Mutation Counts Research Articles

Glycosylation modification identifies novel molecular phenotypes and prognostic stratifications of glioma

Glycosylation modification plays a vital role in tumor progression and is highly associated with glioma prognosis. However, the influence of glycosylation modification on the tumor microenvironment (TME) and omic features of glioma remains unclear. Differentially expressed glycosylation-related genes between adjacent and tumor tissues of The Cancer Genome Atlas and Chinese Glioma Genome Atlas datasets were identified. We performed unsupervised clustering to classify patients into different molecular phenotypes, and analyzed their TME heterogeneity, including immunocyte infiltration, immune pathways and tumor purity. Subsequently, we developed a prognostic predicting system named GlycoScore by stepwise least absolute shrinkage and selection operator-Cox regression to evaluate the modification pattern and its association with somatic mutation, clinical significance, immune fractions and drug resistance. Two clustering clusters were identified and presented distinct clinical outcomes and biological functions characterized by hotand cold tumors respectively. Patients with higher GlycoScores exhibited poor prognosis, less mutation counts, and were more sensitive to chemotherapeutics. We also confirmed that the GlycoScore severed as an independent risk factor. Cancer hallmarks such as cell cycle, hippo, and TGFβ were active in the high-GlycoScore group. The combination of tumor mutation burden and the GlycoScore presented an excellent performance in prognostic stratification. Our study suggests that glycosylation is essential for modeling TME of glioma and the GlycoScore is a promising prognostic signature and indicator of immunotherapeutic efficacy.

Abstract 5679: Application of a hypoxia metric to investigate tumor mutational burden and cancer cell immunogenicity in vitro

Abstract Hypoxia afflicts the microenvironment of solid tumors fueling tumor malignancy. Using an 8-gene in vitro hypoxia signature, we have recently shown that pancreatic cancer patients with highly hypoxic tumors experience worse survival and exhibit markers of an immunosuppressed microenvironment. Simultaneously, we obtained evidence of increased mutation count in more hypoxic tumors, which accommodates previous reports associating hypoxia with the regulation of tumor mutational burden (TMB) and genomic instability. These two features could enhance tumor immunogenicity by increasing neoantigen expression, enabling immune recognition and tumor eradication. In this work our aim is to further elucidate the underlying role of hypoxia in generating TMB and genomic instability, and how it contributes to cancer cells immunogenicity. To this end, we applied our recently identified 8-gene hypoxia signature to score the hypoxic response of different cancer cells, considering their median expression levels of the signature genes upon exposure to hypoxia (1% oxygen). The hypoxia score was then applied as a metric to distinguish between the highest and lowest scoring cells, enabling a novel approach for evaluating the intrinsic impact of hypoxia on immunogenicity. After twenty passages under hypoxic conditions (1% oxygen), scored cancer cells were subjected to whole exome sequencing (WES) using the Ion Chef System. Following data processing using appropriate bioinformatic tools, variants were called in the hypoxia-treated cells using their normoxic (21% oxygen) counterparts as controls. We first demonstrated that hypoxia induced an increase in TMB in all cell lines. More interestingly, higher scoring cells were found to display, on average, higher TMB than lower scoring cells, suggesting that cells that have a lower hypoxia score tend to be less mutable. Copy number alterations, mutational signature and neoantigen load predictions generated from the WES data will be discussed. The microarray analysis of transcriptomic data from these hypoxia-treated cells, as well as the relationship between the high TMB, high neoantigen load and increased activation of immune response pathways will also be illustrated. Citation Format: Raefa Abou Khouzam, Nagwa Ahmed Zeinelabdin, Mohak Sharda, Husam Nawafleh, Ayda shah Mahmood, Munazza Samar Khan, Goutham Hassan Venkatesh, Salem Chouaib, Shyama Prasad Rao. Application of a hypoxia metric to investigate tumor mutational burden and cancer cell immunogenicity in vitro [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5679.

Abstract 5739: Genetic characteristics of the early stage Chinese NSCLC patients

Abstract Background: Lung Cancer, predominantly non-small-cell lung cancer (NSCLC), is the leading cause of cancer-related death in China. Although precision therapies for patients with advanced or metastatic NSCLC have developed over the past two decades, the 5-year survival rate remains low at 0%-26%. Thus, recurrence monitoring and treatment at an early stage are of great significance for improving survival benefits. Here, we explored the genetic characteristics of early stage NSCLC patients to provide additional values to predict recurrence risk and treatment strategies. Methods: Tumor or plasma samples were collected from 146 Chinese NSCLC patients in stage I-III (127 LUAD, 5 LUSC, 2 LUASC, 12 unknown pathology). 733 cancer-related genes were sequenced on NGS platform for variants analysis. Tumor mutation burden (TMB) was defined by all somatic mutation counts in coding region per megabase. Immunohistochemistry (IHC) was performed using antibodies (clone 22C3 or SP263) to analyze PD-L1 expression. PD-L1 positivity was defined as TPS ≥1%. Online databases, such as cBioPortal, TIMER2.0 and Kaplan-Meier Plotter, were also used for analysis. Statistical analysis was performed using the R Software (v 4.0.5). Results: Of 146 NSCLC patients, commonly mutated genes were detected, including EGFR (68%), TP53 (40%), KMT2D (10%), MSH3 (9%), RBM10 (9%), ZFHX3 (9%). Six pathogenic or likely pathogenic fusion variants harbored in common driver genes like EGFR, ALK and ROS1 were identified for the first time. EGFR was more frequently mutated in female patients (P < 0.01), while CDH10 mutations occurred more often in male patients (P < 0.05). Correlation analysis showed RBM10, ZFHX3 and CTNNB1 mutations were respectively co-occurrent with CARD11, ARHGAP5 and APC, reflecting the distinct profile of our cohort when compared to cBioPortal. Patients with TP53mut, MSH3mut or LRP1Bmut had significantly higher TMB (P < 0.05). Compared to PD-L1- group, TP53, TIMELESS, FBXW7 were remarkably mutated in PD-L1+ group. In LUAD, SPTA1mut and TIMELESSmut were noticeably related to CD8+T and B cell infiltration, while FBXW7mut and ZFHX3mut with more M1 macrophages infiltration. Prognostic analysis showed that LUAD patients harboring MSH3mut (P < 0.001) and SPENmut (P = 0.0005) at stage II exhibited significantly shorter RFS. Moreover, we found that cases carried somatic mutations in TP53 signaling pathway had more lymph node metastasis. Conclusion: These findings expanded our understanding of the early stage Chinese NSCLC, hopefully providing valuable guidance for recurrence risk prediction and treatment strategies. Citation Format: Naiquan Mao, Weishen Yao, Liufu Su, Xiaochun Huang, mengli Huang. Genetic characteristics of the early stage Chinese NSCLC patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5739.

Somatic mutation-associated risk index based on lncRNA expression for predicting prognosis in acute myeloid leukemia

ABSTRACT Objectives: Genomic instability has several implications for acute myeloid leukemia (AML) prognosis. This article aims to construct a somatic mutation-associated risk index (SMRI) of genomic instability for AML to predict prognosis and explore the potential determinants of AML prognosis. Methods: We obtained differentially expressed lncRNAs from genomic instability subtypes and selected six lncRNAs to construct the SMRI through multivariate Cox regression analysis. The median SMRI classified patients into high and low SMRI groups. Kaplan–Meier survival analysis was used to clarify the prognostic differences of SMRI subtypes. Receiver operating characteristic curve analysis was performed to elucidate the value of SMRI as a prognostic indicator. Gene set variation analysis, tumor mutation burden (TMB) analysis, immune infiltration, and immune checkpoint expression analysis were performed to investigate possible causes for the differences in prognosis of SMRI subtypes. Results: The high SMRI group exhibited a poor prognosis, which was characterized by elevated levels of TMB, mutation counts (TP53, NPM1, DNMT3A, and FLT3-TKD), CD8+ T cell infiltration, and immune checkpoint (PD-1, PD-L2, CTLA4, LAG3) expression. The SMRI was still associated with prognosis, even after adjustment for age, sex, cytogenetic risk, DNMT3A status, FLT3 status, and NPM1 status. Gene set variation analysis showed that AML with FLT3-ITD mutation, CEBPA mutation, and LSCs (leukemia stem cells) were enriched in the high SMRI group. Conclusion: Our research suggests that the SMRI derived from genomic instability subtypes is a useful biomarker for predicting prognosis and may be beneficial for improving the clinical outcome of patients with AML.

Correlation of BRAF genomic alterations with higher mutation burden and predictive response to immune checkpoint inhibitors among human malignancies.

e14589 Background: The treatment of BRAF-mutated melanoma leads the evolution of management in BRAF-mutated human malignancies from targeted agents to immune checkpoint inhibition. The correlation between BRAF genomic alterations and mutation burden is not well-studied as well as its prognostic role and predictive role to immune checkpoint inhibitors. Methods: Three Public genomic database (AACR GENIE v.11 [N = 119,058], MSKCC pan-cancer [10,080], MSKCC ICI [1,608]) were retrieved from cBioPortal ( https://www.cbioportal.org ). Raw data of cancer types, BRAF status, mutation counts (MC), exposure to immune checkpoints inhibitors (ICIs), and survival were analyzed. Violin plots were used to compare the distribution of MC among BRAF-altered and -wild tumors, and the Kaplan-Meier method was used to estimate survival rates. Statistical significance was set at a two-sided P < 0.05. Results: BRAF genomic alterations exist in 7,361 (6.2%, AACR GENIE v.11), 560 (5.6%, MSKCC pan-cancer) and 165 patients (10.3%, MSKCC ICI). There are 4,851 (4.1%), 357 (3.6%), and 312 (19.4%) patients with melanoma in these database, respectively. AACR GENIE v.11 includes the other two MSKCC databases, and 10.7% (N = 1,074) patients in MSKCC pan-cancer databases are also cases in MSKCC ICI. The median MCs were 7/Mb and 4/Mb, with interquartile ranges of 1 to 1,467 and 1 to 821 in the BRAF-altered and -wild tumors in AACR GENIE v.11. In this three databases, the distribution of MC was different between the BRAF-altered and -wild tumors (p < 0.001). In MSKCC pan-cancer cohort, the median overall survival in BRAF-altered and -wild tumors are 22.8 vs. 26.3 months (logrank p = 0.005). In MSKCC ICI cohort, the median overall survival of BRAF-altered and -wild tumors are 47.0 vs. 17.0 months (logrank p < 0.001). Conclusions: BRAF genomic alterations are associated with higher mutation counts, represent a poor prognostic factor and positively predictive to immune checkpoint inhibition among human malignancies.

Examination of mutation signature and mutation spectrum as predictors of response to immune checkpoint blockade therapy.

2537 Background: In recent years, there has been promising progress in the use of immune checkpoint blockade (ICB) as a treatment for various cancer types such as lung, kidney, bladder, skin, colon, and breast cancer. In order for this modality to have increased success, more precise selection tools are needed to predict which patients will benefit from treatment. Tumor Mutational burden (TMB) has been implicated as a biomarker for ICB response due to the increased tumor immunogenicity present in high TMB samples. However, we hypothesize that the quality of mutations may also have an impact in determining immune response, rather than just the quantity of mutations alone. Methods: A retrospective analysis of 2041 patients across multiple solid tumor types was conducted using an open-access, open-source cancer genomics database (Gao, Sci Signal, 2013), with the goal of assessing the influence of mutational signature on patient response to ICB (including CTLA-4, PD-1, and PDL-1). Patient demographics, treatment variables/outcomes, and tumor variables, including mutation spectrum, and mutation count, were evaluated. A paired two sample test for means was used to analyze data with p < 0.05 considered statistically significant. Response outcomes were determined per RECIST v 1.1. Results: Our data demonstrate that mutational spectrum profiles show distinct patterns when considering response to ICB, independent of TMB. Patients with breast invasive ductal carcinoma who responded to ICB had less C > A mutations ( p = 0.020) and more C > T mutations ( p = 0.017) compared to non-responders. Lung adenocarcinoma responders had less C > T mutations ( p = 0.003) and more C > A mutations ( p = 0.0003) compared to non-responders. Furthermore, both head and neck squamous cell carcinoma and glioblastoma multiforme responders had more T > A mutations ( p = 0.047 and p = 0.011, respectively) than non-responders. However, in other cancer types studied (urothelial, colon, renal, lung squamous, and melanoma), there were no obvious trends to distinguish responders from non-responders. Preferentially mutated gene targets were also considered for all 10 cancer types (including CARD11, SMARCA4, GLI1, PIK3R1, PTPRT, among others), many of which were also tied to the unique mutational signatures observed and which may be separately identified as novel ICB therapy targets. Conclusions: We found that specific mutation type may impact response to ICB in at least 4 cancer types, including breast, lung adenocarcinoma, head and neck squamous cell carcinoma and glioblastoma multiforme. Of note, this trend was not found in other cancer types which are typically found to have higher TMB. Moving away from solely considering the magnitude of mutations in tumor samples and towards identifying specific mutational signatures may aid in providing necessary specificity for selecting patients for ICB cancer therapy.

A pan-cancer analysis of P21 activated kinase (PAK) family genes as potential biomarkers for immune checkpoint therapy.

2614 Background: PAK (P21 activated kinase) is a family of serine threonine kinases, that consist of 6 members, PAKs 1-6, which are positioned at an intersection of multiple signaling pathways implicated in oncogenesis. Recent studies have found relationship between mutations of some PAK family genes and antitumor immunity. Here, we explored the associations of PAK family gene mutations with ICI response based on multidimensional data from multiple solid tumors. Methods: An immunotherapy cohort (Broad/Dana-Farber, Nat Genet 2018, N = 249) within 7 types of tumors as discovery stage, was used to explore the association of PAK family genes mutations with tumor mutation burden (TMB) and efficacy of immunotherapy. To further validate the predictive value of PAK family genes, we employed another ICI-treated cohort, MSKCC cohort (Samstein, Nat Genet 2019, n = 1661), as a validation cohort. TMB was calculated as the total count of nonsynonymous mutations in coding sequence. Results: In discovery cohort, compared to that in the PAK-Wt group, longer OS was observed in the PAK-Mut group (mOS: 31.3 vs 15.4 months, HR = 0.59, 95% CI: 0.36 to 0.96, P= 0.03). In addition, PAK-Mut group had higher TMB ( P< 0.001). Median TMB in PAK-Mut group and PAK-Wt group is 17.79 (IQR, 7.45-36.55) Mut/Mb and 5.47 (IQR, 2.46-9.09) Mut/Mb, separately. In validation cohort, PAK-Mut patients also achieved significant improved OS compared with PAK-Wt patients (mOS: 44.0 vs 17.0 months, HR = 0.51, 95% CI: 0.37 to 0.69, P< 0.001). Moreover, multivariable analysis of validation cohort demonstrated that PAK-Mut was associated with better OS (HR = 0.71; 95%CI, 0.52-0.98; P = 0.036), after adjusting for Age, Gender, Metastasis, Treatment, TMB, Cancer type. Compared with PAK-Wt group (median [IQR]: 5.90 [2.95-10.04]), PAK-Mut group (median [IQR]: 21.19 [11.81-43.29]) also had higher TMB ( P< 0.001). Conclusions: The results indicated that PAK family genes mutation was associated with a higher TMB both discovery and validation cohort. Analysis of discover and validation immunotherapy cohort data showed PAK family was associated with better OS. These findings indicate that these genes mutation may serve as a potential predictive biomarker for ICI in solid tumor patients.

Clinical Analysis of Gene Mutation and Vascular Events in Patients with BCR/ABL Negative Myeloproliferative Neoplasms

To explore the relationship between clinical features, peripheral blood cell count, coagulation function, gene mutation and hemorrhagic events and thrombotic events in essential thrombocythemia (ET), polycythemia vera (PV), and primary myelofibrosis(PMF) patients. Clinical data of 78 patients with ET, PV, and PMF who were admitted to the Second Affiliated Hospital of Chongqing Medical University between September 2019 and August 2020 were retrospectively analyzed. Information about sex, age, gene mutation, peripheral blood cell count, coagulation function, and hemorrhagic and thrombotic events was included, and the influence of these data on the occurrence of hemorrhagic and thrombotic events was estimated. Among the 78 patients with myeloproliferative neoplasms, there were 47 cases of ET, 15 cases of PV, and 16 cases of PMF.A total of 10 patients (12.82%) experienced hemorrhagic events and 27 (34.62%) experienced thrombotic events. Male,patients aged ≥ 60 years, and patients with a JAK2V617F mutation were more likely to experience thrombotic events (P<0.05). Patients with thrombotic events had higher platelet (PLT) counts and fibrinogen (FIB) levels than patients without hemorrhagic-thrombotic events (P<0.05).White blood cell (WBC) count, red blood cell (RBC) count, hemoglobin (HGB) level, prothrombin time (PT), activated partial thromboplastin time (APTT), thrombin time (TT) and international normalized ratio (INR) showed no statistical difference between patients with thrombotic events and patients without hemorrhagic-thrombotic events (P>0.05). There was also no significant difference in the above-mentioned indexes between patients with hemorrhagic events and patients without hemorrhagic-thrombotic events (P>0.05). Among JAK2V617F positive myeloproliferative neoplasm patients, male patients were more likely to have thrombotic events (P<0.05), and patients with thrombotic events had higher platelet counts than those without hemorrhagic-thrombotic events (P<0.05). There was no significant difference in age, white blood cell count, red blood cell count, hemoglobin level, PT, APTT, FIB, TT or INR between patients with thrombotic events and patients without hemorrhagic-thrombotic events (P>0.05). Sex, age, JAK2V617F mutation and platelet count have a certain value for predicting thrombosis in patients with myeloproliferative neoplasms.

Association of DNA damage repair (DDR) mutations (mts) and clinical outcomes in CALGB 90601 (Alliance).

4521 Background: Platinum-based chemotherapy is the standard 1st-line therapy for metastatic urothelial cancer (mUC). C90601 was a randomized phase III trial testing gemcitabine and cisplatin (GC) with bevacizumab (B) or placebo (P) in patients (pts) with untreated mUC. Median overall survival (OS) for GCB vs GCP was 14.5 months (mo) vs 14.3 mo (p=0.14) and median progression-free survival (PFS) was 8 vs 6.7 mo, respectively. DDR mts have been implicated in response and survival in mUC and were investigated in this negative trial. Methods: C90601 enrolled 506 pts randomized 1:1 to GCB or GCP from 7/15/09-12/2/14, with stratification for prior chemotherapy and visceral metastases. Consenting pts submitted archival FFPE tumor specimens and blood for matched germline (g)DNA. Tumor and gDNA were sequenced by MSK-IMPACT, a 468-gene exon capture assay, to detect mts in select DDR genes. The proportional hazards model was used to correlate mts in the DNA helicase ERCC2 (pre-specified hypothesis) and additional DDR gene panels being explored in prospective trials in muscle-invasive disease with OS and PFS, adjusting for tumor mt burden and stratification factors. Mts were categorized as deleterious (del) or non-del using pre-defined published criteria. Results: 208 pts underwent DNA sequencing. Clinical features and PFS/OS were comparable to the 506-pt cohort. Median sequencing coverage was 497X. Median mutation count was 13.2 and 8.8 for DDR mt and wild-type tumors, respectively. A non-significant improvement in OS and PFS was seen in pts with ERCC2 mts (HR 0.70), but the 5.3% frequency of ERCC2 mts was lower than in historical series. Neither del mts (table) nor any mts in DDR genes were associated with PFS/OS. Conclusions: DDR mts were not associated with improved outcomes in C90601. The reliance on archival specimens, lower-than-expected ERCC2 mt frequency, small sample sizes, and tumor genomic heterogeneity may have influenced the predictive capacity of DDR mts in this cohort. Similar analyses are underway in pts who received neoadjuvant chemotherapy prior to cystectomy from completed prospective trials. Support: U10CA180821, U10CA180882, Genentech.[Table: see text]

Prognostic and predictive impact of circulating tumor DNA in advanced gastric cancer treated with immune checkpoint blockade.

e16019 Background: The utility of circulating tumor DNA (ctDNA) as a biomarker in patients with advanced gastric cancer (GC) receiving immune checkpoint blockade (ICB) is uncertain. Methods: Blood samples from advanced GC patients who received ICB were collected. The next-generation sequencing(NGS)–based targeted 733-gene and 61-gene panels (3D Medicines Inc.) were used to characterize somatic genomic alterations for baseline and on-treatment blood samples (4-12 weeks) respectively. For each sample, somatic single-nucleotide variants (SNV) and insertion/deletion mutations (indels) were used to calculate the mean variant allele frequency (VAF) and total mutation count. A VAF of 0.1% was the limit of detection, with the exception of variants detected in both pretreatment and on-treatment samples such that variants with VAF &lt; 0.1% in one sample were retained if the exact variant was also detected in the paired sample with VAF ≥0.1%. The variant genes at baseline were filtered against 61-genes panel list for calculating paired-baseline-VAF. The delta-VAF was calculated per patient as the sum of the on-treatment VAF minus the paired-baseline-VAF for each detected SNV or indels divided by the number of detected SNVs or indels. The percentage of molecular response was calculated per patient as delta-VAF divided by paired-baseline-VAF. Results: A total of 16 patients were was enrolled. The detection rate was 93.8% (15/16). The median total mutation count and baseline-VAF were 8 (range 0-44) and 3.13% (range 0-40.4%) respectively. The patients with top 50% baseline-VAF had shorter median PFS, compared to that with bottom 50% baseline-VAF (2.55 vs 4.85 months, p = 0.022, HR 2.98, 95%CI: 0.96-9.26). For baseline blood samples (N = 16), the median total mutation count and paired-baseline-VAF was 4 and 3.89% (range 0-40.4%) respectively. For on-treatment blood samples (N = 10), the median total mutation count and VAF were 3 (range 1-12) and 5.94% (0.30-37.12%) respectively. All the patients with progressive disease (PD, N = 6) have increased delta-VAF and all patients with stable disease (SD, N = 3) or partial response (PR, N = 1) have decreased delta-VAF. The median delta-VAF for PR, SD and PD were was -6.73%, -2.13% and 3.95% respectively. Compared with paired-baseline and on-treatment-VAF, the median percentage of molecular response for PR, SD and PD were -95.53%, -47.66% (range 7.08%-59.07%) and 138.73% (range 66.40%-347.18%). Compared with patients with increased VAF during treatment, the patients with decreased VAF have longer PFS (median 3.9 vs 1.75 months, p 0.004, HR, 0.24, 95%CI 0.06 to 0.97). Conclusions: This preliminary analysis contributes to understanding the role of ctDNA as a prognostic and predictive biomarker and its potential to complement radiologic endpoints. Further studies are warranted in larger cohort to confirm such observations for better management of GC.

Stepwise evolutionary genomics of early-stage lung adenocarcinoma manifesting as pure, heterogeneous and part-solid ground-glass nodules.

This study was designed to unravel the genomic landscape and evolution of early-stage subsolid lung adenocarcinomas (SSN-LUADs) manifesting as pure ground-glass nodules (pGGNs), heterogeneous ground-glass nodules (HGGNs) and part-solid nodules (PSNs). Samples subjected to either broad-panel next-generation sequencing (NGS) or whole-exome sequencing (WES) were included. Clinicopathologic and genomic features were compared among pGGN, HGGN and PSN, while tumour evolutionary trajectories and mutational signatures were evaluated in the entire cohort. In total, 247 SSN-LUAD samples subjected to broad-panel NGS and 125 to WES were identified. Compared with PSNs, HGGNs had significantly lower tumour mutation count (P < 0.001), genomic alteration count (P < 0.001), and intra-tumour heterogeneity (P = 0.005). Statistically significant upward trends were observed in alterations involving driver mutations and oncogenic pathways from pGGNs to HGGNs to PSNs. EGFR mutation was proved to be a key early event in the progression of SSN-LUADs, with subsequently two evolutionary trajectories involving either RBM10 or TP53 mutation in the cancer-evolution models. This study provided evidence for unravelling the previously unknown genomic underpinnings associated with SSN-LUAD evolution from pGGN to HGGN to PSN, proving that HGGN was an intermediate SSN form between pGGN and PSN genetically.

Tumor mutational burden presents limiting effects on predicting the efficacy of immune checkpoint inhibitors and prognostic assessment in adrenocortical carcinoma

BackgroundAdrenocortical carcinoma (ACC) is a highly malignant urologic cancer and tends to metastasize. Although immune checkpoint inhibitors (ICIs) bring a glimmer of light to conquer ACC, only a fraction of patients have benefit from ICIs treatment. It is well known that tumor mutational burden (TMB) is closely associated with the efficacy and response rate of immunotherapy. However, its roles in ACC were not investigated.MethodsUsing somatic mutations data of 92 ACC samples in TCGA database, we calculated their TMB values by the ‘maftools’ package in R software (Ver 3.6.3). To explore the roles of TMB in ICIs therapy, we have addressed this issue from three perspectives. First, the effects of TMB levels on tumor immune microenvironment (TIM) were analyzed through CIBERSORT algorithm, ssGSEA method and TIMER web server. Second, we investigated the expressive correlations between TMB level and five pivotal immune checkpoints based on Pearson coefficient. Third, the difference in TIDE score between high- and low-TMB groups was compared. The prognostic value of TMB was also evaluated. Besides, GSEA was performed to determine the changes in the activities of signaling pathways caused by TMB.ResultsTMB values in ACC samples were not high. The average of total mutation counts in each sample was only 21.5. High TMB could lead metabolic reprogramming and poor survival outcomes. However, it was unable to affect the infiltration levels of lymphocytes, and failed to facilitate the activities of immune-related pathways. Regarding immune checkpoints (ICs), only PD-L1 upregulation could result in a good prognosis, and TMB level did not correlate with the expressions of other ICs except for LAG3. There was no significant difference in TIDE score between high- and low-TMB groups. Combining the present results and previous study, we speculated that inadequate stimulation for neoantigens formation, intrinsic immune-resistance and special genomic alterations were three possible reasons for TMB limiting functions in TIM and ICIs. Besides, TMB was toughly applied in clinical practice due to its high cost of determination and non-universal definition of high TMB.ConclusionsTMB presents limiting effects on prediction for ICIs efficacy and prognostic assessment for ACC patients.

Clinical Characteristics in the Prediction of Posttreatment Survival of Patients with Ovarian Cancer.

Objective To determine the efficacy of clinical characteristics in the prediction of prognosis in patients with ovarian cancer. Methods Clinical data were collected from 3 datasets from TCGA database, including 1680 cases of ovarian serous cystadenocarcinoma, and were analyzed. Patients with ovarian cancer admitted to our hospital in 2016 were retrieved and followed up for prognosis analysis. Results From the datasets, for patients > 75 years old at the time of diagnosis, histologic grade and mutation count were good predictors for disease-free survival, while for patients > 50 years old at the time of diagnosis, histologic grade, race, fraction genome altered, and mutation count were good predictors for overall survival. In the patients (n = 38) retrieved from our hospital, the longest dimension of lesion (cm) and body weight at admission were good predictors for overall survival. Conclusions Those clinical factors, together with the two predictive equations, could be used to comprehensively predict the long-term prognosis of patients with ovarian cancer.

PD14-09 ANALYZING DIFFERENCES IN SOMATIC MUTATIONS IN LOCALIZED BLADDER CANCER PATIENTS BY RACE AND SEX: AN ANALYSIS OF DATA FROM THE CBIOPORTAL FOR CANCER GENOMICS

You have accessJournal of UrologyCME1 May 2022PD14-09 ANALYZING DIFFERENCES IN SOMATIC MUTATIONS IN LOCALIZED BLADDER CANCER PATIENTS BY RACE AND SEX: AN ANALYSIS OF DATA FROM THE CBIOPORTAL FOR CANCER GENOMICS Kelsey Baker, Yaw Nyame, Mary Redman, Robert Montgomery, Petros Grivas, and Jonathan Wright Kelsey BakerKelsey Baker More articles by this author , Yaw NyameYaw Nyame More articles by this author , Mary RedmanMary Redman More articles by this author , Robert MontgomeryRobert Montgomery More articles by this author , Petros GrivasPetros Grivas More articles by this author , and Jonathan WrightJonathan Wright More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000002546.09AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Disparities in bladder cancer outcomes exist by race/ethnicity and sex. However, limited data exists on differences in tumor biology by these factors. We hypothesize that social, environmental, and biologic differences that exist by race and sex are likely to drive different biologic phenotypes in bladder cancer. This study aimed to evaluate if these differences existed in a large, multi-institutional cohort. METHODS: Nine urothelial cancer studies and 122 genes (113 actionable, 31 DNA damage repair (DDR)) that are commonly associated with bladder cancer were identified in the cBioPortal platform. Patients were excluded if they had low-grade disease or had no demographic data. Multivariable logistic regression, adjusted for age and disease stage, was performed to evaluate the association between mutation status and race/sex. P < 0.05 was used for statistical significance. No adjustments were made for multiple comparisons. RESULTS: Mutation and demographic data for 880 patients was included for analysis. The median age in this cohort was 67 years (range: 25-98), with 216 (25%) female, 423 (48%) White, 74 (8%) non-White, and 383 (44%) unknown race. Muscle-invasive disease was present in 78% of cases. The median total mutation count was 74 (IQR: 11-186). At least one actionable gene mutation and at least one DDR gene mutation was found in 832 (95%) and 392 (45%) patients, respectively. Somatic mutated gene prevalence differed by sex and race (Table 1). Males in this cohort were more likely to have ARID1A, ATM, CHD6, ERBB2, NCOR1, and FLT4 mutations. White race was associated with higher likelihood of having ARID1A, EP300, and TP53 mutations and lower likelihood of having HRAS mutations. CONCLUSIONS: Differences in somatic mutations existed by both race and sex in a large cohort of patients with bladder cancer. These findings are limited by poor representation of females, non-White patients, limited demographic data collection, and retrospective design. Intentional efforts are needed to build representative patient cohorts and more complete demographic data collection to assess the role of social, environmental, and sex-specific biologic drivers of bladder cancer. Source of Funding: This study was supported by institutional funding through the MXD Championships (JLW) © 2022 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 207Issue Supplement 5May 2022Page: e259 Advertisement Copyright & Permissions© 2022 by American Urological Association Education and Research, Inc.MetricsAuthor Information Kelsey Baker More articles by this author Yaw Nyame More articles by this author Mary Redman More articles by this author Robert Montgomery More articles by this author Petros Grivas More articles by this author Jonathan Wright More articles by this author Expand All Advertisement PDF DownloadLoading ...

Comparison of SARS-CoV-2 sequencing using the ONT GridION and the Illumina MiSeq

BackgroundOver 4 million SARS-CoV-2 genomes have been sequenced globally in the past 2 years. This has been crucial in elucidating transmission chains within communities, the development of new diagnostic methods, vaccines, and antivirals. Although several sequencing technologies have been employed, Illumina and Oxford Nanopore remain the two most commonly used platforms. The sequence quality between these two platforms warrants a comparison of the genomes produced by the two technologies. Here, we compared the SARS-CoV-2 consensus genomes obtained from the Oxford Nanopore Technology GridION and the Illumina MiSeq for 28 sequencing runs.ResultsOur results show that the MiSeq had a significantly higher number of consensus genomes classified by Nextclade as good and mediocre compared to the GridION. The MiSeq also had a significantly higher genome coverage and mutation counts than the GridION.ConclusionDue to the low genome coverage, high number of indels, and sensitivity to SARS-CoV-2 viral load noted with the GridION when compared to MiSeq, we can conclude that the MiSeq is more favourable for SARS-CoV-2 genomic surveillance, as successful genomic surveillance is dependent on high quality, near-whole consensus genomes.

High Mutation Burden in ER-Positive/HER2-Negative/Luminal Breast Cancers.

Background: Tumor mutation burden (TMB) is arising as a useful marker of checkpoint inhibitors’ effectiveness in cancer patients in general and has been proposed as predictive in breast cancers. Despite the initial success of checkpoint inhibitors in triple-negative breast cancer, ER-positive breast cancers are less amenable to immunotherapy treatments due to the lower immunogenicity of this subset, associated with lower TMB and less pronounced inflammatory cell infiltration. However, a minority of ER-positive breast cancers do have a higher TMB and could be targets of immune checkpoint inhibitors. Methods: This investigation uses publicly available genomic data to examine ER-positive/HER2-negative or luminal breast cancers with high mutation numbers and compare them with cancers of the same subtype and low mutation numbers. Clinical characteristics and molecular correlates according to mutation numbers are described. Results: ER-positive/HER2-negative and luminal breast cancers with high mutation numbers have a higher prevalence of PIK3CA mutations and in some of the series examined mutations in TP53 and CDH1. A significant proportion of cancers with high mutation numbers carry mutations in microsatellite instability genes and genes involved in DNA damage response. Despite these differences, the prognosis of ER-positive/HER2-negative and luminal breast cancers with high mutation numbers is not significantly different compared to counterparts with lower mutation counts. Conclusions: These data may inform the potential suitability of these cancers for immunotherapy and could guide the development of rational combination therapies based on immune checkpoint inhibitors with other targeted drugs.

Comprehensive characterization of m6A methylation and its impact on prognosis, genome instability, and tumor microenvironment in hepatocellular carcinoma

BackgroundN6-methyladenosine (m6A) RNA regulation was recently reported to be important in carcinogenesis and cancer development. However, the characteristics of m6A modification and its correlations with clinical features, genome instability, tumor microenvironments (TMEs), and immunotherapy responses in hepatocellular carcinoma (HCC) have not been fully explored.MethodsWe systematically analyzed the m6A regulator-based expression patterns of 486 patients with HCC from The Cancer Genome Atlas and Gene Expression Omnibus databases, and correlated these patterns with clinical outcomes, somatic mutations, TME cell infiltration, and immunotherapy responses. The m6A score was developed by principal component analysis to evaluate m6A modifications in individual patients.ResultsM6A regulators were dysregulated in HCC samples, among which 18 m6A regulators were identified as risk factors for prognosis. Three m6A regulator-based expression patterns, namely m6A clusters, were determined among HCC patients by m6A regulators with different m6A scores, somatic mutation counts, and specific TME features. Additionally, three distinct m6A regulator-associated gene-based expression patterns were also identified based on prognosis-associated genes that were differentially expressed among the three m6A clusters, showing similar properties as the m6A regulator-based expression patterns. Higher m6A scores were correlated with older age, advanced stages, lower overall survival, higher somatic mutation counts, elevated PD-L1 expression levels, and poorer responses to immune checkpoint inhibitors. The m6A score was validated as an independent and valuable prognostic factor for HCC.ConclusionM6A modification is correlated with genome instability and TME in HCC. Evaluating m6A regulator-based expression patterns and the m6A score of individual tumors may help identify candidate biomarkers for prognosis prediction and immunotherapeutic strategy selection.

Glycosylation-Related Genes Predict the Prognosis and Immune Fraction of Ovarian Cancer Patients Based on Weighted Gene Coexpression Network Analysis (WGCNA) and Machine Learning.

Background Ovarian cancer (OC) is a malignancy exhibiting high mortality in female tumors. Glycosylation is a posttranslational modification of proteins but research has failed to demonstrate a systematic link between glycosylation-related signatures and tumor environment of OC. Purpose This study is aimed at developing a novel model with glycosylation-related messenger RNAs (GRmRNAs) to predict the prognosis and immune function in OC patients. Methods The transcriptional profiles and clinical phenotypes of OC patients were collected from the Gene Expression Omnibus and The Cancer Genome Atlas databases. A weighted gene coexpression network analysis and machine learning were performed to find the optimal survival-related GRmRNAs. Least absolute shrinkage and selection operator regression (LASSO) and Cox regression were carried out to calculate the coefficients of each GRmRNA and compute the risk score of each patient as well as develop a prognostic model. A nomogram model was constructed, and several algorithms were used to investigate the relationship between risk subtypes and immune-infiltrating levels. Results A total of four signatures (ALG8, DCTN4, DCTN6, and UBB) were determined to calculate the risk scores, classifying patients into the high-and low-risk groups. High-risk patients exhibited significantly poorer survival outcomes, and the established nomogram model had a promising prediction for OC patients' prognosis. Tumor purity and tumor mutation burden were negatively correlated with risk scores. In addition, risk scores held statistical associations with pathway signatures such as Wnt, Hippo, and reactive oxygen species, and nonsynonymous mutation counts. Conclusion The currently established risk scores based on GRmRNAs can accurately predict the prognosis, the immune microenvironment, and the immunotherapeutic efficacy of OC patients.

Disparate genomic characteristics of patients with early-stage lung adenocarcinoma manifesting as radiological subsolid or solid lesions

IntroductionEarly-stage lung adenocarcinoma (LUAD) manifesting as subsolid nodules (SSNs) exhibit more favorable prognosis than solid nodules (SNs). However, the genomic underpinnings behind their indolent tumor behavior remain largely unexplained. MethodsWe identified patients with stage I invasive LUAD who underwent complete surgical resection and broad-panel next-generation sequencing (NGS). Comparative genomic profiling was then performed by radiological subtype (SSNs vs. SNs) regarding the general genomic features, driver genes, oncogenic pathways, therapeutic actionability, and evolutionary trajectory. ResultsIn total, 177 SSN-LUADs and 133 SN-LUADs were included. Compared with SNs, SSN-LUADs possessed lower somatic mutation count (P < 0.001), genomic alteration count (P = 0.002), and intra-tumor heterogeneity (P = 0.006). In terms of driver genes, SSNs harbored more EGFR mutation (77% vs. 62%), but had lower frequencies of genes such as TP53, ARID1A, PIK3CA, CDKN2A, and BRAF (FDR q < 0.1). Besides, RBM10 mutation was independently associated with SSN-LUADs in multivariate analysis (P = 0.033). Three oncogenic pathways (p53, cell cycle, PI3K) were altered with statistical significance in SNs, while only RNA splicing/processing pathway was significantly altered in SSNs (FDR q < 0.1). Also, SSNs had significantly lower number of pathway alterations (P < 0.001). Finally, SSNs and SNs showed distinct evolutionary trajectories regarding somatic mutations during early-stage LUAD progression. ConclusionsThis study performed the first direct comparative genomic profiling in pathologic stage I invasive LUAD by radiological subtype, highlighting a less complex genomic architecture of SSNs, which might be the molecular interpretation of their indolent tumor behavior.

Somatic mutational profiles and germline polygenic risk scores in human cancer

BackgroundThe mutational profile of cancer reflects the activity of the mutagenic processes which have been operative throughout the lineage of the cancer cell. These processes leave characteristic profiles of somatic mutations called mutational signatures. Mutational signatures, including single-base substitution (SBS) signatures, may reflect the effects of exogenous or endogenous exposures.MethodsWe used polygenic risk scores (PRS) to summarize common germline variation associated with cancer risk and other cancer-related traits and examined the association between somatic mutational profiles and germline PRS in 12 cancer types from The Cancer Genome Atlas. Somatic mutational profiles were constructed from whole-exome sequencing data of primary tumors. PRS were calculated for the 12 selected cancer types and 9 non-cancer traits, including cancer risk determinants, hormonal factors, and immune-mediated inflammatory diseases, using germline genetic data and published summary statistics from genome-wide association studies.ResultsWe found 17 statistically significant associations between somatic mutational profiles and germline PRS after Bonferroni correction (p < 3.15 × 10−5), including positive associations between germline inflammatory bowel disease PRS and number of somatic mutations attributed to signature SBS1 in prostate cancer and APOBEC-related signatures in breast cancer. Positive associations were also found between age at menarche PRS and mutation counts of SBS1 in overall and estrogen receptor-positive breast cancer. Consistent with prior studies that found an inverse association between the pubertal development PRS and risk of prostate cancer, likely reflecting hormone-related mechanisms, we found an inverse association between age at menarche PRS and mutation counts of SBS1 in prostate cancer. Inverse associations were also found between several cancer PRS and tumor mutation counts.ConclusionsOur analysis suggests that there are robust associations between tumor somatic mutational profiles and germline PRS. These may reflect the mechanisms through hormone regulation and immune responses that contribute to cancer etiology and drive cancer progression.