens. This is the first study to evaluate PD-L1 expression both before and after chemotherapy in breast cancer and examine its relationship with prognosis. The results suggest that the PD-L1 level may be useful for predicting the prognosis of patients with triple-negative breast cancer who do not have pathological complete responses to neoadjuvant chemotherapy.

Atlas of PD-L1 for Pathologists: Indications, Scores, Diagnostic Platforms and Reporting Systems.

Background. Innovative drugs targeting the PD1/PD-L1 axis have opened promising scenarios in modern cancer therapy. Plenty of assays and scoring systems have been developed for the evaluation of PD-L1 immunohistochemical expression, so far considered the most reliable therapeutic predictive marker. Methods. By gathering the opinion of acknowledged experts in dedicated fields of pathology, we sought to update the currently available evidence on PD-L1 assessment in various types of tumors. Results. Robust data were progressively collected for several anatomic districts and leading international agencies to approve specific protocols: among these, TPS with 22C3, SP142 and SP263 clones in lung cancer; IC with SP142 antibody in breast, lung and urothelial tumors; and CPS with 22C3/SP263 assays in head and neck and urothelial carcinomas. On the other hand, for other malignancies, such as gastroenteric neoplasms, immunotherapy has been only recently introduced, often for particular histotypes, so specific guidelines are still lacking. Conclusions. PD-L1 immunohistochemical scoring is currently the basis for allowing many cancer patients to receive properly targeted therapies. While protocols supported by proven data are already available for many tumors, dedicated studies and clinical trials focusing on harmonization of the topic in other still only partially explored fields are surely yet advisable.

Discordant genomic correlates of PD-L1 expression in lung adenocarcinoma among multiple cohorts using dissimilar PD-L1 testing techniques.

e20526 Background: Programmed death-ligand 1 (PD-L1) is a crucial biomarker predicting efficacious treatment of immune checkpoint inhibitors (ICIs). Previous studies identified distinct staining utility among multiple PD-L1 antibodies, however, whether the genomic correlate of stained PD-L1 is also impacted by different testing assays is largely unknown. Methods: Here, we analyzed 873 lung adenocarcinoma (LUAD) samples by the FDA-approved SP263 and 22C3 antibodies, and discovered that TP53, KRAS, MET, and ALK alterations associated with high SP263/22C3-PD-L1 expression, while EGFR and ERBB2 aberrations linked to SP263/22C3-PD-L1 negativity. Results: We retrieved the data of 873 LUAD patients whose PD-L1 (SP263/22C3) and mutational data. The SP263/22C3-PD-L1 expression was significantly correlated with tumor mutational burden and was greater in metastatic lymph node than lung lesion and distal metastasis. Furthermore, TP53, MET, ALK, KRAS alterations were associated with higher SP263/22C3-PD-L1 expression, while EGFR and ERBB2 alterations were linked to SP263/22C3-PD-L1 negativity. In the TCGA database lacking fusion data except for ALK and using reverse phase protein arrays (RPPA) for quantifying PD-L1 expression, STK11 mutation exhibited negative correlation with PD-L1 mRNA (p &lt; 0.001), but not the RPPA-PD-L1 protein-level expression (p &gt; 0.10), while samples with ALK fusion showed a tendency of higher RPPA-PD-L1 (p = 0.088). Moreover, in the OAK trial providing data of single nucleotide variation and using SP142 (Ventana, Tucson, AZ) assay evaluating both tumor and immune cells, weak correlation between STK11 mutation and SP142-PD-L1 negativity was observed (p = 0.076). Conclusions: Identical usage of staining antibody is the fundament of further comparisons and deeper investigations. The FDA-approved SP263 and 22C3 assays with fixed reagent, machine and protocol may perform more effective and repeatable than LDTs such as the E1L3N method. In addition to the different staining utility among PD-L1 antibodies indicated by previous studies, we further provide insights into the dissimilar genomic correlates of stained PD-L1 by using different PD-L1 quantifying techniques.

PCR-based analysis of PD-L1 RNA expression in lung cancer: comparison with commonly used immunohistochemical assays

BackgroundPD-L1 testing is currently performed by immunohistochemistry (IHC). We questioned whether the results of PCR-based measurement of PD-L1 RNA expression correlate with IHC scores obtained by different commercial assays. Materials and methods167 consecutive non-squamous non-small cell lung carcinomas (NSCLCs) were analyzed for PD-L1 RNA expression and 22C3, SP263, and SP142 IHC scoring using recommended cut-offs. RNA expression was divided into low, moderate, and high categories. ResultsRNA and protein expression demonstrated moderate correlation as continuous variables. Using prespecified RNA cut-offs, PCR testing showed a high negative predictive value towards the IHC analysis: the share of PD-L1 protein-negative tumors among cases classified as PD-L1-low by the PCR test reached 92–99% for all three antibodies. Meanwhile, about half of cases with moderate to high PD-L1 RNA expression had IHC staining in less than 1% tumor cells as determined by 22C3 or SP263 antibodies. Among the 51 discordant cases, which had <1% tumor staining by both 22C3 and SP263 clones but high RNA level, 29 (57%) showed ≥1% positive immune cells by SP263 and/or 22C3, 14 cases (27%) had detectable IHC expression in 0.1–0.9% tumor or immune cells by SP263 and/or 22C3, and 8 (16%) were entirely negative by IHC. ConclusionSome NSCLCs demonstrate readily detectable PD-L1 expression on the level of RNA, but fall below commonly accepted cut-offs by IHC. It remains to be studied whether these discrepancies are attributed to technical or biological reasons. Clinical sensitivity of these tumors to immune therapy deserves additional investigations.