bluuebunny/crossref_metadata_2025 huggingface dataset:
PD-1 inhibitor combined with radiotherapy and GM-CSF as salvage therapy in patients with chemotherapy-refractory metastatic solid tumors.
Authors: Kong Yuehong , Zhao Xiangrong , Zou Li , Xing Pengfei , Ma Yifu , Tian Ye , Zhang Liyuan
Published: May 2020

e15173

Background: Patients with refractory metastatic solid tumors have poor prognoses and few therapeutic options. The application of PD-1 inhibitor has led to a paradigm shift in the treatment of advanced cancer, but its efficacy as a monotherapy is limited. Previous evidence has shown that the anti-tumor effects might be reinforced when PD-1 inhibitor combines with radiotherapy or granulocyte macrophage-colony stimulating factor (GM-CSF). An exploratory study was conducted to assess the clinical efficacy and safety of PD-1 inhibitor combined with radiotherapy and GM-CSF (PRaG regimen) for the treatment of refractory patients. Methods: Participants had multimetastatic solid tumors progressing beyond at least first-line chemotherapy.They were treated with hyperfractionated radiotherapy (3 doses of 8Gy or 5 doses of 5Gy) for each metastatic site. On the second day after radiotherapy, PD-1 inhibitor 200 mg once was intravenously administered and GM-CSF 200 µg daily was subcutaneously injected for 2 weeks. This course was repeated every 3 weeks, targeting a second metastatic site. Triple-combination therapy. (PRaG regimen) was given for at least 2 cycles. After combination therapy, maintenance with PD-1 inhibitor was administered until disease progression or unacceptable toxicity. The main outcome measures included safety, toxicity, progression-free survival (PFS) and objective response rate (ORR) after three cycles of treatment. Results: A total of 16 patients were enrolled.The median number of metastatic lesions was 7.5(95%CI, 5.3 to 19.5) and the median sum of the longest diameter of all measurable lesions was 161.7mm (95%CI, 104.1 to 279.3mm).All patients completed two cycles or more of triple-combination therapy. The ORR was 20% and the median PFS was 3.3 months (95%CI, 2.3to 7.2months) at the time of evaluation. Treatment-related adverse events of any grade occurred in 13 (81%) patients, and grade 3 and higher adverse event like pneumonitis occurred in 2 (11.0%) patients. Conclusions: The PRaG regimen is well tolerated with acceptable toxicity. For patients with resistant metastatic solid tumors, low efficacy and high cytotoxicity are commonly observed in conventional therapy. With the new chemo-free PRaG regimen, myelosuppression was not observed and gastrointestinal side effects remained low. So, PRaG regimen is considered as a promising salvage treatment for patients with chemotherapy-refractory metastatic solid tumor. The suggested mechanism might involve radiosensitization of immunotherapy. Clinical trial information: ChiCTR1900020175 .
Robust prediction of immune checkpoint inhibition therapy for non-small cell lung cancer.
Authors: Zhang Yiqun , Zhao Guodong , He Ning , Wang Lingyu , Wang Jianfei , Jin Ge , Lin Rongbo , Yang Yadong
Published: May 2020

e15192

Background: Immune checkpoint inhibition (ICI) has been validated and applied in advanced lung cancer patients. However Oonly part of patients response to ICI, thus optimizing ICI would be emergent. Based on public non-small cell lung cancer (NSCLC) cohorts’ data, we expected to build a multivariable model which would improve the efficacy of predicting ICIs response. Methods: 2 NSCLC and 1 pan-cancer cohorts treated with ICIs were collected from cBioPortal (luad_mskcc_2015, n = 203; nsclc_pd1_msk_2018, n = 33; tmb_mskcc_2018, n = 1661, pan-cancer). Another NSCLC cohort (cancer_cell_2018) was collected from MD. Hellmann’s study (Cancer Cell, 2018, n = 75). We incorporated features including tumor mutation burden (TMB), PD-L1 expression, Mutant-Allele Tumor Heterogeneity (MATH), and mutational status of cancer-related pathways. And we used Bayesian Regularization for Feed-Forward Neural Networks ( brnn) method to fit models of predicting clinical response. Results: The correlation analysis showed that MATH was independent of TMB (R = 0.141, p = 0.01252) and PD-L1 expression (p = 0.432, ANOVA test among high, median and low PD-L1 expression group) in 3 merged NSCLC cohorts. We randomly selected 70% of cancer_cell_2018 cohort as training set, and the AUC improved from 0.5702/0.5191/0.6728 to 0.6893/0.527/0.8554 (in cancer_cell_2018 validation set, luad_mskcc_2015, and nsclc_pd1_msk_2018 cohort, respectively) after incorporating MATH to TMB + PD-L1 model. Additionally, we identified that mutational status of DDR, HRR, MAPK, Wnt, JAK, PI3K, and NFKB pathway was significantly associated with overall survival in tmb_mskcc_2018 cohort. After incorporating the mutation status of these pathways, the model exhibited further improvement in prediction efficacy (AUC reached 0.8595/0.5634/0.7831). When using luad_mskcc_2015 cohort as training set, the model performance improvement was also observed after joining pathway mutation information into TMB + PD-L1 + MATH model (AUC improved from 0.5742/0.6339 to 0.7305/0.7 in predicting luad_mskcc_2015 and cancer_cell_2018 cohort). Conclusions: We built a multivariable model with robust performance in predicting ICI response in independent NSCLC cohorts. With the accumulation of larger datasets, further studies are warranted to refine the predictive performance of the approach.
CD8A and HAPLN3 expression profiling to reveal an immunologic subtype of bladder cancer with favorable survival.
Authors: Ma Jingjiao , He Ning , Wang Jianfei , Wang Lingyu , Jin Ge , Lin Rongbo , Yang Yadong
Published: May 2020

e15193

Background: At present, immune checkpoint inhibitors (ICI) have demonstrated robust anti-tumor activity with a favorable safety in bladder cancer (BLCA). However, less than half of BLCA patients respond to immunotherapy. We aimed to further delineate the immunologic subtype of BLCA to pick patients who might benefit from ICI treatment. Methods: The RNA-seq and clinical data were collected from The Cancer Genome Atlas (TCGA) data portal (n = 408) and Gene Expression Omnibus (GEO) (GSE48075, n = 142; GSE32894, n = 308). TCGA and GSE48075 cohort are the discovery datasets, and GSE32894 cohort is used for validation. According to the expression of every specific gene ( Gi), patients were divided into two groups using median FPKM as cutpoint value, namely Gi_high and Gi_low, respectively. Combined with CD8A expression, all the patients were divided to four groups as follows: CD8A_high/ Gi_high, CD8A_high/ Gi_low, CD8A_low/ Gi_high, and CD8A_low/ Gi_low. Then we calculated the pearson correlation coefficients between Gi and 13 immune checkpoint genes ( CD274, IDO1, CTLA4, LAG3, CD276, VTCN1, CD70, HAVCR2, CD40, CD47, TNFRSF18, TIGIT, and TNFSF14). Overall or disease free survival of the four groups were compared using the survfit function from the survival R package and P values from log-rank tests were reported. Results: In the discovery cohort, we found CD8A_high/ HAPLN3_low group showed the best overall survival, whereas the CD8A_low/ HAPLN3_high showed the worst (TCGA, p = 0.013; GSE48075, p = 0.012). Additionally, HAPLN3 was positively correlated with 5/13 immune-checkpoint genes: CD274, IDO1, CTLA4, LAG3, HAVCR2 (r > 0.6, p < 0.01). And the same observations were extended in the validation cohort (GSE32894): CD8A_high/ HAPLN3_low group exhibited a favorable survival (p = 0.00059), and HAPLN3 had a strong correlation with the same 5 immune-checkpoint genes (r > 0.6, p < 0.01). Conclusions: The combination of high CD8A and low HAPLN3 expression identified a subtype of BLCA patients with favorable survival and these findings may further help refining the selection of BLCA patients for immunotherapy.
RNA pathway enrichment in serum-based mass spectroscopy prognostic analyses.
Authors: German Benjamin D , Borgia Jeffrey Allen , Naqib Ankur D , Bonomi Philip D. , Batus Marta , Basu Sanjib , Fidler Mary J.
Published: May 2020

e15197

Background: Veristrat(VS) good and poor labels were found to be prognostic in the front-line NSCLC setting with or without immune checkpoint inhibitors. Little is known about RNA expression data corresponding to these good and poor prognostic labels. Methods: Raw RNA seq expression data was obtained from TEMPUS XP RNA analysis. Sequences were filtered for allowing only genes that had an overall count of at least 20 across all samples. Samples were then processed through the EdgeR analysis in the R programming environment. Along with a Z-scored heatmap, PCA plot was also generated. Differential abundance analysis was done further filter out the genes. Benjamini-Hochberg correction was done and only genes with corrected p-value of < 0.05 were selected for further pathway analysis. 242 genes with corrected p-value < 0.05 were used. Pathway analysis was performed using the Broad Institute GSEA (Gene Set Enrichment Analysis) and DAVID (Database for Annotation, Visualization and Integrated Discovery). The gene list was ran using KEGG database in GSEA and using KEGG, BioCarta and Reactome pathway databases. Functional analyses against the KEGG and REACTOME database were reported out to determine the enriched pathways in the VS poor and VS good groups. Kaplan-Meier and Logrank were used to compare overall survival in the poor vs. good label cohorts of patients. Results: 26 patients had VS label and RNA expression data available. Nine patients had VS poor label and 17 had veristrat good label. 6(66%) VS poor 13(76%) VS good patients received anti PD1 agents during the course of their therapy. Overall survival was not significantly different between good and poor label groups in this small series. 242 genes had significantly different abundance in the VS poor vs good patients with the majority (211) over expressed in the poor label patients. Significant pathways enriched in the VS poor group included the intrinsic pathway of fibrin clot formation, platelet degranulation (DAVID), steroid hormone biosynthesis and dilated cardiomyopathy (KEGG). Systemic lupus erythematous pathway was enriched in the VS good group (KEGG). Conclusions: In this small set of front-line patients with RNA pathway analyses and known VS poor or good labels, differential expression of acute phase reactants was found in VS poor patients. DAVID and KEGG pathways were differentially enriched and may reveal potential targets to mitigate prognosis in poor risk NSCLC.
Bioinformatic profiling of tumor immunity from patient biopsies to predict survival and response to immunotherapy.
Authors: Mahmoud Yamil Damian , Veigas Florencia , Merlo Joaquin , Balzarini Monica , Rocha Dario , Rabinovich Gabriel Adrian , Fernandez Elmer , Girotti Maria Romina Romina
Published: May 2020

e15198

Background: Immunotherapies have revolutionized cancer treatment, but responses are not universal and patients who initially respond to therapy develop resistance. The accurate quantification of tumor-infiltrating immune cells holds the promise to reveal the role of the immune system in human cancers and its involvement in tumor escape mechanisms and response to therapy. We present MIXTURE, a new algorithm for tumor immune cell-type proportions deconvolution from transcriptomic data that overcomes competitive methods and revealed novel associations of immune cell types with patient survival and immunotherapy response. Methods: We applied MIXTURE to transcriptomic data from BRCA (n = 1095), LUAD (n = 506), SKCM (n = 472), HNSC (n = 499) and COAD (n = 521) cohorts from TCGA and five published datasets of melanoma patients treated with anti-PD-1/anti-CLTA-4. Results: The analysis of TCGA breast cancer biopsies showed high proportions of M2-macrophages associated with poor patient survival (p < 0.001). In contrast, we found that proportions of follicular T helper cells were associated with better outcome (p < 0.001). We observed a differential immune composition in biopsies of lung adenocarcinoma patients with mutations in TP53 (WT; n = 247; Mut; n = 259) and EGFR (WT, n = 438; Mut, n = 68) related with their different response to immunotherapy (p < 0.05). We found correlations between immune cells proportions and current biomarkers of response to immunotherapy such as TMB, intratumoral heterogeneity, MSI and PD-L1 expression in the TCGA cohorts (p < 0.05). The meta-analysis of melanoma patients treated with immunotherapy showed a distinct immune infiltrate in patients who responded to anti-PD-1 with an increase of immune effector cells such as CD8, CD4 memory activated and gamma-delta T cells, and a decrease in immunosuppressive M2-macrophages (p < 0.05; R = 81; NR = 107). According with the latest findings, we observed higher presence of B cells in responders to anti-PD-1 on-treatment (p = 0.033; R = 31; NR = 23) and in baseline of responders to anti-CTLA-4 (p = 0.028; R = 14; NR = 26). Interestingly, patients that previously progressed to anti-CTLA-4 showed a differential immune profile that was associated with response to anti-PD-1 (p < 0.05; Ipi-Prog = 59; Ipi-Naïve = 102). Conclusions: We demonstrated the potential of MIXTURE to understand the tumor immune microenvironment and its relationship with patient survival and response to immunotherapies. MIXTURE is available for the wider scientific community as web application and as packages for R and Python.
targetSCAPE and ultraSCAPE: Simultaneous identification and deep profiling of human antigen-specific T cells and other immune cell subsets by mass cytometry.
Authors: Abel Brian , Kairi Faris , Nardin Alessandra , Newell Evan , Fehlings Michael
Published: May 2020

e15204

Background: During clinical trial immune monitoring, especially in the field of immunotherapy, it is critical to collect in-depth phenotypic information from multiple immune cell populations in order to assess the biological activity of the immunotherapy, to identify biomarkers of response or disease progression, and/or to identify new drug targets. However, patient samples such as peripheral blood mononuclear cells or tissues, are often scarce and current methods face limitations in either achieving a sufficient depth of analysis and/or cell throughput. Methods: In order to identify therapy-relevant antigens and to facilitate a concurrent in-depth characterization of T cells directed towards these targets, immunoSCAPE leverages the high-dimensional immune profiling capabilities of mass cytometry and a unique methodology allowing the identification and characterization of rare antigen-specific T-cell subsets (targetSCAPE). By implementing a new cutting-edge technology that combines flow and mass cytometry in parallel with a combinatorial live cell barcoding strategy, we further increased the high-dimensional phenotyping capacities to over 100 different marker molecules on up to four different immune cell subsets simultaneously within the same sample. Results: We isolated 4 different immune cell populations from a single sample and combined 3 different phenotypic panels consisting of 35 makers each together with a combinatorial tetramer multiplex and phenotyping panel for deep profiling of myeloid cells, NK cells, B cells and T cells. We demonstrate the potential of this novel immuno-phenotyping method, by tracking virus-specific T cells while simultaneously characterizing 4 immune cell subsets with over 100 distinct phenotypic markers from a single sample, which is currently impossible employing modern flow cytometers or classical mass cytometry methods. Conclusions: With its ability to provide an unprecedented picture of the immune status within a single sample, including T cell specificity information and in-depth profiling of relevant immune cell subsets, ultraSCAPE in combination with targetSCAPE can provide detailed insights on the effects of immunotherapy on the immune cell population. Information learned from in-depth immune phenotyping of several immune cell subsets such as T, B, NK and myeloid cell subsets can be leveraged for the development of novel diagnostics, biomarker discovery and monitoring therapeutic strategies in immunotherapy clinical trials.
Balixafortide (a CXCR4 antagonist) plus eribulin in HER2 negative metastatic breast cancer: Dose-response analysis of efficacy from phase I single-arm trial.
Authors: Kaufman Peter A. , Pernas Simon Sonia , Martin Miguel , Gil-Martin Marta , Gomez Pardo Patricia , Lopez-Tarruella Sara , Manso Luis , Ciruelos Eva , Perez-Fidalgo Jose Alejandro , Hernando Cristina , Ademuyiwa Foluso Olabisi , Weilbaecher Katherine N. , Mayer Ingrid A. , Pluard Timothy J. , Martinez Garcia Maria , Vahdat Linda T. , Ringeisen Francois Philippe , Bobirca Alexandra , Cortes Javier
Published: May 2020

e15209

Background: Balixafortide (B) is a potent, selective antagonist of the chemokine receptor CXCR4. High CXCR4 levels correlate with aggressive metastatic phenotypes and poor prognosis in metastatic breast cancer (MBC). Efficacy and safety data were published recently from the Phase 1 trial investigating B + eribulin (E) in patients with HER2 negative MBC1. We report the final efficacy analyses from this trial, including assessment of dose-response. Methods: In this single-arm, dose escalation trial, patients (pts) received E + increasing doses of B using a 3+3 design in 3 parts: Part I (cohorts received low E doses); Part II (dose-escalation cohort for B [1−5.5mg/kg] + 1.4mg/m2 E); Expanded Cohort (EC; 5.5mg/kg B + 1.4mg/m2 E) to confirm safety and efficacy. Results: At entry, all 56 women (age range 33−82 years) were HER2-negative (IHC and/or FISH), CXCR4 positive. The majority were Caucasian. Most pts were heavily pre-treated in the metastatic setting (line of chemotherapy on study: 29% 2nd line, 50% 3rd line, 21% 4th line). 75% were hormone receptor positive and 23% had triple negative breast cancer. Conclusions: A consistent dose response effect for B + E was suggested in heavily pretreated pts with HER2 negative MBC across all efficacy endpoints. A comparison of these efficacy results, and particularly response data, with single agent data for E in similar populations2, 3 showed that pts in the EC had a more profound benefit observed consistently throughout all efficacy endpoints. Further data and analysis will be forthcoming for presentation. 1. 3 patients from Part II also included in EC because they received the B dose selected for EC (5.5mg/kg). 2. Part I was an initial safety run-in with lower E doses, and so is not included in the table. 1. Pernas S et al. Lancet Oncol. 2018; 19: 812−24 2. Cortes J et al. Lancet. 2011; 377: 914−923 3. Kaufman PA et al. J Clin Oncol. 2015; 33: 594−601. Clinical trial information: NCT01837095 . [Table: see text]
The landscape of chromosome 11q13 amplification in Chinese solid tumor patients and hyperprogressive disease (HPD) clinical example.
Authors: Yu Jieli , Wang Minghui , Song RongFeng , Li Shuben , Wang Hongbiao , Xie Jianjiang , Zhou Chengzhi , Li Kunsong , Chi Liangjie , Huang Jian , Shen Zhuojian , Gu Weiguang , Li Yun , LU Zhen-hai , Xue FangQin , Liu Dan , Shi Xiaoliang , Liu Angen , Zhang Lin
Published: May 2020

e15222

Background: Multiple biomarkers are thought to be effective guides in selecting immune checkpoint inhibitors, such as Tumor mutational burden (TMB), PD-L1 and MSI/dMMR. Immunotherapy may be miraculous effective in some patients but many other patients experienced poor prognosis and even tumor overgrowth after immunotherapy in real practice. Previous studies have reported that 11q13 (CCND1, FGF3, FGF4, and FGF19) amplifications were associated with HPD. However, the characterization of chromosome 11q13 amplification in Chinese solid tumor patients are not clear. Methods: A total of 10167 Chinese solid tumor patients’ FFPE and matching blood samples sequenced by next-generation sequencing (NGS) targeting 450 cancer genes were included in this study. Genomic alterations including single nucleotide variants (SNV), insertions and deletions, copy number variations and fusions were assessed. The testing was carried out by a College of American Pathologists (CAP) accredited and Clinical Laboratory Improvement Amendments (CLIA) certified laboratory. Results: Chromosome 11q13 amplifications were identified in 679 (6.7%) Chinese solid tumor patients. The top six tumor types with 11q13 amplification were esophageal carcinoma 43.8%, melanoma 17%, urothelial carcinoma 16%, breast carcinoma 13.4%, head and neck carcinoma 13.1% and hepatocellular carcinoma 9.4%. In 2257 patients with TMB-H (10muts/Mb) / PD-L1 expression / MSI samples, chromosome 11q13 copy number gain were identified in 184 (8.2%) patients. Chromosome 11q13 amplification and MSI were independent events and didn't occur simultaneously. In TMB-H patients, the detection rate of 11q13 amplification was 8.5%, while in PD-L1 positive patients, the amplification rate of 11q13 was 7.4%. In patients with TMB-H and PD-L1 positive, the detection rate of 11q13 amplification was 9.4%. In our clinical practice, we saw example of HPD exist in a lung squamous cell carcinoma patient who was treated with multiple lines of therapy and then used the Nivolumab. The patient underwent genetic testing and was found to be chromosome 11q13 amplified. Conclusions: The proportion of 11q13 amplification was different in different types of solid tumor. Esophageal carcinoma, melanoma and urothelial carcinoma were top three types of solid tumors with 11q13 amplification. The 11q13 amplification did not coincide with MSI but it overlapped with PD-L1 expression or TMB-H. Together, these results may provide immunotherapy guidance in clinical practice.
Efficacy and hepatotoxicity of Infliximab in resolving steroid-refractory immune-related adverse events (irAEs) in a real-world setting.
Authors: Pimentel Muniz Thiago , Araujo Daniel Vilarim , Yang Anjie , Sorotsky Hadas , Keshavarzi Sareh , Butler Marcus O. , Saibil Samuel , Spreafico Anna , Hogg David
Published: May 2020

e15227

Background: Infliximab is used to treat steroid-refractory immune-related adverse events (irAEs) induced by immune checkpoint inhibitors (ICI). Current dogma holds that infliximab is contraindicated in immune hepatitis; steroid refractory disease should be managed with mycophenolate mofetil (MMF). This is based on anecdotal reports of infliximab-induced hepatotoxicity in patients with rheumatologic diseases who received multiple infusions of the drug. In this series of 56 consecutive cases of infliximab-treated patients, we assessed the efficacy of infliximab in resolving specific steroid-refractory irAEs and evaluated the risk of hepatotoxicity from this drug. Methods: We reviewed consecutive patients treated with infliximab for steroid-refractory irAE at a tertiary cancer center between January 2010 and February 2019. To judge hepatotoxicity, we used the Wilcoxon signed rank test to compare the mean value of ALT, AST and total bilirubin (BT) between 0-4 weeks before and after infliximab therapy. We compared factors associated with infliximab efficacy by logistic regression. A p value < 0.05 was considered statistically significant. Results: Mean age was 61.9 years. The majority had a diagnosis of melanoma (62%); 45% had a combination of two ICI before toxicity onset, of whom 73% received an anti-PD1 and an anti-CTLA-4. Colitis was the most common toxicity (66%), followed by pneumonitis (11%). Infliximab was used to treat immune related hepatitis (ir-hepatitis) in 1 patient (2%). Median number of infliximab doses was 1 (1-3), with resolution of toxicity in 76% of patients. Colitis was likely to resolve with infliximab in comparison to other irAE [OR 0.2(95% CI 0.05-0.73)]. Univariate logistic regression did not demonstrate statistical difference in toxicity outcome when ICI were given either as monotherapy (p = 0.3) or combination (p = 0.97). Mean BT, ALT and AST levels before and after infliximab were not statistically different (see table), with no evidence of infliximab-induced hepatotoxicity. The patient treated for ir-hepatitis had a complete recovery, with no further hepatotoxicity. Conclusions: Infliximab was effective in treating irAE and was not associated with hepatotoxicity in this dose-limited setting. Infliximab may be an alternative to MMF in steroid-refractory ir-hepatitis and should be tested prospectively in a randomized trial. [Table: see text]
Key issues in the development and approval of new anti-cancer immune therapies.
Authors: Blunt Al , Gyorffy Steve , Messerschmidt Gerald L.
Published: May 2020

e15232

Background: Immunotherapy (IT) is now an established treatment option for a select group of cancer patients, and holds great promise for broader application. However, the number of new IT drug approvals has remained low, despite hundreds of new immune therapies entering clinical trials. Methods: We examined both successes and failures of immune-therapeutics (small molecules, large molecules and cellular therapies) to develop insight into this trend. We searched multiple public data sources including clinicaltrials.gov, PubMed, and other large data bases to identify informative trials and primary endpoints. A key area of our research focus was the rate of objective tumor responses following treatment with the immunotherapy agents. Results: Our main observation was that most clinical trials in solid tumors were characterized by a low rate of objective response. Tumor shrinkage is known to be the final manifestation of a multi-step process that begins with the therapeutic product’s ability to bind to its target, and the subsequent engagement of pathways that result in tumor infiltration and tumor cell destruction. A critical step is a sustained immune effector cell attack on the tumor cells. However, a growing body of evidence supports the concept that both non-cellular and cellular immune therapies are active primarily in the setting of a “very warm” or “hot” tumor microenvironment (TME). We found evidence of a positive correlation between objective responses and the degree of ‘hotness’ of the TME. Importantly, our research indicates that study drugs were typically successful in precisely engaging their targets in the associated pathway. High response appears to require selection of appropriate populations with respect to tissue/organism level pathways that lead to successful tumor destruction. Conclusions: The success of IT drugs requires clinical trial designs based on insight into both the study drug’s target pathway, and the impact of key organism level physiologic processes, (e.g., TME) that interfere with optimal anti-tumor activity. [Table: see text]
Lack of association of immune checkpoint protein expression with T stage or stage grouping in colorectal cancer (CRC): A tissue microarray (TMA) based analysis of clinical data.
Authors: Levy Rachel , Acuna-Villaorduna Ana , Maitra Radhashree , Goel Sanjay
Published: May 2020

e15241

Background: Immunotherapy is a valuable therapeutic intervention in multiple cancer types. In CRC, it is currently limited to those with microsatellite instability–high expression. Thus, immunotyping of CRC patients is critical to identifying potential targets for drug development. In this study, we profiled 171 CRC patient tumor samples to assess the expression of three immune check-point regulators (HHLA2, B7H3, and PD-L1), and 3 clinically useful biomarkers (CK7, CK20, and CDX2) and their association with T stage and stage grouping. Methods: TMA was created from 171 samples of CRC diagnosed between 2000 and 2016 in a major urban cancer center. Formalin fixed paraffin embedded (FFPE) tissues were used for immunohistochemistry (IHC). Antibodies specific for IHC staining were used for HHLA2, B7H3, PD-L1, CK7, CK20, and CDX2. Each specimen was scored from 4-12, based on multiplication of tumor tissue staining value (1 if 1-25%, 2 if 26-50%, 3 if 51-75%, and 4 if 76-100% of area visualized) by the intensity (1-3X). Expression of the proteins was sub divided into two or three groups, depending on the mean expression. Association between the six proteins and pathologic features including T stage and stage grouping (based on the AJCC stage at time of diagnosis) was assessed using chi-square or Fisher Exact tests. Results: Of the 171 patients, 62 (36.3%) presented with a primary cancer that recurred at a later date, and 109 (63.7%) had metastatic disease at presentation. Twenty patients were excluded from statistical analysis due to lack of staging data. The % of tumors with positive expression was 38.1 (HHLA), 37.6 (B7H3), 54.7 (PD-L1), 20.9 (CK7), 52.5 (CK20), and 58.2 (CDX2). There was no significant association between expression of each of the 6 proteins (HHLA2, B7H3, PD-L1, CK7, CK20, and CDX2) and the T stage (T1-2, T3 or T4) or stage grouping (stage 1-2 or stage 3). Consistent with clinical expectation, the expression of CK20 was higher than CK7 (p < 0.05). Conclusions: Expression of HHLA2, B7H3, PD-L1, CK7, CK20, and CDX2 were not associated with T stage or stage grouping of CRC samples in our study. These data suggest that these proteins are expressed early in the onset of CRC. Studies to determine their predictive and prognostic implications are underway.
Single cell dissociation from mouse tumor samples and analysis on BD FACSLyric Flow Cytometer.
Authors: Huang Wei , Pereira Nate , Neuhoefer Patrick Thomas , Middlebrook Aaron , Ghanekar Smita
Published: May 2020

e15245

Background: Single cell multiomics analysis of tumor cells and tumor infiltrating immune cells using high parameter flow cytometry and single cell genomics technologies is a powerful approach in the investigation of tumor biology and in the discovery of new drug targets and biomarkers. The preparation of single cells from solid tumor samples often involves a physical dissociation step in combination with an enzymatic digestion step. Methods: The effect of the tumor cell dissociation process on the phenotypic properties of single cells isolated from solid tumor samples were analyzed using a 12-color reagent panel on the BD FACSLyric flow cytometer. Specifically, spontaneous pancreatic tumor samples from genetically modified mice were processed using a manual dissociation process with a scalpel blade as well as an integrated tumor sample dissociation process with the prototype Singulator instrument with various protocol settings. The BD Horizon Dri Tumor & Tissue Dissociation Reagent (TTDR) reagent was used for the enzymatic step in all sample preparations. The yield, viability, and frequency of tumor cells and different subsets of leukocytes were measured for each tumor-derived single cell sample preparation. On the Singulator system, cell yield reached a plateau with highest viability at the 30-min process time and a higher percentage of tumor cells was obtained compared to the manual sample processing method. Results: The relative frequencies of T cells, B cells, myeloid cells and macrophages also changed in different sample preparation methods. However, the spatial heterogeneity in the tumor section and cell composition may contribute to these differences and would require more in-depth analysis. In summary, these results suggest that there are many variables in the tumor sample single cell dissociation process that could impact the downstream study results, particularly for multiomics studies. Conclusions: This study highlights the need for optimization and standardization of the sample processing methodologies for downstream analysis. Disclaimers: For Research Use Only. Not for use in diagnostic or therapeutic procedures. Class 1 Laser Product. BD, the BD Logo, FACSLyric, and Horizon are trademarks of Becton, Dickinson and Company. © 2020 BD and its subsidiaries. All rights reserved.
Identifying biomarkers to guide immunotherapy treatment of cancer of unknown primary.
Authors: Mileshkin Linda R. , Posner Atara , Pattison Andrew , Balachander Shiva , Etemadmoghadam Dariush , Fellowes Andrew , Wong Hui Li , Dow Eryn , deFazio Anna , Gao Bo , Karapetis Christos Stelios , Singh Madhu Sudan , Collins Ian M. , Steer Christopher , Warren Mark Andrew , Karanth Narayan , Bowtell David , Prall Owen W.J. , Tothill Richard
Published: May 2020

e15252

Background: Cancer of unknown primary (CUP) is a metastatic cancer where the primary tissue of origin (ToO) evades detection despite extensive clinical investigation. Recent approaches to the management of CUP have involved gene-expression profiling to resolve the likely ToO and mutational profiling to identify targeted treatments. Current evidence is that both of these methods can have some clinical benefit but only in a minority of cases. Immunotherapy may also be effective in CUP, but clinical trials looking at this are yet to report and the best way to select patients who may benefit is uncertain. Methods: Through a national cohort study called Solving Unknown Primary Cancer (SUPER), 245 CUP patients had molecular profiling performed and clinical follow-up. Variant detection from DNA sequencing using a targeted panel of 386 genes (SureSelect Agilent) was used to calculate tumour mutation burden (TMB). Additionally, a custom gene expression assay including 225 genes associated with ToO and 35 immune genes (SUPERDx, nCounter NanoString) was used to classify the likely ToO and immune profile the tumours. We used the genomic data from 217 of these patients to explore interdependencies between immune gene-expression patterns and TMB. We also retrospectively reviewed the outcomes of patients treated with anti-PD1/PD-L1 immune checkpoint inhibition (ICI) therapy in whom genomic data was available. Results: TMB was weakly correlated with a gene-expression score for immune response (IR) based on averaged z-score values of six genes (CD8A, IFNG, PRF1, PD-L1, GZMA, GZMB) (Spearman rho = 0.21) with many outliers exhibiting low TMB but high IR scores. Among twenty patients treated with ICIs, with both molecular data and response data available, 6 had a partial response (PR), 5 had stable disease (SD) and 9 had progressive disease (PD). We found that an elevated IR score was more predictive of ICI response than TMB. Most patients with either PR or SD to ICI treatment had a high IR score (9/10), and most patients with PD had low IR scores. However, only 3/10 patients with TMB available and a PR or SD had a high TMB ( > 10). Conclusions: Our results demonstrate that ICI therapy may benefit some CUP patients and that an immune response gene-expression scoremay predict ICI treatment response.
Immune- and tumor-intrinsic gene expression profiles of response or resistance to tislelizumab as monotherapy or in combination with chemotherapy in non-small cell lung cancer (NSCLC).
Authors: Desai Jayesh , Wang Jie , Zhou Qing , Zhao Jun , Deva Sanjeev , Tan Wei , Ma Xiaopeng , Zhang Yun , Shen Zhirong , Wu Xikun , Leaw Shiangjiin , Wu Yanjie , Wu Yi-Long
Published: May 2020

e15258

Background: Tislelizumab, an anti-PD-1 monoclonal antibody, showed clinical benefit for patients (pts) with NSCLC alone (NCT02407990, CTR20160872) and in combination with chemotherapy (NCT03432598). Gene expression profiles (GEP) associated with response and resistance to tislelizumab in these studies were assessed. Methods: The GEP of baseline tumor samples from 59 nonsquamous (NSQ) and 42 squamous (SQ) NSCLC pts treated with tislelizumab monotherapy (mono) as ≥1L treatment, and 16 NSQ and 21 SQ pts treated with tislelizumab plus chemotherapy (combo) as 1L treatment were assessed using the 1392-gene HTG GEP EdgeSeq panel. NSQ and SQ cohorts were analyzed separately due to distinct GEP features shown by PCA and t-SNE clustering. Results: Tislelizumab mono and combo showed antitumor activity in NSCLC (Table). In 80 biomarker-evaluable samples, inflamed tumor signatures (inflammatory GEP; antigen presentation GEP) were associated with longer overall survival (log-rank test, NSQ mono: P=0.04, 0.003; NSQ combo: P=0.05, 0.02; SQ combo: P=0.06, 0.06). Monotherapy non-responders (NRs) were clustered into 2 subgroups (NR1, NR2) with distinct GEPs. Compared with responders, NR1 had proliferation signatures (elevated cell cycle [CC] and DNA repair) in both NSQ ( P=0.2, 0.02) and SQ ( P=0.03, 0.4) cohorts, trending toward low inflamed tumor signatures. In NR pts receiving combo, CC and DNA repair signatures were not enriched, and high CC and DNA repair scores were observed in some SQ combo responders versus NRs ( P=0.2, 0.02). NR2 had higher M2 macrophage and Treg cell signatures versus responders in both NSQ and SQ mono, despite high inflamed tumor and low proliferation signatures. NR2 also had increased expression of genes related to immune regulation and angiogenesis, including PIK3CD, CCR2, CD244, IRAK3, and MAP4K1 ( P<0.05) in NSQ, and PIK3CD, CCR2, CD40, CD163, MMP12, VEGFC, and TEK ( P<0.05) in SQ. Conclusions: Clinical benefit in pts with NSCLC receiving tislelizumab (mono or combo) was associated with high inflamed tumor signatures, while elevated immune suppressive cell signatures may indicate resistance. High proliferation signatures were associated with resistance to monotherapy, but not to combination therapy. Both immune- and tumor-intrinsic factors may be considered for validation in future clinical trials. [Table: see text]
PD-L1 expression landscape and its relationship with tumor mutation burden in Chinese cancer patients.
Authors: Yang Haihua , Cui Longgang , Zhang Yuzi , Zhao Zhengyi , Bai Yuezong , Xie Wenzhuan
Published: May 2020

e15267

Background: Little is known about the pan-cancer PD-L1 expression landscape in Chinese patients although PD-L1 expression has been approved by FDA as a diagnosis for anti-PD-(L)1 therapy in several types of cancer. We did a cross-sectional analysis to assess the PD-L1 expression landscape in Chinese patients and its relationship with Tumor mutation burden (TMB). Methods: Tissue samples were collected from more than 8,000 consecutive cases in China between January, 2017, and August, 2019 and were analyzed by 3D Medicines, a College of American Pathologists (CAP)-accredited and Clinical Laboratory Improvement Amendments (CLIA)-certified laboratory. The method for NGS sequencing and tumor mutational burden (TMB) measurement were described previously. Clinical data and PD-L1 expression profiles were obtained from 8,063 patients whose tissue samples assed quality control. IHC staining for PD-L1 expression was performed using PD-L1 IHC 22C3 pharmDx assay (Dako North America, Carpentaria, CA, U.S.) or Ventana PD-L1 SP263 assay (Ventana Medical Systems, Tucson, AZ, U.S.). PD-L1 expression was determined using Tumor Proportion Score (TPS), the percentage of viable tumor cells stained. Results: PD-L1 expression was examined for 8,063 tissue samples collected from more than 18 different types of solid tumors. There were 4,866 (60%) male and 3,197 (40%) female patients. Their median age was 59 (IQR range, 50-66) years. Given the significance of different cut-points of PD-L1 expression in predicting clinical outcomes, expression levels of PD-L1 were arranged into the following intervals: < 1%, 1%-5%, 5%-50% and ≥50% for each cancer type. Small cell lung cancer (SCLC) had the lowest and Squamous Carcinoma of Head and Neck (HNSC) had the highest levels of PD-L1 expression. Spearman correlation analysis indicated no correlation between PD-L1 and tumor mutational burden (TMB) for Chinese cancer patients (R = 0.1, P < 0.01), which is in line with the previous reports that PD-L1 and TMB were two independent predictors in immunotherapy. Conclusions: The landscape of PD-L1 expression among Chinese cancer population in this study will further assist the utilization of PD-L1 as a predictive biomarker in clinical practice.
Real-world observation of the efficacy and prognostic factors analysis of apatinib for patients with advanced gastric cancer.
Authors: Shen Qiwen , WANG Mingyun , Peng Xiaobao , Tan Siyi , Xie Jiaqi , Du Juan , Liu Qin , Wei Jia , Liu Baorui , Yang Yang
Published: May 2020

e15508

Background: Apatinib is the first oral anti-angiogenic agent approved for the treatment of advanced gastric cancer in China. With the development of marketing promotion, the real world observation of the efficacy and prognostic factors analysis of apatinib become the urgent need for clinical decision. The aim of this study is to evaluate the efficacy and safety of apatinib in patients with advanced gastric cancer, and the association between some prognostic factors and clinical outcomes. Methods: We collected the data of 153 patients with advanced gastric cancer, who were treated with apatinib after failure of at least one regimen chemotherapy from December 2014 to June 2019. Treatment response, progression-free survival(PFS), overall survival(OS) and treatment-related adverse events were evaluated. Results: The median PFS for 153 patients was 3.1m (95% confidence interval, CI 2.863-3.337). The median OS was 6.0m (95% CI 4.392-7.608). Adverse events of grade 3 or above happened in 47.6 percent of patients and 61.7 percent of whom stopped treatment. Multivariate analysis showed that the number of metastatic sites was the independent factor simultaneously predicting DCR (disease control rate, P = 0.008) and OS (P = 0.007). Patients spending more than one year from diagnosis to treatment with apatinib had better DCR (P = 0.032) and longer PFS (P = 0.015). Liver metastasis (P = 0.007) and combined systemic therapy (P = 0.036) were significantly associated with DCR, but not associated with PFS or OS. Conclusions: Some patients with advanced gastric cancer can benefit from apatinib. This study help us to identify patients who has potential benefit from apatinib.
Association between central and peripheral circulating tumor cell (CTC) clusters in oral squamous cell carcinoma (OSCC): A prospective, observational study.
Authors: Bagadia Ritvi K , Rao Vishal Uchila Shishir , Balakrishnan Ajay , George Abhijith , Kumar Prashant
Published: May 2020

e15548

Background: Around 90% of cancer-related mortalities are caused by tumor metastasis. CTC clusters, which constitute an intermediate stage of metastasis, have not been studied extensively in head & neck cancers. The mortality rate of oral cancers remains alarmingly high, despite multimodality treatment. The aim of the study is to identify the presence of CTC clusters in patients with Oral Squamous Cell Carcinoma (OSCC) and to correlate their presence with clinical and pathological factors. Methods: Fifty patients diagnosed with histologically proven OSCC, treatment naïve, and underwent surgery at HCG Cancer Centre, Bangalore, were consented and enrolled in the study. An IRB-approved protocol allowed for the collection of 10 ml of blood from central (jugular) and peripheral veins intra-operatively, prior to tumor removal. The culturing of CTC clusters was done using ellipsoidal microwell plates maintained at hypoxic conditions, at the Institute of Bioinformatics, Bangalore. After fourteen days of culturing, the cells were fixed and stained for DAPI, Pan-CK and CD45. The CTC clusters were classified into Loose, Tight and very Tight based on the median gray values obtained from DAPI staining on ImageJ software. Clinical data was collected from patient records and subjected to analysis using Descriptive statistics. Results: From the 50 patients included in the study, 22 (44%) patients exhibited tight clusters in central blood, while only 13 (26%) patients exhibited tight clusters in peripheral blood. A higher clinical stage was observed in a greater percentage of patients with tight clusters in central blood (early: 45.5% versus late: 54.5%), but the same findings could not be inferred with pathological staging (early stage: 59.1% versus late stage: 40.1%). No significant correlation with adverse pathological features was noted. Conclusions: This observational study provides an insight into the varying biological behaviours of similarly grouped cancers, which is based on the standard TNM staging. The study forms the basis for the hypothesis of tight clusters in the central and peripheral circulation, correlating with loco-regional and distant metastasis respectively, thus leading to poorer disease-free and overall survival rates.
Biomarker screening for the early detection of prostate cancer using an exosomal-enrichment RNA liquid biopsy test.
Authors: Roberts Douglas , Mitsock Emily , Ogunlusi Olubode , Yu Seth , Skog Johan
Published: May 2020

e15553

Background: Prostate cancer is one of the most common cancers in men, with approximately 10% of all new cancer cases and ~5% of all cancer deaths in 2019. The standard test for prostate cancer is the Prostate Serum Antigen (PSA) test. The PSA test suffers from low specificity (20-40%) in patients including ‘grey zone’ levels (4-10 ng/mL); moreover, the PSA test fails to identify patients with high-risk cancers. Previously we developed ExoDx Prostate Intelliscore (EPI), a urine-based exosome prostate cancer test optimized to rule out the need for a biopsy (risk stratification for high-grade prostate cancer). This study utilized a next generation exosome-based test that specifically enriches a subtype of prostate cancer exosomes from urine. Early detection of prostate cancer via a non-invasive method is desirable and the identification of patients with high-risk cancer is critical. Here we describe the development of a prostate-specific urinary exosome test for the identification of patients with prostate cancer. Methods: We have developed a prostate-specific enrichment method to isolate exosomes of prostate origin from urine samples. Using an affinity-based method against surface marker proteins found on prostate cells, we were able to selectively enrich for exosomes shed by the prostate gland with demonstrated specificity. Subsequent analysis of exosomal nucleic acids enables a promising panel of gene expression biomarkers capable of distinguishing patients with prostate cancer from healthy individuals. Results: RNA from prostate cancer enriched exosomes was compared to total exosomes from urine. Enrichment of prostate cancer specific exosomes significantly enhanced the RNA signature compared to total urine exosomes. Conclusions: Prostate cancer tests have recently been developed for RNA signatures in urine. Exosomes provide a source of nucleic acids as they are actively shed continuously from living cells as part of their normal life cycle. The urine exosomes can be used for total RNA transcriptome analysis and are therefore a very rich source of biomarkers for prostate cancer that can be tailored to different clinical indications. An affinity-based enrichment for tissue-specific exosomes allow for better resolution of the gene expression profile from the tissue of interest and reduces the RNA targets from non-relevant processes of the bladder and kidneys. The gene signature identified in this ongoing study could potentially provide a non-invasive molecular means for the early diagnosis of prostate cancer.