Distinct Genomic Landscape Research Articles

The genomic landscape of papillary thyroid carcinoma on next-generation sequencing in patients undergoing total thyroidectomy.

Thyroidectomy is increasingly performed for suspected malignancy. This cohort study aimed to identify genetic markers associated with malignancy and determine the molecular landscape of papillary thyroid carcinoma (PTC) through next-generation sequencing (NGS) in patients undergoing total thyroidectomy. Among 116 surgical candidates, 103 patients (age=42.9±13.7years; Male: Female=14:89) with benign or malignant thyroid nodules were eligible. Live thyroid tissue samples harvested intraoperatively with adequate DNA and RNA yield were subjected to NGS on the Illumina NovaSeq 6000 platform for genomic and transcriptomic analysis, respectively. Histopathology comprised 20 malignant (19.4%) and 83 benign (80.6%) cases, including 16 PTC (15.5%) cases. On NGS, single nucleotidepolymorphisms (SNPs) in NTRK1 at NC_000001.11:156879016 on chromosome 1 and ALK at NC_000002.12:29717663 on chromosome 2 were frequent in malignant lesions (p<0.05). A SNP in ALK at NC_000002.12:29193706 was consistently a homozygous alternate alleleacross the cohort. DNA-sequencing of PTC lesions identified recurrentsomatic mutations in BRAF (100%), ALK (56.3%), RET (18.8%), PIK3CA (12.5%), NTRK1 (12.5%), NTRK2 (87.5%), NTRK3(12.5%), NRAS(6.3%), and PTCH1 (31.3%) genes. RNA sequencing revealednovelfusiongenes, including MKRN1-BRAF, RN7SL1-CDH1, IRF2BPL-MED12, MED12-IRF2BPL, CPM-MDM2, and AC005895.3-PDGFRB. Inreceiveroperative characteristics analysis, the AUCs ofALKmutationpredictingrecurrence and metastases were 0.818 and 0.783. This Indian study identified novel somatic mutations and fusion genes in PTC, revealing a distinct genomic landscape with implications in precision diagnostics and personalized therapies. NGS with intraoperative live sampling shows promise in prognostication and therapeutic optimization of advanced/metastatic PTC cases. CTRI/2020/09/027607dt 04/09/2020; REF/2020/08/036119.

Large-scale analysis of CDH1 mutations defines a distinctive molecular subset with treatment implications in gastric cancer.

Although histological and molecular classifications have been extensively studied for gastric cancer (GC), targeted therapies for GC remain limited. CDH1 mutations (MT) are characteristic of genomically stable GC and are associated with poor prognosis, but lack effective or targeted therapies. Here, we showed the overall mutation frequency of CDH1 was 9.7% (155 of 1596). CDH1-MT GC showed significantly lower rates of PD-L1 positivity (CPS score ≥1) than CDH1-wildtype (WT) GC (56.7% vs. 73.3%, p < 0.05). Compared to CDH1-WT GC, mutations of ARID1A, WRN, POT1, CDK12, and FANCC were significantly higher, while TP53 and APC were significantly lower in CDH1-MT GC (p < 0.05); The rates of KRAS and HER2 amplifications were significantly lower, while CRKL and IGF1R amplifications were significantly higher in CDH1-MT GC, compared to CDH1-WT GC (p < 0.05). Frequently altered genes in CDH1-MT GC were especially enriched in DNA damage repair and cell cycle checkpoint pathways. Inhibition of E-cadherin sensitized GC cell lines to PARP and Wee1 inhibitors by disrupting DNA damage repair pathway and cell cycle checkpoint. This is the largest study to investigate the distinct genomic landscape of CDH1-MT GC. Our data indicated GC patients with CDH1 mutations could potentially benefit from agents targeting PARP and Wee1.

Local cryptic diversity in salinity adaptation mechanisms in the wild outcrossing Brassica fruticulosa

It is normally supposed that populations of the same species should evolve shared mechanisms of adaptation to common stressors due to evolutionary constraint. Here, we describe a system of within-species local adaptation to coastal habitats, Brassica fruticulosa, and detail surprising strategic variability in adaptive responses to high salinity. These different adaptive responses in neighboring populations are evidenced by transcriptomes, diverse physiological outputs, and distinct genomic selective landscapes. In response to high salinity Northern Catalonian populations restrict root-to-shoot Na+ transport, favoring K+ uptake. Contrastingly, Central Catalonian populations accumulate Na+ in leaves and compensate for the osmotic imbalance with compatible solutes such as proline. Despite contrasting responses, both metapopulations were salinity tolerant relative to all inland accessions. To characterize the genomic basis of these divergent adaptive strategies in an otherwise non-saline-tolerant species, we generate a long-read-based genome and population sequencing of 18 populations (nine inland, nine coastal) across the B. fruticulosa species range. Results of genomic and transcriptomic approaches support the physiological observations of distinct underlying mechanisms of adaptation to high salinity and reveal potential genetic targets of these two very recently evolved salinity adaptations. We therefore provide a model of within-species salinity adaptation and reveal cryptic variation in neighboring plant populations in the mechanisms of adaptation to an important natural stressor highly relevant to agriculture.

Genomic characterization reveals distinct mutational landscapes and therapeutic implications between different molecular subtypes of triple-negative breast cancer

Triple-negative breast cancer (TNBC) has high heterogeneity, poor prognosis, and limited treatment success. Recently, an immunohistochemistry-based surrogate classification for the “Fudan University Shanghai Cancer Center (FUSCC) subtyping” has been developed and is considered more suitable for clinical application. Seventy-one paraffin-embedded sections of surgically resected TNBC were classified into four molecular subtypes using the IHC-based surrogate classification. Genomic analysis was performed by targeted next-generation sequencing and the specificity of the subtypes was explored by bioinformatics, including survival analysis, multivariate Cox regression, pathway enrichment, Pyclone analysis, mutational signature analysis and PHIAL analysis. AKT1 and BRCA1 mutations were identified as independent prognostic factors in TNBC. TNBC molecular subtypes encompass distinct genomic landscapes that show specific heterogeneities. The luminal androgen receptor (LAR) subtype was associated with mutations in PIK3CA and PI3K pathways, which are potentially sensitive to PI3K pathway inhibitors. The basal-like immune-suppressed (BLIS) subtype was characterized by high genomic instability and the specific possession of signature 19 while patients in the immunomodulatory (IM) subtype belonged to the PD-L1 ≥ 1% subgroup with enrichment in Notch signaling, suggesting a possible benefit of immune checkpoint inhibitors and Notch inhibitors. Moreover, mesenchymal-like (MES) tumors displayed enrichment in the receptor tyrosine kinase (RTK)-RAS pathway and potential sensitivity to RTK pathway inhibitors. The findings suggest potential treatment targets and prognostic factors, indicating the possibility of TNBC stratified therapy in the future.

Characterization and impact of non-canonical WNT signaling on outcomes of urothelial carcinoma.

Non-canonical WNT family (WNT5A pathway) signaling via WNT5A through ROR1 and its partner, ROR2, or Frizzled2 (FZD2) is linked to processes driving tumorigenesis and therapy resistance. We utilized a large dataset of urothelial carcinoma (UC) tumors to characterize non-canonical WNT signaling through WNT5A, ROR1, ROR2, or FZD2 expression. NextGen Sequencing of DNA (592 genes or WES)/RNA (WTS) was performed for 4125 UC tumors submitted to Caris Life Sciences. High and low expression of WNT5A, ROR1, ROR2, and FZD2 was defined as ≥ top and <bottom quartile of transcripts per million (TPM), respectively. Gene expression profiles were analyzed for a transcriptional signature predictive of response to immunotherapy. Mann-Whitney U and X2/Fisher Exact tests were applied where appropriate, with p-values adjusted for multiple comparisons (p < 0.05). Real-world overall survival (OS) was obtained from insurance claims data. WNT5A pathway gene expression varied significantly between primary versus metastatic sites: WNT5A (25.2 vs. 16.8 TPM), FZD2 (3.2 vs. 4.05), ROR1 (1.7 vs. 2.1), and ROR2 (2.4 vs. 2.6) p < 0.05 for all. Comparison of high- and low-expression subgroups revealed variation in the prevalence of TP53, FGFR3, and RB1 pathogenic mutations, as well as increasing T cell-inflamed scores as expression of the target gene increased. High gene expression for ROR2 (HR 1.31, 95% CI 1.15-1.50, p < 0.001) and FZD2 (HR 1.16, 95% CI 1.02-1.32, p = 0.024) was associated with worse OS. Distinct genomic and immune landscapes for the four investigated WNT5A pathway components were observed in patients with UC. External validation studies are needed.

Assessing the genomic feature of Chinese patients with ampullary adenocarcinoma: potential therapeutic targets

BackgroundsAmpullary adenocarcinoma (AMPAC) is a rare malignancy, treated as pancreatic or intestinal cancer based on its histologic subtype. Little is known about the genomic features of Chinese patients with AMPAC.Materials and methodsWe enrolled 145 Chinese AMPAC patients in our local cohort and performed a compressive somatic and germline genetic testing using a 156 gene panel. Expression of PD-L1 (clone 28 − 8) was also assessed in tumor specimens from 64 patients.ResultsThe frequency of genetic alterations (GAs) in Chinese patients with AMPAC was found to be distinctive, with TP53, KRAS, SMAD4, APC, CTNNB1, ARID1A, and CDKN2A emerged as the most frequently mutated genes. Comparing with Western patients, significant differences were observed in the prevalence of PIK3CA and ARID2. Furthermore, the incidence of MSI-H was lower in the Chinese cohort, with only two patients identified as MSI-H. Conversely, 11 patients (8.27%) had pathogenic/likely pathogenic germline alterations, all of which were in the DNA damage response (DDR) pathway. In our cohort, 34.48% (22/64) of patients exhibited positive PD-L1 expression in tumor cells, and this expression was associated with GAs in CTNNB1 and BLM. Importantly, over three-fourths of Chinese AMPAC patients in our study had at least one actionable GA, with more than one-fifth of them having actionable GAs classified as Level 3. These actionable GAs were primarily involved in the DDR and PI3K pathways. Notably, GAs in the DDR pathway were detected in both Chinese and Western patients, and regardless of their functional impact, these alterations demonstrated enhanced overall survival rates and higher tumor mutational burden (TMB) levels.ConclusionThese findings underscore the distinct genomic landscape of Chinese AMPAC patients and highlight the potential for targeted therapies based on the identified GAs.

Genomic analysis of oesophageal carcinoma (EC) to identify recurrent mutations in histone methyltransferases as a distinctive subset.

379 Background: Histone-lysine N-methyltransferase 2 (KMT2) family proteins methylate lysine 4 on the histone H3 tail at important regulatory regions in the genome and thereby impart crucial functions through modulating chromatin structures and DNA accessibility, which is associated with tumorigenesis and immune tolerance, indicating its possible correlation with the efficacy of immunotherapy. Recurrent mutations of KMT2 have been identified in EC, but data addressing the molecular features of KMT2 mutated (MT) EC are lacking. We aimed to understand the molecular profile of KMT2 -MT EC. Methods: A total of 787 oesophageal carcinoma [adenocarcinoma (EAC), N=604; squamous cell carcinoma (ESCC), N=183] were analyzed using next-generation sequencing (NGS) and immunohistochemistry (Caris Life Sciences, Phoenix, AZ). Tumor mutational burden (TMB) was calculated based on somatic nonsynonymous mutations, and mismatch repair deficiency (dMMR)/microsatellite instability-high (MSI-H) status was evaluated by a combination of IHC, Fragment analysis and NGS. Results: The mutation spectrum of KMT2A/C/D was significantly different between ESCC and EAC ( KMT2A, 2.7% vs. 1.2%; KMT2C, 3.8% vs. 1.2%; KMT2D, 14.2% vs. 2.0%, P &lt;0.05). KMT2-MT EAC tumors showed significantly higher mutation rates in CIC (13% vs 1.1%), NF1 (12.5% vs 2.2%), FBXW7 (12% vs 3.4%), ATM (11.5% vs 2.6%) and BRCA1 (11.5% vs 1%, all q&lt;0.05), compared to KMT2-WT tumors. However, in ESCC there were no significant differences in gene mutations between the KMT2-MT and WT groups. Significantly higher rates of dMMR/MSI-H (19.2% vs 1.0% in EAC, 5.4% vs. 0% in ESCC, both p&lt;0.01) were seen in both KMT2-MT ESCC and EAC, compared to KMT2-WT cohorts, respectively. TMB was significantly higher in both KMT2-MT ESCC (median: 10 vs 8.6 mut/Mb, P=0.042) and EAC (23.9 vs 8.0 mut/Mb, P&lt;0.0001), compared to KMT2-WT cohorts, respectively. A numerically longer immune-related overall survival was observed in KMT2 -MT EC patients, compared to KMT2 -WT EC patients (9.05 vs 6.98 months, HR=0.744, 95%CI: 0.52-1.07, P =0.11). Conclusions: This is the largest study to investigate the distinct genomic landscapes between KMT2 -MT and WT EC to date. Our data showed the KMT2 -MT EC has a distinctive genetic profile, indicated by higher TMB, and higher frequency of dMMR/MSI-H and gene mutations involved in DDR and epigenetic regulation. Understanding these molecular characteristics may be informative in the development of effective treatment strategies in KMT2 -MT EC.

Distinct genomic landscapes in radiation-associated angiosarcoma compared with other radiation-associated sarcoma histologies.

MYC amplifications have been frequently detected in radiation (RT)-associated angiosarcomas (ASs) by low-resolution molecular methods. However, large-scale next-generation sequencing (NGS) studies to investigate the genomic landscape of RT-AS are scarce, particularly compared with other RT-associated sarcomas. We performed a detailed comparative genomic investigation of RT-AS versus other RT-associated histotypes, as well as sporadic sarcomas with similar histologies. Our institutional targeted DNA-NGS assay database was searched for RT-associated sarcomas. Clinical outcome data, pathologic diagnosis, and the types and frequencies of genomic alterations, including single nucleotide variants (SNVs) and copy number alterations (CNAs), were analyzed. The cohort consisted of 82 patients, 68 (83%) females and 14 (17%) males, aged 37-88 (mean 64) years. Forty-four RT-ASs (38 from breast) and 38 RT sarcomas of other histologies, including 12 malignant peripheral nerve sheath tumors (RT-MPNSTs), 14 undifferentiated pleomorphic sarcomas (RT-UPSs), and 12 osteosarcomas (RT-OSs), were included. Median time intervals from radiation to initial diagnosis in RT-AS (8.0 years) were significantly lower than those in RT-MPNST and RT-UPS (12.5 and 18.5 years), respectively. Each RT-sarcoma histotype harbored distinct mutations and CNAs. RT-associated AS had more frequent MYC, FLT4, CRKL, HRAS, and KMT2D alterations than sporadic AS (enriched in TP53, KDR, ATM, ATRX), whereas the mutational landscapes of MPNST, UPS, and OS were similar in both RT and non-RT settings. CDKN2A/B deletions and TP53 alterations were infrequent in RT-AS compared with other RT sarcomas. Among RT sarcomas, RT-AS harbored the lowest fraction of genome altered (FGA), while RT-MPNST showed the highest FGA. RT-AS had the lowest insertion:SNV and deletion:SNV ratios, while RT-UPS had the highest. The predominant mutational signatures were associated with errors in DNA repair and replication. In conclusion, RT-AS has a distinct genomic landscape compared with other RT sarcomas and sporadic AS. Potential molecular targets for precision medicine may be histotype-dependent. © 2023 The Pathological Society of Great Britain and Ireland.

Response to programmed cell death protein 1 antibody in patients with Epstein-Barr virus-associated biliary tract cancer.

4071 Background: Epstein-Barr virus-associated biliary tract cancer (EBVaBTC) has a distinct genomic landscape and increased infiltration of CD3+ and CD8+ T cells. However, the efficacy of immunotherapy in EBVaBTC remains largely unknown. The aim of this study is to assess the efficacy of programmed cell death protein 1 (PD-1) antibody in EBVaBTC. Methods: Consecutive patients with metastatic biliary tract cancer diagnosed at our institution from January 2017 to December 2021 were identified. In situ hybridization was performed to detect EBV. The objective response to PD-1 antibody was assessed. We also analyzed the immune microenvironment of EBVaBTC by multiplex immunofluorescence staining, and performed the proteomic characterization of EBVaBTC. Results: A total of 131 patients with BTC who received PD-1 antibody were identified, of which 9 (6.9%) had EBVaBTC. All EBVaBTC cases were intrahepatic cholangiocarcinoma. EBVaBTC was not associated with deficient mismatch repair but with lymphoepithelioma-like carcinoma and hepatitis B virus infection. Four (44.4%) patients achieved a partial response, and the remaining five (55.6%) patients had stable disease. The response lasted for at least 12 months in 8 (88.9%) patients with EBVaBTC, including 4 patients received PD-1 antibody monotherapy. Blood EBV-DNA was detectable in all 9 patients (100%) with EBVaBTC. Multiplex immunofluorescence staining revealed more abundant CD3+ and CD8+ T cells infiltration in EBVaBTC than that of mismatch repair deficient BTC. A total of 8,379 proteins were identified by proteomics. Comparison of EBVaBTC and paired normal tissue revealed 547 differentially expressed proteins. Enrichment analysis demonstrated that differentially expressed proteins in the EBVaBTC were associated with EBV infection and T cell activation. We also observed the expression of immune checkpoint other than PD-1, including LAG-3 and TIM-3. Conclusions: Identification of EBVaBTC may represent a subset of patients with promising response to immunotherapy. [Table: see text]

Effect of germline mutations on somatic alteration landscapes in BRCA-associated cancers.

10509 Background: The interaction between germline alterations in homologous recombination repair (gHRR) genes and tumoral genomic landscapes is poorly understood. We hypothesized that gHRR alterations may influence somatic mutational landscapes in BRCA-associated cancers. Methods: Using a large de-identified real-world dataset of patients undergoing matched tumor/normal next-generation DNA sequencing (Tempus xT), we compared somatic genomic landscapes of tumors arising in individuals with germline HRR mutations (g BRCA1, g BRCA2, g PALB2, g ATM, g CHEK2) versus their sporadic counterparts, across four BRCA-associated cancers (breast, ovary, pancreas, prostate). Results: In breast cancer (N = 6955; gHRR-altered 5.9%, sporadic 94.1%), somatic TP53 muts were enriched in g BRCA1 pts, and depleted in g ATM and g CHEK2 pts (all P &lt; 0.001). ESR1 muts were depleted in g BRCA1 pts, and enriched in g ATM and g CHEK2 pts (all P &lt; 0.05). PIK3CA muts were depleted in g BRCA1/ 2 pts, and enriched in g ATM and g CHEK2 pts (all P &lt; 0.05). FGFR1 muts were depleted in g BRCA1/2 and g PALB2 pts, and enriched in g ATM pts (all P &lt; 0.01). HER2 and CDH1 alterations were depleted in g BRCA1 pts (P &lt; 0.05). In ovarian cancer (N = 4244; gHRR-altered 6.2%, sporadic 93.8%), somatic TP53 muts were enriched in g BRCA1/ 2 pts, and depleted in g ATM and g CHEK2 pts (all P &lt; 0.05). ESR1 muts were enriched in g ATM pts (P &lt; 0.001). PIK3CA muts were depleted in g BRCA1/ 2 pts, and enriched in g ATM pts (all P &lt; 0.05). KRAS muts were depleted in g BRCA1/ 2 pts (P &lt; 0.05). In pancreatic cancer (N = 5386; gHRR-altered 3.9%, sporadic 96.1%), somatic TP53 muts were enriched in g BRCA1 pts, and depleted in g ATM and g PALB2 pts (all P &lt; 0.05). KRAS muts were enriched in g BRCA1/2 and g ATM pts (P &lt; 0.05). Unexpectedly, ESR1 muts were enriched in g PALB2 and g CHEK2 pts (P &lt; 0.05). In prostate cancer (N = 4378; gHRR-altered 4.5%, sporadic 95.5%), somatic TP53 muts trended higher in g BRCA1 pts, and lower in g ATM and g PALB2 pts (P = NS). TMPRSS2-ERG fusions were depleted in g BRCA1 and g BRCA2 pts (P &lt; 0.05). FOXA1 muts were enriched in g BRCA2 pts (P &lt; 0.05), while BRAF and CDK12 muts were enriched in g CHEK2 pts (P &lt; 0.05 and P = 0.07). Median TMB was higher in BRCA-associated cancers with g BRCA1/2 and gPALB2 muts relative to sporadic cancers (all P &lt; 0.05), but not in pts with g ATM or g CHEK2 muts. There were no differences in MMR mutations or MSI status. Conclusions: Across four BRCA-associated cancers, TP53 muts are enriched in BRCA1 pts and depleted in g ATM pts. In breast/ovarian cancers, PIK3CA muts are depleted in g BRCA1/ 2 pts, while ESR1 muts are enriched in g ATM pts. KRAS muts are enriched in g BRCA1/2-altered pancreas cancers, but are depleted in g BRCA1/2-altered ovarian cancers. g BRCA2-altered prostate cancers are enriched in FOXA1 muts, and depleted in ERG fusions. These data suggest that gHRR-mutated cancers have distinct genomic landscapes compared to their sporadic counterparts; this may influence therapeutic considerations.

Characterization and impact of Wnt5A signaling on outcomes of urothelial carcinoma.

560 Background: Active Wnt signaling via WNT5A through ROR1 and its partner, ROR2, or WNT5A/frizzled 2 (FZD2) is linked to processes driving tumorigenesis, disease progression and therapy resistance. The role of this pathway in the pathogenesis of urothelial carcinoma (UC) is not fully elucidated. In adult tissue, ROR1 is largely absent, which makes it ideal for targeted therapies, with several ROR1 targeting agents in early clinical development. We utilized a large dataset of molecularly characterized UC tumors to investigate the significance of Wnt5a/ ROR1, ROR2 or FZD2 transcriptional expression. Methods: NextGen Sequencing of DNA (592 genes or WES)/RNA (WTS) was performed for 4743 UC tumors submitted to Caris Life Sciences (Phoenix, AZ). PD-L1 expression (SP142; Positive (+): ³2+, ³%5) was tested by IHC. Gene expression profiles were analyzed for a transcriptional signature predictive of response to immunotherapy (T cell-inflamed; Bao, 2020). WNT5a, ROR1, ROR2, and FZD2-high and -low expression were defined as ³ top and &lt; bottom quartile of transcripts per million (TPM), respectively. Mann-Whitney U and X2/Fisher-Exact tests were applied where appropriate, with P-values adjusted for multiple comparisons ( q &lt; .05). Real-world overall survival (OS) information was obtained from insurance claims data and Kaplan Meier estimates were calculated for molecularly defined cohorts. Results: We observed similar expression of these WNT5A signaling pathway genes between upper (N=795) and lower tract UC (N=3,204): WNT5A (22.7 v. 22.2 median TMP (mTPM), q = .18), FZD2 (3.4 v 3.5, q = .93), ROR1 (2.0 v. 1.7, q = .05), and ROR2 (2.1 v 2.5, q &lt; .01). WNT family gene expression varied significantly between primary (N=2,756) and metastatic sites (N=1,361): WTN5A (25.2 v 16.8 mTPM), FZD2 (3.2 v 4.05), ROR1 (1.7 v 2.1), and ROR2 (2.4 v 2.6) for primary vs. metastatic sites respectively ( q &lt; .05 for all). Comparison of high- and low-expression subgroups revealed variation in the prevalence of TP53, FGFR3 and RB1 mutations, as well as PDL1+ staining and T cell-inflamed scores (Table). High gene expression for ROR2 (HR 0.66, 95% CI 0.56-0.78, p &lt; .001) and FZD2 (HR 0.75, 95% CI 0.63-0.89, p &lt; .001) was associated with worse OS compared to low gene expression. No significant difference in OS was observed for WNT5A (HR 0.97, 95% CI 0.82-1.15, p &lt; .76) and ROR1 (HR 0.86, 95% CI 0.72-1.01, p &lt; .068). Conclusions: Distinct genomic and immune landscapes for the four investigated WNT pathway components were observed and should be leveraged to identify combination therapies that complement the current pipeline of WNT pathway-targeting drugs. [Table: see text]

High-Grade Sarcomas with Myogenic Differentiation Harboring Hotspot PDGFRB Mutations

PDGFRB-activating mutations have been reported in pediatric myofibroma and myofibromatosis. However, recurrent gain-of-function PDGFRB mutations have not been documented in sarcomas with myogenic differentiation. Driven by occasional sarcomas harboring PDGFRB mutations, we investigated their prevalence and clinicopathologic and genomic features in a large cohort of sarcomas. An institutional targeted DNA next-generation sequencing database was searched for sarcomas with myogenic differentiation harboring hotspot PDGFRB gene alterations. Among 3300 patients with sarcomas, 21 (0.6%) patients were identified (17 women, 4 men) with an age range of 35 to 88 years. The site distribution included 13 gynecologic tract (12 uteri, 1 vagina), 4 bone and soft tissue, and 4 viscera. All except 1 were high grade. Most patients were diagnosed as sarcomas with myogenic differentiation based on partial staining for 1 or more muscle markers, whereas 6 were labeled as leiomyosarcoma (LMS). Most tumors showed monomorphic spindle morphology, with either heterogeneous features of myofibroblastic and smooth muscle differentiation or an undifferentiated phenotype. Hormone receptors were negative in all uterine cases. PDGFRB immunostaining in all cases tested was strong and diffuse, whereas PDGFRA was negative/focal. The most frequent PDGFRB mutations were exon 12 (43%), exon 14 (N666K/S/T) (38%), and exon 18 (D850Y/H/V or insertion/deletion) (19%). The most frequent co-existing genetic alterations (26% to 37%) occurred in CDKN2A/B, TP53, TERT, and MED12. Moreover, PDGFRB-mutant sarcomas had an overall distinct genomic landscape compared with both uterine and soft tissue LMS control groups. These tumors were associated with a highly aggressive clinical course, with frequent distant metastases (81%) and death (76%), regardless of anatomic location, and worse overall survival compared with the 2 LMS control groups. This is the first study documenting recurrent hotspot PDGFRB alterations in high-grade sarcomas, which show a predilection for uterine location and myogenic differentiation that fall short of the diagnostic criteria for LMS. Further studies are needed to investigate the therapeutic potential of kinase inhibitors in this group of tumors.

TRNA-Derived RNA Fragments Are Novel Biomarkers for Diagnosis, Prognosis, and Tumor Subtypes in Prostate Cancer.

tRNA-derived RNA fragments (tRFs) are a novel class of small ncRNA that are derived from precursor or mature tRNAs. Recently, the general relevance of their roles and clinical values in tumorigenesis, metastasis, and recurrence have been increasingly highlighted. However, there has been no specific systematic study to elucidate any potential clinical significance for these tRFs in prostate adenocarcinoma (PRAD), one of the most common and malignant cancers that threatens male health worldwide. Here, we investigate the clinical value of 5'-tRFs in PRAD. Small RNA sequencing data were analyzed to discover new 5'-tRFs biomarkers for PRAD. Machine learning algorithms were used to identify 5'-tRF classifiers to distinguish PRAD tumors from normal tissues. LASSO and Cox regression analyses were used to construct 5'-tRF prognostic predictive models. NMF and consensus clustering analyses were performed on 5'-tRF profiles to identify molecular subtypes of PRAD. The overall levels of 5'-tRFs were significantly upregulated in the PRAD tumor samples compared to their adjacent normal samples. tRF classifiers composed of 13 5'-tRFs achieved AUC values as high as 0.963, showing high sensitivity and specificity in distinguishing PRAD tumors from normal samples. Multiple 5'-tRFs were identified as being associated with the PRAD prognosis. The tRF score, defined by a set of eight 5'-tRFs, was highly predictive of survival in PRAD patients. The combination of tRF and Gleason scores showed a significantly better performance than the Gleason score alone, suggesting that 5'-tRFs can offer PRAD patients additional and improved prognostic information. Four molecular subtypes of the PRAD tumor were identified based on their 5'-tRF expression profiles. Genetically, these 5'-tRFs PRAD tumor subtypes exhibited distinct genomic landscapes in tumor cells. Clinically, they showed marked differences in survival and clinicopathological features. 5'-tRFs are potential clinical biomarkers for the diagnosis, prognosis, and classification of tumor subtypes on a molecular level. These can help clinicians formulate personalized treatment plans for PRAD patients and may have similar potential applications for other disease types.

Genomic characterization reveals distinct mutation landscapes and therapeutic implications in neuroendocrine carcinomas of the gastrointestinal tract.

Neuroendocrine carcinomas of the gastrointestinal tract (GI-NECs) remain a disease of grim prognosis with limited therapeutic options. Their molecular characteristics are still undefined. This study aimed to explore the underlying genetic basis and heterogeneity of GI-NECs. Comprehensive genomic analysis using whole-exome sequencing was performed on 143 formalin-fixed, paraffin-embedded samples of surgically resected GI-NEC with a thorough histological evaluation. Mutational signatures, somatic mutations, and copy number aberrations were analyzed and compared across anatomic locations and histological subtypes. Survival analysis was conducted to identify the independent factors. In total, 143 GI-NECs were examined: the stomach, 87 cases (60.8%); the esophagus, 29 cases (20.3%); the colorectum, 20 cases (14.0%); and the small intestine, 7 cases (4.9%). Eighty-three (58.0%) and 60 (42.0%) cases were subclassified into small cell and large cell subtypes, respectively. GI-NECs showed distinct genetic alterations from their lung counterparts and non-neuroendocrine carcinomas in the same locations. Obvious heterogeneity of mutational signatures, somatic mutations, and copy number variations was revealed across anatomic locations rather than histological subtypes. Except for tumor protein p53 (TP53) and retinoblastoma 1 (RB1), the most frequently mutated genes in the stomach, esophagus, colorectum, and small intestine were low-density lipoprotein receptor-related protein 1B (LRP1B), notch receptor 1 (NOTCH1), adenomatosis polyposis coli (APC), catenin beta 1 (CTNNB1), respectively. Mutations in the WNT-β-catenin, NOTCH and erythroblastic leukemia viral oncogene B (ERBB) pathways were prevalently identified in gastric, esophageal, and colorectal NECs, respectively. Importantly, 104 (72.7%) GI-NECs harbored putative clinically relevant alterations, and non-gastric location and RB1 bi-allelic inactivation with copy number alterations were identified as two independent poor prognostic factors. Furthermore, we found that tumor cells in GI-NECs first gain clonal mutations in TP53, RB1, NOTCH1 and APC, followed by subsequent whole-genome doubling (WGD) and post-WGD clonal mutations in LRP1B, CUB and Sushi multiple domains 3 (CSMD3), FAT tumor suppressor homolog 4 (FAT4) and erb-b2 receptor tyrosine kinase 4 (ERBB4), and finally develop subclonal mutations. GI-NECs harbor distinct genomic landscapes and demonstrate significant genetic heterogeneity across different anatomic locations. Moreover, potentially actionable alterations and prognostic factors were revealed for GI-NECs.

Genomic landscape of multiple Bowen's disease using whole-exome sequencing.

The genomic landscape of Bowen's disease (BD), with multiple manifestations, has not yet been determined. This study aimed to investigate the genomic alterations in multiple BD. We performed whole-exome sequencing of BD lesions (n=9) and matched germlines collected from three patients with multiple (≥3) BD to detect somatic and germline mutations. We found a median of 64 somatic mutations in each sample (range 20-267). UV-signature mutations accounted for 64.9% (median, range 26.0%-82.1%) of point mutations. Putative driver mutations were found in five BDs (RB1 p.R445*, ARID2 p.R274*, TP53 p.Y163D/p.Y205D/p.R342*, KMT2C p.R4549C) but not in the other four lesions. Somatic mutations were not shared between multiple BD lesions collected from the same patient, indicating a different clonal origin. We also found no known pathogenic germline mutations in cancer-related genes. The mutational signature analysis revealed that UV signatures (SBS7a/7b) and age-related signatures (SBS1/5) were the main active signatures. Copy number alterations (CNAs) were found in two BDs: one with extensive CNA regions (21.7% of the genome), including driver genes (PIK3CA/SOX2/TP63 and MYC gain, and CDKN2A loss), and the other with 1q gain. Our study revealed that multiple BD lesions harbor distinct genomic landscapes, suggesting that they have different risks of malignant progression.

Genomic landscape of advanced prostate cancer patients with BRCA1 versus BRCA2 mutations as detected by comprehensive genomic profiling of cell-free DNA.

BRCA1-mutated prostate cancer has been shown to be less responsive to poly (ADP-ribose) polymerase (PARP) inhibitors as compared to BRCA2-mutated prostate cancer. The reason for this differential response is not clear. We hypothesized this differential sensitivity to PARP inhibitors may be explained by distinct genomic landscapes of BRCA1 versus BRCA2 co-segregating genes. In a large dataset of 7,707 men with advanced prostate cancer undergoing comprehensive genomic profiling (CGP) of cell-free DNA (cfDNA), 614 men harbored BRCA1 and/or BRCA2 alterations. Differences in the genomic landscape of co-segregating genes was investigated by Fisher’s exact test and probabilistic graphical models (PGMs). Results demonstrated that BRCA1 was significantly associated with six other genes, while BRCA2 was not significantly associated with any gene. These findings suggest BRCA2 may be the main driver mutation, while BRCA1 mutations tend to co-segregate with mutations in other molecular pathways contributing to prostate cancer progression. These hypothesis-generating data may explain the differential response to PARP inhibition and guide towards the development of combinatorial drug regimens in those with BRCA1 mutation.

1383P Genomic landscape of advanced prostate cancer patients with BRCA1 versus BRCA2 mutations as detected by comprehensive genomic profiling (CGP) of cell-free DNA (cfDNA)

BRCA1-mutated prostate cancer has been shown to be less responsive to poly (ADP-ribose) polymerase (PARP) inhibitors as compared to BRCA2-mutated prostate cancer. The reason for this differential response is not clear. We hypothesized this differential sensitivity to PARP inhibitors may be explained by distinct genomic landscapes of BRCA1 versus BRCA2 co-segregating genes.

Molecular map of chronic lymphocytic leukemia and its impact on outcome.

Recent advances in cancer characterization have consistently revealed marked heterogeneity, impeding the completion of integrated molecular and clinical maps for each malignancy. Here, we focus on chronic lymphocytic leukemia (CLL), a B cell neoplasm with variable natural history that is conventionally categorized into two subtypes distinguished by extent of somatic mutations in the heavy-chain variable region of immunoglobulin genes (IGHV). To build the 'CLL map,' we integrated genomic, transcriptomic and epigenomic data from 1,148 patients. We identified 202 candidate genetic drivers of CLL (109 new) and refined the characterization of IGHV subtypes, which revealed distinct genomic landscapes and leukemogenic trajectories. Discovery of new gene expression subtypes further subcategorized this neoplasm and proved to be independent prognostic factors. Clinical outcomes were associated with a combination of genetic, epigenetic and gene expression features, further advancing our prognostic paradigm. Overall, this work reveals fresh insights into CLL oncogenesis and prognostication.

Exome sequencing reveals a distinct somatic genomic landscape in breast cancer from women with germline PTEN variants

Germline PTEN variants (PTEN hamartoma tumor syndrome [PHTS]) confer up to 85% lifetime risk of female breast cancer (BC). BCs arising in PHTS are clinically distinct from sporadic BCs, including younger age of onset, multifocality, and an increased risk of second primary BCs. Yet, there is no previous investigation into the underlying genomic landscape of this entity. We sought to address the hypothesis that BCs arising in PHTS have a distinct genomic landscape compared to sporadic counterparts. We performed and analyzed exome sequencing data from 44 women with germline PTEN variants who developed BCs. The control cohort comprised of 497 women with sporadic BCs from The Cancer Genome Atlas (TCGA) dataset. We demonstrate that PHTS-derived BCs have a distinct somatic mutational landscape compared to the sporadic counterparts, namely second somatic hits in PTEN, distinct mutational signatures, and increased genomic instability. The PHTS group had a significantly higher frequency of somatic PTEN variants compared to TCGA (22.7% versus 5.6%; odds ratio [OR] 4.93; 95% confidence interval [CI] 2.21 to 10.98; p<0.001) and a lower mutational frequency in PIK3CA (22.7% versus 33.4%; OR 0.59; 95% CI 0.28 to 1.22; p= 0.15). Somatic variants in PTEN and PIK3CA were mutually exclusive in PHTS (p= 0.01) but not in TCGA. Our findings have important implications for the personalized management of PTEN-related BCs, especially in the context of more accessible genetic testing.

Abstract 5995: Tumor genomic landscape in patients (pts) with and without liver metastasis in metastatic urothelial carcinoma (mUC)

Abstract Background: Presence of liver metastasis is associated with a poor prognosis in mUC. According to the prognostic model by Sonpavde et. al, post platinum treated mUC pts with liver metastasis had a 50% higher risk of death on immunotherapy compared to pts without liver metastasis (PMID 32552295). Herein, we hypothesize that pts with liver metastasis harbor a distinct tumor genomic landscape compared to pts without liver metastasis. Methods: In this IRB-approved retrospective study, mUC pts with and without liver metastasis at baseline and available tumor comprehensive genomic profiling (CGP) from a CLIA-certified laboratory were included. Variants of unknown significance and mutated genes present in ≤ 5% of pts were excluded. A Chi-square test was used to compare the frequency of mutated genes between both groups. The p-value was adjusted for false discovery, significance was preset at 0.25. Results: 114 pts were included. 16/114 pts had a liver metastasis. Median age was 69 and 67 years, male/female 12/4 and 72/26, smokers/non-smokers 6/10 and 46/52, for pts with and without liver metastasis respectively. The most frequent genomic alterations (GAs) were seen in TP53, CDKN2B and FGFR3. Pts with liver metastasis had significantly higher GAs in MYC (unadjusted p=0.009, adjusted p=0.24) compared to pts without liver metastasis. The median number of GAs per patient and the tumor mutation burden were not significantly different between groups. Conclusion: Our results indicate GAs in MYC are significantly enriched in pts with liver metastasis compared to those without liver metastasis. These hypothesis generating data require external validation. Genes Liver Metastasis: N=16 Non-Liver Metastasis: N=98 Adjusted P-value MYC 3(18.8%) 3(3.1%) 0.24 ERBB3 3(18.8%) 5(5.1%) 0.62 FGFR3 5(31.2%) 16(16.3%) 0.73 RICTOR 2(12.5%) 4(4.1%) 0.73 TP53 7(43.8%) 58(59.2%) 0.73 ERBB2 3(18.8%) 9(9.2%) 0.73 CDKN2B 7(43.8%) 31(31.6%) 0.73 ARID1A 3(18.8%) 21(21.4%) 0.90 RB1 3(18.8%) 24(24.5%) 0.90 KDM6A 2(12.5%) 19(19.4%) 0.90 Citation Format: Nishita Tripathi, Yeonjung Jo, Nicolas Sayegh, Beverly Chigarira, Blake Nordblad, Vinay Mathew Thomas, Benjamin Haaland, Roberto Nussenzveig, Sumati Gupta, Neeraj Agarwal, Umang Swami, Benjamin L. Maughan. Tumor genomic landscape in patients (pts) with and without liver metastasis in metastatic urothelial carcinoma (mUC) [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 5995.