Somatic Gene Research Articles

Different Mutational Landscapes in Human Papillomavirus–Induced and Human Papillomavirus–Independent Invasive Penile Squamous Cell Cancers

Penile squamous cell carcinomas (SCC) are rare cancers that arise after transforming human papillomavirus (HPV) infections or independent of HPV in the background of chronic dermatoses. Limited knowledge about genetic alterations driving penile carcinogenesis comes from studies of mainly small cohorts of typically mixed etiology. In this comparative genetic study of HPV-induced and HPV-independent invasive penile SCC of 156 patients from a single institution in a low-incidence country, hotspots of 50 cancer-relevant genes were analyzed with targeted next-generation sequencing. Seventy-nine of 156 SCC were classified as HPV induced, and 77 of 156 SCC arose independent of HPV. Only 28 (35%) of 79 HPV-induced penile SCC, but 69 (90%) of 77 HPV-independent SCC carried somatic gene mutations. PIK3CA, FGFR3, and FBXW7 mutations occurred in both groups in similar numbers as seen in other human cancers. In contrast, mutations in TP53 (44/77; 57%), CDKN2A (35/77; 45%), and HRAS (13/77; 17%) genes occurred with one exception of a HIV positive patient exclusively in HPV-independent SCC with a frequent co-occurrence of TP53 and CDKN2A mutations (28/77; 42%). Mutations in multiple genes occurred in 9 (11%) of 79 HPV-induced SCC versus 47 (62%) of 77 HPV-independent SCC (χ2; P < .001). More than one mutation per gene (multi hits) was characteristic for HPV-independent SCC in 14 (18%) of 77 compared with only 3 (4%) of 79 HPV-induced SCC (χ2; P < .001). The total number of mutations in HPV-induced penile SCC (47 mutations) was significantly lower than that in HPV-independent SCC (143 mutations; Welsh test; P < .001). The presence of somatic driver gene mutations did not correlate with the age of patients, histology, or tumor stage of the primary SCC in either etiologic group, suggesting that acquisition of driver gene mutations is an early event after invasion. This large cohort analysis identified characteristic differences in mutational landscapes for the 2 etiologies. While genetic mutations in tumor suppressor genes drive HPV-independent penile carcinogenesis, oncogenic action of E6 and E7 substitute for mutations in HPV-induced SCC. A subgroup of patients with advanced SCC may be candidates for targeted therapy and clinical trials, although the majority of advanced penile SCC remain a therapeutic challenge.

Molecular findings in myeloid neoplasms

AbstractThe proliferation of new molecular technologies in recent years has greatly advanced our knowledge of the genetics that underlie hematologic cancers. Particularly, with the advent and wide‐implementation of next‐generation sequencing (NGS), a host of somatic (and some germline) gene mutations have been identified as significant in the classification, prognostication, and treatment of the spectrum of myeloid neoplasms. These driver and disease modifier mutations now play a prominent role in the updated international diagnostic guidelines of acute myeloid leukemia (AML), myelodysplastic syndromes/neoplasms (MDS), and myeloproliferative neoplasms (MPN). As high‐throughput technologies such as NGS increasingly become standard in the genetic evaluation of myeloid disorders, it is critical that clinicians understand the clinical relevance of these mutations in order to further personalize patient care. In this review we discuss some of the most essential somatic and cytogenetic findings.

Barley stripe mosaic virus-mediated somatic and heritable gene editing in barley (Hordeum vulgare L.).

Genome editing strategies in barley (Hordeum vulgare L.) typically rely on Agrobacterium-mediated genetic transformation for the delivery of required genetic reagents involving tissue culture techniques. These approaches are genotype-dependent, time-consuming, and labor-intensive, which hampers rapid genome editing in barley. More recently, plant RNA viruses have been engineered to transiently express short guide RNAs facilitating CRISPR/Cas9-based targeted genome editing in plants that constitutively express Cas9. Here, we explored virus-induced genome editing (VIGE) based on barley stripe mosaic virus (BSMV) in Cas9-transgenic barley. Somatic and heritable editing in the ALBOSTRIANS gene (CMF7) resulting in albino/variegated chloroplast-defective barley mutants is shown. In addition, somatic editing in meiosis-related candidate genes in barley encoding ASY1 (an axis-localized HORMA domain protein), MUS81 (a DNA structure-selective endonuclease), and ZYP1 (a transverse filament protein of the synaptonemal complex) was achieved. Hence, the presented VIGE approach using BSMV enables rapid somatic and also heritable targeted gene editing in barley.

Genomic landscape and expression profile of consensus molecular subtype four of colorectal cancer.

Compared to other subtypes, the CMS4 subtype is associated with lacking of effective treatments and poorer survival rates. A total of 24 patients with CRC were included in this study. DNA and RNA sequencing were performed to acquire somatic mutations and gene expression, respectively. MATH was used to quantify intratumoral heterogeneity. PPI and survival analyses were performed to identify hub DEGs. Reactome and KEGG analyses were performed to analyze the pathways of mutated or DEGs. Single-sample gene set enrichment analysis and Xcell were used to categorize the infiltration of immune cells. The CMS4 patients had a poorer PFS than CMS2/3. CTNNB1 and CCNE1 were common mutated genes in the CMS4 subtype, which were enriched in Wnt and cell cycle signaling pathways, respectively. The MATH score of CMS4 subtype was lower. SLC17A6 was a hub DEG. M2 macrophages were more infiltrated in the tumor microenvironment of CMS4 subtype. The CMS4 subtype tended to have an immunosuppressive microenvironment. This study suggested new perspectives for exploring therapeutic strategies for the CMS4 subtype CRC.

Single-cell multi-gene identification of somatic mutations and gene rearrangements in cancer.

In this proof-of-concept study, we developed a single-cell method that provides genotypes of somatic alterations found in coding regions of messenger RNAs and integrates these transcript-based variants with their matching cell transcriptomes. We used nanopore adaptive sampling on single-cell complementary DNAlibraries to validate coding variants in target gene transcripts, and short-read sequencing to characterize cell types harboring the mutations. CRISPR edits for 16 targets were identified using a cancer cell line,and known variants in the cell line were validated using a 352-gene panel. Variants in primary cancer samples were validated using target gene panels ranging from 161 to 529 genes. A gene rearrangement was also identified in one patient, with the rearrangement occurring in two distinct tumor sites.

Necrotic related-lncRNAs: Prediction of prognosis and differentiation between cold and hot tumors in head and neck squamous cell carcinoma.

Treatment of head and neck squamous cell carcinoma (HNSCC) is a substantial clinical challenge due to the high local recurrence rate and chemotherapeutic resistance. This project seeks to identify new potential biomarkers of prognosis prediction and precision medicine to improve this condition. A synthetic data matrix for RNA transcriptome datasets and relevant clinical information on HNSCC and normal tissues was downloaded from the Genotypic Tissue Expression Project and The Cancer Genome Atlas (TCGA). The necrosis-associated long-chain noncoding RNAs (lncRNAs) were identified by Pearson correlation analysis. Then 8-necrotic-lncRNA models in the training, testing and entire sets were established through univariate Cox (uni-Cox) regression and Lasso-Cox regression. Finally, the prognostic ability of the 8-necrotic-lncRNA model was evaluated via survival analysis, nomogram, Cox regression, clinicopathological correlation analysis, and receiver operating characteristic (ROC) curve. Gene enrichment analysis, principal component analysis, immune analysis and prediction of risk group semi-maximum inhibitory concentration (IC50) were also conducted. Correlations between characteristic risk score and immune cell infiltration, immune checkpoint molecules, somatic gene mutations, and anti-cancer drug sensitivity were analyzed. Eight necrosis-associated lncRNAs (AC099850.3, AC243829.2, AL139095.4, SAP30L-AS1, C5orf66-AS1, LIN02084, LIN00996, MIR4435-2HG) were developed to improve the prognosis prediction of HNSCC patients. The risk score distribution, survival status, survival time, and relevant expression standards of these lncRNAs were compared between low- and high-risk groups in the training, testing and entire sets. Kaplan-Meier analysis showed the low-risk patients had significantly better prognosis. The ROC curves revealed the model had an acceptable predictive value in the TCGA training and testing sets. Cox regression and stratified survival analysis indicated that the 8 necrosis-associated lncRNAs were risk factors independent of various clinical parameters. We recombined the patients into 2 clusters through Consensus ClusterPlus R package according to the expressions of necrotic lncRNAs. Significant differences were found in immune cell infiltration, immune checkpoint molecules, and IC50 between clusters, suggesting these characteristics can be used to evaluate the clinical efficacy of chemotherapy and immunotherapy. This risk model may serve as a prognostic signature and provide clues for individualized immunotherapy for HNSCC patients.

High-LET-Radiation-Induced Persistent DNA Damage Response Signaling and Gastrointestinal Cancer Development.

Ionizing radiation (IR) dose, dose rate, and linear energy transfer (LET) determine cellular DNA damage quality and quantity. High-LET heavy ions are prevalent in the deep space environment and can deposit a much greater fraction of total energy in a shorter distance within a cell, causing extensive DNA damage relative to the same dose of low-LET photon radiation. Based on the DNA damage tolerance of a cell, cellular responses are initiated for recovery, cell death, senescence, or proliferation, which are determined through a concerted action of signaling networks classified as DNA damage response (DDR) signaling. The IR-induced DDR initiates cell cycle arrest to repair damaged DNA. When DNA damage is beyond the cellular repair capacity, the DDR for cell death is initiated. An alternative DDR-associated anti-proliferative pathway is the onset of cellular senescence with persistent cell cycle arrest, which is primarily a defense mechanism against oncogenesis. Ongoing DNA damage accumulation below the cell death threshold but above the senescence threshold, along with persistent SASP signaling after chronic exposure to space radiation, pose an increased risk of tumorigenesis in the proliferative gastrointestinal (GI) epithelium, where a subset of IR-induced senescent cells can acquire a senescence-associated secretory phenotype (SASP) and potentially drive oncogenic signaling in nearby bystander cells. Moreover, DDR alterations could result in both somatic gene mutations as well as activation of the pro-inflammatory, pro-oncogenic SASP signaling known to accelerate adenoma-to-carcinoma progression during radiation-induced GI cancer development. In this review, we describe the complex interplay between persistent DNA damage, DDR, cellular senescence, and SASP-associated pro-inflammatory oncogenic signaling in the context of GI carcinogenesis.

Homologous recombination-related (HRR) gene mutation profiling and the association with TMB in Chinese patients with esophageal and gastric cancer.

e16000 Background: Homologous recombination deficiency (HRD) is associated with tumorigenesis and could predict the response of PARP inhibitor (PARPi) therapy and immunotherapy. Loss of function or deleterious mutations in homologous recombination-related (HRR) genes contribute to HRD phenotype. Methods: Tumor tissue samples from 1237 Chinese patients with esophageal and gastric cancer were sequenced with 551 cancer-related genes. Based on the clinical evidence, 15 HRR genes ( ATM, BRCA1, BRCA2, BARD1, BRIP1, CDK12, CHEK1, CHEK2, FANCL, PALB2, RAD51B, RAD51C, RAD51D, RAD54L, PPP2R2A) were selected. 623 patients with WES data from the TCGA datasets were included for analysis. Results: In the Chinese cohort, 25.1% (62/247) esophageal cancer (EC) patients and 20.71% (205/990) gastric cancer (GC) patients exhibited somatic HRR (sHRR) gene mutations. The most frequently mutated genes were CHEK2 (7%) in EC and ATM (8%) in GC. The germline HRR (gHRR) gene mutations were identified in 48.16% (118/245) EC patients and 50.66% (498/983) GC patients. BRCA2 was the most common mutant gene (22% in EC and 9% in GC). 4.84% EC patients and 4.14% GC patients carried both germline and somatic HRR gene mutations. The sHRR-Mut patients had a higher median TMB compared to the sHRR-WT patients (EC, 8.09 vs 4.41, p &lt; 0.001; GC, 8.09 vs 4.26, p &lt; 0.001). However, the median TMB was similar between the gHRR-Mut and the gHRR-WT patients (EC, 5.67 vs 5.67, p = 0.903; GC, 4.96 vs 4.41, p = 0.356). In the TCGA cohort, 15.68% (29/185) EC patients and 23.97% (105/438) GC patients exhibited sHRR gene mutations. The most common mutant gene was ATM (5% in EC and 10% in GC). The sHRR-Mut patients had a higher median TMB compared to the sHRR-WT patients (EC, 4.78 vs 2.75, p &lt; 0.001; GC, 21.23 vs 2.65, p &lt; 0.001). For the TCGA gastric cancer cohort, the patients with sHRR-Mut had a significantly better median overall survival compared to the ones with sHRR-WT (46.9 months vs 26.5 months, p = 0.043). Conclusions: Our data reported the genetic landscape of HRR genes in Chinese esophageal and gastric cancer patients. Patients with somatic HRR gene mutations had a significantly elevated TMB level. In gastric cancer, patients carried somatic HRR gene mutations had a significantly prolonged OS.

Homologous recombination-related (HRR) gene mutation profiling and the association with TMB in Chinese patients with pancreatic cancer.

e16252 Background: Homologous recombination deficiency (HRD) is associated with tumorigenesis and could predict the response of PARP inhibitor (PARPi) therapy and immunotherapy. Loss of function or deleterious mutations in homologous recombination-related (HRR) genes contribute to HRD phenotype. Methods: Tumor tissue samples from 669 Chinese patients with pancreatic cancer were sequenced with 551 cancer-related genes. Based on the clinical evidence, 15 HRR genes ( ATM, BRCA1, BRCA2, BARD1, BRIP1, CDK12, CHEK1, CHEK2, FANCL, PALB2, RAD51B, RAD51C, RAD51D, RAD54L, PPP2R2A) were selected. 495 patients with WES data from the TCGA datasets were included for analysis. Results: In the Chinese cohort, 10.46% (70/669) patients exhibited somatic HRR (sHRR) gene mutations. ATM (4%) was the most common mutant gene, followed by BRCA2 (3%), BARD1 (1%), BRCA1 (1%), BRIP1 (1%), RAD51D (1%), CHEK2(1%). The germline HRR (gHRR) gene mutations were identified in 55.67% (118/245) patients. BRCA2 (23%) was the most common mutant gene, followed by ATM (10%), BRCA1 (9%), BRIP1 (9%), PALB2 (8%), BARD1 (7%), FANCL (3%). 1.49% (10/669) patients carried both germline and somatic HRR gene mutations. The sHRR-Mut patients had a higher median TMB compared to the sHRR-WT patients (3.97 vs 2.13, p&lt;0.001). However, the median TMB was similar between the gHRR-Mut and the gHRR-WT patients (2.13 vs 2.13, p=0.49). In the TCGA cohort, 9.09% (45/495) patients exhibited sHRR gene mutations. The most common mutant gene was ATM (4%), followed by CDK12 (2%) BRCA2 (2%), BARD1 (1%). The sHRR-Mut patients had a higher median TMB compared to the sHRR-WT patients (1.03 vs 0.71, p&lt;0.001). Totally, the somatic mutant frequency of HRR genes was similar between Chinese cohort and TCGA cohort (10.46% vs 9.09%, p = 0.113). For the TCGA cohort, the patients with sHRR-Mut had a similar median progression free survival compared to the ones with sHRR-WT (74.4 months vs not reached, p = 0.299). Conclusions: Our data reported the genetic landscape of HRR gene in Chinese pancreatic cancer patients. Patients with somatic HRR gene mutations had a significantly elevated TMB level. Our data suggest that patients with altered HRR genes may be rational candidates for precision oncology treatment and provide new opportunities to predict the tumor response to multiple treatments.

Genomic characterization of FGFR-altered solid tumors using a clinically annotated pan-cancer repository.

3074 Background: Somatic mutations, gene fusions, and amplifications in FGFR1/2/3 are found across solid tumors. FGFR2/3 fusions and select mutations have matched FDA-approved therapies in intrahepatic cholangiocarcinoma (iCCA) and urothelial carcinoma (UC). Identification of FGFR-altered tumors may inform patient selection for future drug development. Methods: All solid tumors that underwent MSK-IMPACT, a DNA-based NGS panel with coverage of FGFR1-4, were queried for FGFR1-4 alterations. FGFR structural variants (SV) were manually reviewed and compared to RNA sequencing data (RNAseq, Archer FusionPlex) when available. Alteration (alt) frequencies of iCCA and UC tumors with target FGFR alt (level 1 OncoKB evidence), non-target FGFR alt (lacking level 1 evidence) and no FGFR alt were compared using chi-squared tests and odds ratios. Euclidean distance was calculated between cohorts based on prevalence of frequently altered genes (&gt;5%). Consensus clustering was used to identify optimal number of clusters and compare genomic profiles between clusters. Results: At data lock of June 1, 2022, 5695 tumors among 48,416 patients harbored an FGFR alteration (11.8%). FGFR SV were identified in 347 tumors on MSK-IMPACT. Of these 111/140 (79.2%) with RNAseq were positive. 98% (203/207) of SV without RNAseq were considered likely activating (canonical SV, n=125; in-frame inclusive of kinase domain, n=78). The most frequent fusion was FGFR3-TACC3 (n=124), followed by fusions unique to a single patient (n=100). Fusions were discovered in &gt;15 tumor types, most frequently in iCCA (n=93), UC (n=50) and high-grade glioma (n=44). Among 1176 iCCA tumors, 9.9% (n = 117) had a target FGFR2 fusion. Target FGFR tumors were enriched for concurrent BAP1 alt (18%) compared to tumors without FGFR alt (58/969, 5.9%) and with non-target FGFR alt (7/90, 7.8%). Both groups without target alt were enriched in TP53, ARID1A and IDH1 alts compared to target group. Four optimal genomic clusters were identified, with FGFR2 fusions enriching in cluster 4 (29% vs 3%, 7% and ≤1%); 21 non-target FGFR alt were also in cluster 4. Among 2720 UC tumors, 24.7% (n=672) had target FGFR alt (select FGFR3 mutations and FGFR2/3 fusions). Three genomic clusters were identified, with target FGFR enriched in cluster 3 (42% target FGFR vs cluster 1: 11%, cluster 2: 18%) and non-target FGFR alterations enriched in cluster 1 (13% vs. cluster 2: 9.4 %, cluster 3: 7.0%); 68 non-target FGFR alt were in cluster 3. Tumors with non-target FGFR alterations and no FGFR alterations had close genomic association (Euclidean distance: 0.18). Conclusions: Activating FGFR fusions, which occur across tumor types with marked diversity in fusion partners, present opportunities for future drug development. Non-target FGFR alterations in iCCA and UC, particularly those that cluster with target FGFR alterations, may be candidates for future FGFR-targeted strategies.

Clinical and genomic characteristics between DDR germline and somatic mutations in pan-cancer.

e15139 Background: The defects DNA-damage repair (DDR) genes would drive tumor formation and are associated with increased genomic instability and tumor mutational burden (TMB) in cancer. Although, alterations of DDR genes are common in tumor, the differences between the germline and somatic alterations are poorly characterized. Methods: The DNA sequencing data of 540 genes from 6174 pan-cancer patients were retrospectively collected and 276 DDR genes were analyzed. The variations were annotated as pathogenic (P), likely pathogenic (LP), and nonpathogenic (NP) according to ACMG (American College of Medical Genetics) guideline. The patients were divided into 3 groups: DDR-germline (P&amp;LP germline variants, N = 766), DDR-somatic group (only P&amp;LP somatic variants, N = 2923) and non-DDR group (NP variants, N = 2485). Results: In pan-cancer, 12.41% patients have germline DDR gene mutations, of which tumors with high germline mutation frequency include ovarian cancer (27/102, 26.47%), glioma (22/136, 16.18%), breast cancer (25/159, 15.72%), genitourinary cancer (17/114, 14.91%), biliary tract cancer (42/284, 14.79%), pancreatic cancer (28/193, 14.51%), colorectal cancer (95/727, 13.07%), head-neck tumors (11/85, 12.94%) and esophagus cancer (16/124, 12.90%). Meanwhile, 47.34% patients have somatic DDR gene mutations, of which tumors with high somatic mutation frequency include esophagus cancer (89/124, 71.77%), colorectal cancer (488/727, 67.13%), endometrial cancer (49/73, 67.12%), head-neck tumors (50/85, 58.82%), pancreatic cancer (111/193, 57.51%), gastric carcinoma (233/415, 56.14%), ovarian cancer (53/102, 51.96%) and biliary tract cancer (138/284, 48.59%). The most commonly germline alterations were found in TP53 (33%), PTEN (14%) and BRCA2 (14%), while in the somatic mutations, TP53 (52%) showed the highest frequency, PTEN (9%) and MSH3 (8%) followed. In pairwise comparisons of the three groups’ actionable mutations, DDR-somatic group was more likely to have alterations in TP53 (OR = 5.68), AKT3 (OR = 5.81) and APC (OR = 1.42) compared with DDR-germline group. Taken non-DDR group as reference, mutations in TP53 (OR = 5.09), PTEN (OR = 7.65), BRCA2 (OR = 5.32) were more common in DDR-germline group, but TP53 (OR = 28.90), PTEN (OR = 7.73), APC (OR = 2.68) in DDR-somatic group. The tumor mutational burden (TMB) was statistically different among the three groups (p &lt; 2.22e−16). The DDR-somatic group exhibits the highest median TMB (5.68) compared with 1.42 in DDR-germline and 2.13 in non-DDR group. Conclusions: The differences in the mutation profile between the DDR-germline and DDR-somatic groups and the distinct actionable genes may indicate the different target-therapy choices. Besides, DDR-somatic group exhibited the highest TMB, which might indicate patients with somatic DDR alterations may better benefit from the immunotherapy.

Somatic hereditary gene panel analysis: A pilot study from Uzbekistan.

e15044 Background: Molecular diagnostics is gaining importance in routine clinical practice. Availability of actionable genetic changes in hereditary panel of genes has acquired even larger interest. This study is an attempt evaluate the limited somatic genetic alterations in small population of Uzbek patients. Methods: Somatic tumor sample material was analyzed using FOUNDATIONONER CDx (F1CDx) - the NGS based in vitro diagnostic device for detection of substitutions, insertion and deletion alterations(indels), and copy numbers alterations (CNAs) in 324 genes and select gene rearrangements, as well as genomic signatures. Our focus of hereditary panel included following genes-TP53, CDH1, PTEN, STK11, PALB2, CHEK2, ATM, NBN, BARD1, BRCA1 and BRCA2. Results: Out of 76 patients analyzed 50 were found have at list one abnormal gene from the above hereditary panel. TP53 was frequent pathogenic mutation in 50 of 76 patients. Commonest hereditary gene alterations were seen in ATM gene -12/50(6%). BRCA2 mutations existed in 7 of 50(3,5%), PTEN – 6/50(3%), PALB-2 – 4/50(2%), CHEK-2 – 5/50(2,5%), NBN – 5/50(2,5%), STK11 – 3/50(1,5%), CDH1 – 2/50(1%), BRCA1-2(1%), BARD1 – 1/50(0,5%). Some patients had multiple gene combinations. One of the patients had four gene abnormality-PTEN, PALB2, CHEK2, and BRCA2.Three patients have 3(1,5%) somatic hereditary mutations, 23(11,5%) patients have two abnormal hereditary genes and 38(19%) patients had changes in single gene only. Important actionable mutations (BRCA, PALB2 and CHEK2 were found in 15(11,4%) patients. Conclusions: This is the first exploratory analysis of somatic hereditary multi-gene panel. About 11% of the patients were found to have actionable gene mutations.

The spectrum of homologous recombination deficiency in cervical cancer: Is it also a cervical issue?

5543 Background: Tumors deficient in homologous recombination DNA damage repair (HR-DDR) demonstrate sensitivity to therapeutic agents targeting the HR-DDR pathway, such as poly-ADP ribose phosphate (PARP) inhibitors. The landscape of homologous recombination deficiency (HRD) is not well-characterized for cervical cancer; thus, we sought to analyze the prevalence of somatic pathogenic gene variants (PGVs) in HRD genes in cervical cancers. Methods: The American Association for Cancer Research’s (AACR) Project Genomics Evidence Neoplasia Information Exchange (GENIE) database version 13.0 was queried for all cervical cancer tumors via cBioPortal (http://genie.cbioportal.org). This is a publicly available, multi-institutional database of next-generation sequencing (NGS) genomic profiles of multiple tumors. PGV frequencies of 27 genes involved in HR-DDR were descriptively reported for cervical cancer tumors, with histologic stratification: ATM, ARID1A, ATRX, BRCA1, BRCA2, BARD1, BRIP1, BLM, BAP1, CHEK1, CHEK2, FANCA, FANCC, FANCD2, FANCE, FANCF, FANCG, FANCL, MRE11, NBN, PALB2, RAD50, RAD51, RAD51B, RAD51C, RAD51D, WRN. Results: A total of 857 cervical tumors from 817 patients were included for analysis. At least one somatic PGV in an HRD gene was found in 16.5% of all tumors (141/857). When stratified by histological subtypes with at least 25 tumors sequenced, highest frequencies of ≥1 PGV in HRD genes were observed in mucinous cervical cancer (13/66, 19.7%), cervical adenocarcinoma (48/262, 18.3%), squamous cell cervical cancer (53/349, 15.2%), adenosquamous cervical cancer (6/43, 14.0%) and neuroendocrine cervical cancers (5/42, 11.9%). Substantial rates of PGVs in HRD genes were also observed in histological subtypes with fewer than 25 sequenced tumors. Across all tumors, HRD genes with the highest frequencies of PGVs were ARID1A (9%), BAP1 (3%), ATM (1.6%) and BRCA1 (1.6%). Conclusions: NGS data demonstrate a substantial rate of somatic PGVs in HRD genes in cervical cancers, including among HPV related subtypes. These data suggest the need to expand routine functional HRD status assessment to cervical cancers in order to further characterize the landscape of HRD in these tumors. Furthermore, genetically driven clinical trials are warranted to evaluate the efficacy of HRD-targeted therapies such as PARP inhibitors, ataxia telangiectasia, and Rad-3 related kinase (ATR) inhibitors. [Table: see text]

The impact of cause of mismatch repair deficiency and other molecular markers on clinical outcomes with the use of durvalumab in advanced endometrial cancer in the phase 2 PHAEDRA trial (ANZGOG1601).

5604 Background: In the single arm phase 2 PHAEDRA trial, MMR deficiency (dMMR) was predictive of response to durvalumab (1500mg IV Q4W), with an objective tumor response rate (OTR; defined by iRECIST) of 47% in dMMR compared with 3% in MMR-proficient (pMMR) advanced endometrial cancer (AEC). This substudy of the PHAEDRA trial investigates MMR molecular subtypes and other genomic tumor features and their correlation with treatment outcomes. Methods: Testing was performed to determine molecular subtypes of dMMR, including germline pathogenic variants in DNA MMR genes (Lynch syndrome), somatic biallelic MMR gene inactivation through somatic mutation and somatic hypermethylation of the MLH1 gene promoter. DNA from formalin-fixed paraffin-embedded tumor tissue and matched peripheral blood was available from 41/71 (25 dMMR, 16 pMMR) participants for testing on a custom capture-based 298-gene targeted panel (2.005Mb) including the MMR genes and key somatic AEC driver genes. The derived tumor genomic features included microsatellite instability (MSI), tumor mutational burden (TMB), COSMIC v.3.2 tumor mutational signatures and insertion/deletion (Indel) somatic mutation count. Results: Of the 71 patients recruited, 35 were dMMR and 36 were pMMR. Median follow-up was 44 vs 52 months in dMMR vs pMMR participants, respectively. The dMMR molecular subtypes were 4 (11.4%) Lynch syndrome, 4 (11.4%) somatic MMR mutation, 25 (71.4%) MLH1 methylated and 2 (5.7%) dMMR-uncategorised. The OTR rate was 100% (4/4; 95%CI 40-100%) for Lynch, 75% (3/4; 95%CI 22-99%) for somatic MMR mutations and 40% (10/25; 95%CI 22-61%) for MLH1 methylated groups. The median TMB (assessed in 41/71) was higher in those with a confirmed radiological response (37, IQR:26-50) vs non-responders (16, IQR:9-25; p &lt; 0.001). Within the MLH1 methylated group, TMB was also higher in responders vs non-responders (40 v 21; p = 0.03). Somatic mutations in KRAS, PTEN, PIK3CA, ARID1A and TP53 were not associated with OTR rate. Conclusions: dMMR- MLH1 methylated AEC demonstrated greater heterogeneity in OTR to single agent durvalumab than the dMMR-Lynch and dMMR-somatic MMR mutation molecular subtypes. Higher TMB was seen in responders, and specifically within dMMR- MLH1 methylated responders, compared to non-responders. Clinical trial information: ANZGOG1601.

Reproduction of Sheep through Nuclear Transfer of Somatic Cells: A Bibliometric Approach.

Somatic cell nuclear transfer (SCNT) is a reproductive biotechnology with great potential in the reproduction of different species of zootechnical interest, including sheep. This study aimed to carry out a bibliometric analysis of scientific papers published on the application of SCNT in sheep reproduction during the period 1997-2023. The search involved the Science Citation Index Expanded and Social Sciences Citation Index databases of the main collection of the Web of Sciences with different descriptors. A total of 124 scientific papers were analyzed for different bibliometric indicators using the VOSviewer software. Since 2001, the number of SCNT-related papers that have been published concerning sheep reproduction has increased and it has fluctuated in ensuing years. The main authors, research groups, institutions, countries, papers, and journals with the highest number of papers related to the application of SCNT in sheep reproduction were identified, as well as the topics that address the research papers according to the terms: somatic cell, embryo, oocyte, gene expression, SCNT, and sheep.

Description of technique for endoscopic ultrasound-guided tissue acquisition in pancreatic cancer for comprehensive molecular profiling: Results of a randomized trial.

e16306 Background: As theoretically actionable genomic lesions are found in almost 50% of pancreatic cancers which can impact clinical management in up to 30% of patients, there is increased focus on molecular profiling. Although EUS-FNB is an established method for achieving pathological diagnosis in pancreatic cancers, the technique of tissue acquisition for comprehensive molecular profiling (CMP) has not been described. Methods: Patients proven to have pancreatic adenocarcinoma using rapid onsite evaluation (ROSE) at EUS were randomized to undergo two or three dedicated FNB passes for CMP. Tissue was procured using 22G Franseen needle adopting the fanning technique and stylet-retraction maneuver. Genomic DNA and total RNA were extracted from formalin-fixed paraffin-embedded (FFPE) cell blocks (Qiagen AllPrep). Next-generation sequencing (NGS) was performed (Archer Panels) for analyzing 69 gene DNA mutations (single nucleotide variants, insertions, and deletions) and 53 RNA somatic oncogenic gene fusions. Main outcome measure was the proportion of specimens in which adequate DNA and RNA were extracted for CMP. Results: 33 patients (mean age 77.0 years [SD 7.7], female = 20) with pancreatic cancer (location = head/uncinate 22; other 11) were randomized to two (n = 17) or three (n = 16) FNB passes. While sufficient DNA was extracted from all 33 FFPE cell blocks, adequate RNA was extracted from 15 of 16 in the three pass (93.8%) vs. 16 of 17 in the two pass cohort (94.1%) (p = 0.99). There was no significant difference in the mean concentration of DNA (two passes 10.7 ng/ul [SD 7.1] vs. three passes 7.9 ng/ul [SD 4.4]; p = 0.19) or RNA (two passes 37.1 ng/ul [SD 26.5] vs. three passes 28.9 ng/ul [SD 13.2]; p = 0.29) between the groups. While somatic oncogene RNA fusion (LDAH-ETV1) predictive of metastatic disease was identified in one specimen, DNA mutations were identified in all 33 specimens that included one BRCA1 mutation that warranted Oxaliplatin-based chemotherapy. Conclusions: Specimens of adequate quantity and of high quality for comprehensive molecular profiling in pancreatic cancer can be procured in nearly 95% of patients by performing two dedicated passes using the 22G Franseen needle, adopting the fanning maneuver and stylet-retraction technique. Clinical trial information: NCT05043532 .

Homologous recombination-related (HRR) gene mutation profiling and the association with TMB in Chinese patients with urological cancer.

e16561 Background: Homologous recombination deficiency (HRD) is associated with tumorigenesis and could predict the response of PARP inhibitor (PARPi) therapy and immunotherapy. Loss of function or deleterious mutations in homologous recombination-related (HRR) genes contribute to HRD phenotype. Methods: Tumor tissue samples from 484 Chinese patients with urological cancer were sequenced with 551 cancer-related genes. 957 patients with WES data from the TCGA datasets were included for analysis. Results: In the Chinese cohort, 35.88% (47/131) bladder cancer (BCa) patients, 15.47% (28/181) renal cell carcinoma (RCC) and 37.21% (64/172) prostate cancer (PCa) patients exhibited somatic HRR (sHRR) gene mutations. The most frequently mutated genes were BRCA2 (12% in BCa, 5% in RCC) and CDK12 (17% in PCa), respectively. The germline HRR (gHRR) gene mutations were identified in 62.6% (82/131) BCa, 55.33% (96/180) RCC and 60.62% (97/160) PCa patients. The most frequently mutated gene was BRCA2 (24% in BC, 18% in RCC, 26% in PCa). 7.6% (10/131) BCa patients, 1.7% (3/181) RCC patients and 2.9% (5/173) Pca patients carried both germline and somatic HRR gene mutations. In the TCGA cohort, 38.69% (159/411) BCa patients, 11.62% (43/370) RCC and 6.25% (11/176) PCa patients exhibited somatic HRR (sHRR) gene mutations. The most frequently mutated gene was ATM (14% in BCa, 4% in RCC, 5% in PCa). Interestingly, for the patients with PCa, the somatic mutant frequency of HRR genes in Chinese cohort was higher than that in TCGA cohort (37.21% vs 6.25%, p &lt; 0.001). The sHRR-Mut patients had a higher median TMB compared to the sHRR-WT patients (Chinese cohort: BCa 13.97 vs 4.96, p &lt; 0.001; RCC, 4.96 vs 2.84, p &lt; 0.001; PCa, 5.64 vs 2.84, p &lt; 0.001, TCGA cohort: BCa, 7.21 vs 3.63, p &lt; 0.001; RCC, 1.68 vs 1.42, p = 0.003; PCa, 1.03 vs 0.71, p &lt; 0.001). However, the median TMB was similar between the gHRR-Mut and the gHRR-WT patients (Chinese cohort, BCa 6.38 vs 7.09 p = 0.646; RCC, 3.55 vs 2.84, p = 0.229; PCa, 3.55 vs 2.84, p = 0.111). Conclusions: Our data reported the genetic landscape of HRR gene in Chinese urological cancer patients and suggested that patients with altered HRR genes may be rational candidates for precision oncology treatment and provide new opportunities to predict the tumor response to multiple treatments.

Distinguishing primary mucinous ovarian tumors from metastases of non-gynecologic mucinous cancers: Can we leverage next-generation sequencing?

5582 Background: Primary mucinous ovarian cancers (MOC) are histopathologically challenging to differentiate from ovarian metastases of non-gynecologic origin, with this distinction being critical for appropriate management and prognosis. We compared the somatic gene variant landscape of MOC to that of non-gynecologic mucinous tumors. Methods: Data were extracted from the American Association for Cancer Research’s (AACR) Project Genomics Evidence Neoplasia Information Exchange (GENIE) database version 13.0 via cBioPortal. This publicly available, multi-institutional database provides next generation sequencing (NGS) genomic profiles of tumors. We queried this database for samples of MOC, mucinous colorectal cancer (MCRC), mucinous appendiceal cancer (MAC), mucinous breast cancer (MBC) and gastric type mucinous cancer (GMC). Frequencies of somatic gene variants including mutations, copy number alterations and structural variants were compared using Chi-squared or Fischer’s exact tests, using the Benjamini-Hochberg method to control for multiple hypothesis testing with q-values reported. Results: A total of 883 tumors were included for analysis: 358 MCRC, 268 MAC, 157 MOC, 59 MBC and 41 GMC samples. Compared to MAC, MOC samples had higher variant frequencies of CDKN2A (33.3% vs 0.4%, q&lt;0.001), CDKN2B (24.0% vs 0.0%, q&lt;0.001), TP53 (64.3% vs 23.5%, q&lt;0.001), ERBB2 (14.3% vs 1.1%, q&lt;0.001) and CDK12 (13.2% vs 0.0%, q&lt;0.001), whereas GNAS variants were more common in MAC (45.5% vs 5.7%, q&lt;0.001). Compared to MCRC, MOC samples had higher variant frequencies of CDKN2A (33.3% vs 2.3%, q&lt;0.001), CDKN2B (24.0% vs 3.7%, q&lt;0.001), TP53 (64.3% vs 42.9%, q&lt;0.001), KRAS (69.4% vs 51.1%, q&lt;0.001) and ERBB2 (14.3% vs 5.3%, q&lt;0.001), whereas MCRC had higher variant frequencies of 57 genes, with the largest differentials among APC (48.8% vs 2.6%, q&lt;0.001), SMAD4 (25.2% vs 5.8%, q&lt;0.001) and TCF7L2 (19.0% vs 0.0%, q&lt;0.001). Samples of MOC had significantly higher rates of KRAS variants compared to GMC (69.4% vs 31.7%, q&lt;0.001) and lower rates of STK11 variants (1.9% vs 22.0%, q&lt;0.001). Compared to MBC, MOC samples had higher variant rates of CDKN2A (33.3% vs 3.4%, q&lt;0.001), TP53 (64.3% vs 10.2%, q&lt;0.001) and KRAS (69.4% vs 0.0%, q&lt;0.001), whereas MBC samples had higher variant frequencies of 11 genes, with the largest differentials among GATA3 (32.1% vs 0.8%, q&lt;0.001), FGF3 (30.4% vs 2.4%, q&lt;0.05) and CCND1 (28.1% vs 1.6%, q&lt;0.001). Conclusions: NGS demonstrates that MOCs carry a distinct genetic signature compared to mucinous tumors of non-gynecologic origin, most commonly with significantly higher variant frequencies of CDKN2A, CDKN2B and lower variant frequencies of GNAS, APC, STK11 and GATA3. This provides rationale for prospective studies evaluating genetic signatures as an adjunct to histopathology in the diagnosis of primary MOC.

Genomic comparison of MET exon 14 skipping and MET amplified non-small cell lung cancer.

8553 Background: In non-small cell lung cancer (NSCLC), the MET tyrosine kinase receptor can be dysregulated by mutations and/or gene amplification. The most common MET mutation is in exon 14 (METex14), leading to impaired receptor degradation and increased MET-mediated signaling causing sustained tumor proliferation. MET amplification also leads to continued MET signaling and oncogenesis and can be a mechanism of resistance to targeted therapy. We sought to compare the genomic landscape of METex14 and high-level MET amplified tumors, both of which can be targeted with tyrosine kinase inhibitors. Methods: We analyzed 18,047 NSCLC tumors (any stage/subtype) sequenced with Tempus xT assay (DNA-seq of 648 genes at 500x coverage, full transcriptome RNA-seq). Tumors were queried for METex14 mutations, high MET amplification (METamp), defined as copy number gain (CNG) ≥10, and other MET mutations (METother). Immuno-oncology (IO) biomarkers were compared across MET-altered and MET wild-type (METwt) groups. The prevalence of somatic gene alterations was compared similarly with the false-discovery rate (FDR) adjustment. Results: A total of 276 (1.53%) METex14, 138 (0.76%) METamp, 27 (0.15%) METother, and 17,606 (97.56%) METwt tumors were identified. Across groups, patients with METex14 were older, more likely to be female, and nonsmokers. METex14 exhibited the lowest tumor mutational burden (TMB) and lowest neoantigen tumor burden (NTB). PD-L1 positivity rates were higher in METex14 tumors compared to METamp and METwt tumors. MET log10 gene expression was highest in METamp. METamp exhibited the lowest proportion of CD4 T cells and the highest proportion of NK cells. Compared to METex14, METamp exhibited increased prevalence of TP53 (83% vs 37%), TFEC (46% vs 1.1%), CFTR (37% vs 1.4%), EGFR (31% vs. 7.6%), WNT2 (11% vs 0.4%), KEAP1 (14% vs 0.7%), and STK11 (8% vs 1.4%), and decreased prevalence of MDM2 (4.3% vs. 14%), and FRS2 (2.9% vs. 12%). Conclusions: In this large population-based analysis of MET-altered NSCLC, METex14 tumors exhibited differences in IO biomarkers and the somatic landscape compared to non-METex14 NSCLC tumors. Variations in immune profiles may affect immunotherapy selection in MET-altered NSCLC and require further exploration. [Table: see text]

Biomarkers of response to sacituzumab govitecan (SG) and efficacy after treatment with enfortumab vedotin (EV) in advanced urothelial carcinoma (aUC): Analysis of the UNITE study.

4572 Background: SG is an antibody-drug conjugate (ADC) with accelerated FDA approval for aUC refractory to platinum-based chemotherapy and immune checkpoint inhibitor (ICI). SG activity following treatment with EV, the most commonly used ADC in the post-platinum/ICI setting, and biomarkers of response to SG are not well described. We hypothesized that SG would be active after EV in aUC. Methods: Patients (pts) treated with SG monotherapy who had next-generation sequencing (NGS), were identified in the retrospective UNITE study. Observed response rate (ORR) was determined for evaluable pts with scans after ≥1 SG cycle. All pts were assessed for baseline clinical characteristics, as well as molecular biomarkers including tumor mutational burden, somatic gene alterations (alt) in ≥ 10% of pts and presence of ≥1 DNA damage response (DDR) mutations. ORRs were compared with Chi-squared test. Kaplan Meier method and cox proportional hazard (cph) models were used to assess progression-free survival (PFS) and overall survival (OS) from SG start. Pt characteristics and molecular biomarkers were evaluated in univariate analyses (UVA), followed by evaluation of biomarkers in separate multivariate cph analyses (MVA), while accounting for clinical features. Results: Among 90 pts treated with SG at 9 US sites, 78 had NGS data. Median age at SG start was 68, 65 (72%) men, 73 (81%) Caucasian, 56 (62%) with pure urothelial histology, 60 (67%) primary bladder tumor and 67 (74%) ECOG PS 0/1. Most (n=84, 93%) received SG after EV, and 33 (37%) had ≥4 prior therapies for aUC. ORR with SG was 23% (15/66); 24% in pts with prior CR/PR/SD on EV and 14% after primary PD on EV. Median follow-up from SG start was 8.7 mos, PFS and OS were 3.5 mos and 7.1 mos. In pts with NGS, ORRs to SG were higher in MTAP altered pts (50% vs 19%; p = 0.05). In UVA, low neutrophil to lymphocyte ratio (NLR), high Hgb, no prior smoking history, &lt; 3 prior therapies, as well as BRCA2 and DDR alts were associated with longer OS (p&lt;0.05 for all). In MVA, pts with alt in either TP53 or MDM2 had longer OS. Conclusions: SG has notable activity after EV in heavily pretreated pts with aUC, with ORR consistent with SG phase 2 trial data in post-platinum/ICI only pts. We identified several clinical and genomic biomarkers, including TP53/MDM2 alt, that are potentially associated with improved outcomes with SG treatment. [Table: see text]