Presumed Pathogenic Variants Research Articles

Prognostic Value of Tumor Mutational Burden in Follicular Lymphoma Patients Treated with Immunochemotherapy

INTRODUCTION: Follicular lymphoma (FL) is the most prevalent indolent non-Hodgkin lymphoma subtype. It is characterized by frequent relapses, and patients who progress or relapse within 24 months (POD24) of front-line immunochemotherapy (ICT) have a poor outcome. Several prognostic indices have been proposed to identify high-risk patients, but none of them are currently used in clinical practice to guide treatment. Tumor mutational burden (TMB), defined as the number of somatic mutations/megabase (mut/Mb), has been proposed as a biomarker for predicting response to immune checkpoint inhibitors in several types of cancer. Therefore, this study aimed to explore the role of TMB in FL patients treated with frontline ICT. METHODS: The study included a total of 103 patients with FL grade 1-3a treated with frontline ICT. TMB was determined by Next Generation Sequencing (NGS) in 98 diagnostic lymph node biopsies with enough material available, with the Oncomine Tumor Mutational Load Assay (Thermo Fisher). This assay covers 1.65Mb across 409 genes frequently mutated in cancer, allowing an accurate estimation of TMB. Sequencing was performed on the Ion GeneStudio S5 system with >300X mean read depth. Only exonic non-synonymous variants remaining after manual variant refinement to exclude artefacts were considered. In addition, the mutational profile of 56 of the 98 patients was analyzed using a QIAgen custom DNA panel which covered 64 genes frequently mutated in FL. Sequencing was performed using NextSeq (Illumina) with >3000X mean read depth. Variants were analyzed and interpreted for pathogenicity using several databases, and only pathogenic or presumed pathogenic variants (P/PP) were considered. Statistical analysis was performed using SPSS and R. RESULTS: The final cohort included 94 patients, as in 4/98 patients (4%) TMB could not be accurately estimated due to a high frequency of artefacts caused by cytosine deamination. Median age at diagnosis of 61 years (range 24-86), 51% males and 49% females. Patients with advanced disease stage at diagnosis represented 87% of the cohort. Median TMB was 5.08mut/Mb (range 1.69-13.80 mut/Mb), and all cases genetically profiled presented at least one P/PP variant. The number of P/PP mutations correlated with TMB at diagnosis (p=0.004). TMB did not correlate with clinical parameters, such as age at diagnosis (p=0.891), histologic grade (p=0.243), Ann Arbor stage (p=0.200), ECOG (p=0.190), FLIPI (p=0.363), FLIPI2 (p=0.693), or LDH levels (p=0.483). However, lower TMB at diagnosis was associated with the absence of the t(14;18)(q32;q21) translocation (mean 3.93 vs 5.42 mut/Mb, p=0.006). The analysis of the mutational status of the 64 genes frequently mutated in FL revealed that mutations in genes involved in cellular migration ( GNA13, GNAI2) were associated with higher TMB values (4.50 vs 7.56 mut/Mb, p=0.001). An statistically significant association between low TMB and shorter progression-free survival (PFS) (p=0.011) was found when stratifying patients into two groups: high basal TMB (>2.55mut/Mb) and low basal TMB (≤2.55mut/Mb) (Figure 1). 2-year PFS was 54.5% (95%CI: 33.2-89.5%) in low-TMB patients and 84.9% (95%CI: 77.3-93.7%) in high-TMB patients. 5-year PFS was 39.0% (95%CI: 19.7-77.0%) in low-TMB patients and 70.5% (95%CI: 60.0-82.9%) in high-TMB patients. The proportion of POD24 cases was higher in the low-TMB group (46.1% vs 16.9%, p=0.039), none of them being refractory to treatment (7/70 in the high-TMB group). Genetically, the low-TMB group was enriched in patients with mutations affecting genes involved in the mTOR signaling pathway ( ATP6AP1, ATP6AP2, ATP6V1B2, RRAGC) (50.0% vs 8.7%, p=0.006). CONCLUSIONS: FL patients harboring t(14;18) or mutations in genes involved in migration have higher TMB values at diagnosis. Patient stratification based on TMB values allows the identification of a subgroup of FL patients with ≤2.55 mut/Mb with shorter PFS after treatment with frontline ICT. The prognostic usefulness of TMB should also be explored at relapse, in particular in patients receiving novel immunotherapies such as bispecific antibodies and CAR T-cell therapies, that are being approved for relapsed/refractory FL. ACKNOWLEDGMENTS: Work supported by Institut de Salud Carlos III-Federación Española de Enfermedades Raras (ISCIII-FEDER) grant PI19/00034, GLD18/00117, 2017SGR205, PT20/00023, and Xarxa de Banc de Tumors de Catalunya.

Rare Saposin A deficiency: Novel variant and psychosine analysis

Saposin A is a post-translation product of the prosaposin (PSAP) gene that serves as an activator protein of the galactocerebrosidase (GALC) enzyme, and is necessary for the degradation of certain glycosphingolipids. Deficiency of saposin A leads to a clinical picture identical to that of early-infantile Krabbe disease caused by GALC enzyme deficiency. Galactosylsphingosine, also known as psychosine, is a substrate of the GALC enzyme that is known to be elevated in classic Krabbe disease. We present the case of an 18-month-old male with clinical and radiological findings concerning for Krabbe disease who had preserved GALC enzyme activity and negative GALC gene sequencing, but was found to have a homozygous variant, c.257 T > A (p.I86N), in the saposin A peptide of PSAP. Psychosine determination on dried blood spot at 18 months of age was elevated to 12 nmol/L (normal <3 nmol/L). We present this case to add to the literature on the rare diagnosis of atypical Krabbe disease due to saposin A deficiency, to report a novel presumed pathogenic variant within PSAP, and to suggest that individuals with saposin A deficiency may have elevated levels of psychosine, similar to children with classic Krabbe disease due to GALC deficiency.

Detection of Pathogenic Germline Variants Among Patients With Advanced Colorectal Cancer Undergoing Tumor Genomic Profiling for Precision Medicine.

Genomic profiling of colorectal cancer aims to identify actionable somatic mutations but can also discover incidental germline findings. The purpose of this study was to report the detection of pathogenic germline variants that confer heritable cancer predisposition. This was a retrospective study. The study was conducted at a tertiary-referral institution. Between 2012 and 2015, 1000 patients with advanced cancer underwent targeted exome sequencing of a 202-gene panel. The subgroup of 151 patients with advanced colorectal cancer who underwent matched tumor-normal (blood) sequencing formed our study cohort. Germline variants in 46 genes associated with hereditary cancer predisposition were classified according to a defined algorithm based on in silico predictions of pathogenicity. Patients with presumed pathogenic variants were examined for type of mutation, as well as clinical, pedigree, and clinical genetic testing data. We measured detection of pathogenic germline variants. A total of 1910 distinct germline variants were observed in 151 patients. After filtering, 15 pathogenic germline variants (9.9%) were found in 15 patients, arising from 9 genes of varying penetrance for colorectal cancer (APC (n = 2; 13%), ATM (n = 1; 6%), BRCA1 (n = 2; 13%), CDH1 (n = 2; 13%), CHEK2 (n = 4; 27%), MSH2 (n = 1; 7%), MSH6 (n = 1; 7%), NF2 (n = 1; 7%), and TP53 (n = 1; 7%)). Patients with pathogenic variants were diagnosed at a younger age than those without (median, 45 vs 52 y; p = 0.03). Of the 15 patients, 7 patients (46.7%) with variants in low/moderate- penetrant genes for colorectal cancer would likely have not been tested based on clinical and pedigree criteria, where 2 harbored clinically actionable variants (CDH1 and NF2, 28.5% of 7). This study was limited by its small sample size and advanced-stage patients. Tumor-normal sequencing can incidentally discover clinically unsuspected germline variants that confer cancer predisposition in 9.9% of patients with advanced colorectal cancer. Precision medicine should integrate clinical cancer genetics to inform and interpret the actionability of germline variants and to provide follow-up care to mutation carriers. See Video Abstract at http://links.lww.com/DCR/A906.

Characteristics of colorectal cancer (CRC) patients with BRCA1 and BRCA2 mutations.

606 Background: Between 3-5% of CRC patients have BRCA1/2 pathogenic mutations. This study aims to identify associations between BRCA1 and BRCA2 mutations and clinical characteristics in CRC. Methods: A total of 6396 CRC tumor samples were tested with Next-Generation Sequencing (NGS) on a 592-gene panel, pathogenic or presumed pathogenic variants were counted as mutations (mt). Microsatellite instability (MSI) and tumor mutational burden (TMB) were tested by NGS. Statistical correlations were investigated using ANOVA, Chi-square and t-test. Results: Among tumors sampled, 53% derived from male patients and median age was 60 years. BRCA1 mt were detected in 1.1% (n = 72) of tumors, while BRCA2 in 2.8% (n = 179). BRCA1 mt were more frequent in women (W;65%) than men (M;35%) (p = 0.0019) while no relationship with sex was seen for BRCA2 mt (42% F vs. 58% M). No significant associations with age were noticed. Majority of pathogenic mt in BRCA1 (52%; n = 34) and BRCA2 (62%; n = 103) occurred in MSI-High (MSI-H) cases. MSI-H pts had more frameshift mt in both BRCA1/2 than MSS pts. MSS cases had lower rates of BRCA1 and 2 pathogenic mt (44% and 37%, respectively). Right-sided tumors were significantly associated with BRCA1 (p = 0.0056) and BRCA2 (p &lt; 0.0001) mt in MSI-H cases only. BRCA1/2 mt were associated with higher TMB in all CRCs, including MSI-H and MSS cases (p &lt; 0.001). POLE mt (n = 31) were associated with higher BRCA1/2 mt rates (9.6%, 55% respectively). Among MSS cases with POLE wild-type status, BRCA1 (p = 0.0269) and BRCA2 (p = 0.0151) mt were associated with high TMB and combining both BRCA1/2 mutations led to an even higher TMB (3.6%; p = 0.001). Conclusions: This is the first study to show that BRCA1/2 mutations are more frequent in MSI-H, and independently associated with higher TMB, pathogenic POLE mutations, and right-sided tumors in MSI-H CRCs. Given their relationship with TMB, the presence of BRCA1/2 mutations may be potential predictive biomarkers for checkpoint or PARP inhibitors in CRC, a finding that should be prospectively evaluated.

Abstract A190: Management of germline findings revealed throughout the course of tumor-normal whole genome sequencing in oncology

Abstract Background: The Personalized OncoGenomics (POG) project at the BC Cancer Agency utilizes tumor-normal whole genome sequencing (WGS) to understand key driver pathways and guide personalized treatment decisions. Analysis of the germline data can reveal variants; these may be presumed pathogenic, presumed benign, or of unknown significance (VUS). We have developed a process for evaluating and returning presumed pathogenic variants in known cancer susceptibility genes to patients, for counseling and validation in a clinical-accredited laboratory. Methods: Patients receive germline cancer-related information as part of the consent process for participation in the POG program. A subcommittee comprising medical geneticists, bioinformaticians, pathologists, oncologists, and an ethicist review the germline results. Any variants suspicious of being an artifact undergo a technical validation step. Presumed pathogenic findings of known cancer susceptibility genes are returned to the patient by their treating oncologist and patients are referred to the Hereditary Cancer Program (HCP), for genetic counseling and clinical confirmation. Results: From June 2012-January 2017, 466 patients have consented to the project. To date, 39 cases (8.4%) had at least one variant that was deemed pathogenic, and 86 cases had at least one VUS in a known cancer susceptibility gene. 11 out of 23 cases (47.8%) with high-penetrance mutations were already known to HCP. All VUS were reviewed by the subcommittee, taking into consideration the VUS and clinical context. 8 of the subjects with pathogenic results and 3 with VUS were known to HCP before POG data were generated. A VUS in 7 cases (1.5%) was returned after review. Conclusions: The number of pathogenic variants in known cancer susceptibility genes is consistent with published oncology results. We created a process to manage clinically relevant germline findings discovered during the course of genomic research to ensure appropriate care for patients. Genetic counseling within HCP and validation of variants in the clinically accredited Cancer Genetics Laboratory enables seamless return of research-generated clinically relevant germline results to affected subjects. Citation Format: Howard J. Lim, Kasmintan A. Schrader, Sean Young, Jessica M T Nelson, Alexandra Fok, Erin Pleasance, Martin Jones, Yaoqing Shen, Linlea Armstrong, Alice Virani, Shahrad Rassekh, Rebecca Deyell, Stephen Yip, Robyn Roscoe, Aly Karsan, Marco Marra, Janessa J. Laskin. Management of germline findings revealed throughout the course of tumor-normal whole genome sequencing in oncology [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr A190.

Neurodevelopmental Disorders Caused by De Novo Variants in KCNB1 Genotypes and Phenotypes

Knowing the range of symptoms seen in patients with a missense or loss-of-function variant in KCNB1 and how these symptoms correlate with the type of variant will help clinicians with diagnosis and prognosis when treating new patients. To investigate the clinical spectrum associated with KCNB1 variants and the genotype-phenotype correlations. This study summarized the clinical and genetic information of patients with a presumed pathogenic variant in KCNB1. Patients were identified in research projects or during clinical testing. Information on patients from previously published articles was collected and authors contacted if feasible. All patients were seen at a clinic at one of the participating institutes because of presumed genetic disorder. They were tested in a clinical setting or included in a research project. The genetic variant and its inheritance and information on the patient's symptoms and characteristics in a predefined format. All variants were identified with massive parallel sequencing and confirmed with Sanger sequencing in the patient. Absence of the variant in the parents could be confirmed with Sanger sequencing in all families except one. Of 26 patients (10 female, 15 male, 1 unknown; mean age at inclusion, 9.8 years; age range, 2-32 years) with developmental delay, 20 (77%) carried a missense variant in the ion channel domain of KCNB1, with a concentration of variants in region S5 to S6. Three variants that led to premature stops were located in the C-terminal and 3 in the ion channel domain. Twenty-one of 25 patients (84%) had seizures, with 9 patients (36%) starting with epileptic spasms between 3 and 18 months of age. All patients had developmental delay, with 17 (65%) experiencing severe developmental delay; 14 (82%) with severe delay had behavioral problems. The developmental delay was milder in 4 of 6 patients with stop variants and in a patient with a variant in the S2 transmembrane element rather than the S4 to S6 region. De novo KCNB1 missense variants in the ion channel domain and loss-of-function variants in this domain and the C-terminal likely cause neurodevelopmental disorders with or without seizures. Patients with presumed pathogenic variants in KCNB1 have a variable phenotype. However, the type and position of the variants in the protein are (imperfectly) correlated with the severity of the disorder.

Abstract 2779: Assessing germline and somatic alterations in DNA repair pathway in cancer

Abstract Introduction: DNA repair genes are involved with repair of single strand breaks as well as double strand breaks which ultimately maintain the genome integrity. The genes involved with DNA repair are frequently deregulated in cancer and these defects can be exploited therapeutically. Our study aims to explore somatic and germline changes in DNA repair genes across multiple cancer types. Methods: Genomic profiling was performed on 226 tumor samples from 24 cancer types with ages ranging 19-82 years. We performed targeted exome sequencing of 562 genes in tumor and paired normal DNA which included 112 genes associated with cancer predisposition syndromes. In addition to identifying genomic changes in 33 key DNA repair genes (ATM, ATR, ARID1A, BAP1, BARD1, BRCA1, BRCA2, BRIP1, CDK12, CHEK1, CHEK2, ERCC2, ERCC3, ERCC4, FANCA gene family, MMR genes, NBS1, PALB2, PTEN, RAD50, RAD51, TP53 and POLE), we also characterized germline mutations using public databases (ClinVar, Ensembl). Results: Profiled specimens included breast cancer (17.5%), ovarian cancer (15%), CRC (11%), NSCLC (10%), uterine (6%), sarcoma (6%) and other cancers. The most commonly deleted or mutated (somatic) DNA repair genes included TP53 (55%), PTEN (16%), ARID1A (13%), FANCA (11%), ERCC2 (10%), and ATM, BAP1, CDK12, CHEK2 at 6% each and BRCA1/2 combined at (9%). Deleterious mutations in MMR genes were noted in 3.5% (8/226) which included 1 case of germline PMS2-S46I mutation. Interestingly, a 55 yr old male African American CRC patient harbored germline XPC-P334H mutation along with somatic MLH1-Y548fs. No mutations were found in FANCC, FANCF, FANCG and RAD51. Germline analysis revealed a total of 47 pathogenic and presumed pathogenic variants out of which 61.7%(29/47) were DNA repair genes. A total of 13% (30/226) of cases harbored germline variants in DNA repair genes. Germline events included 9 cases with BRCA1 and BRCA2 clinically significant mutations. Presence of loss of heterozygosity (LOH) at the BRCA locus in 5/9 cases and somatic mutations in 3/9 cases were noted in germline BRCA mutated samples. Germline and somatic BRCA1 and BRCA2 alterations were present in 10% (22/226) of total cohort and as expected, the majority were present in breast and epithelial ovarian cancer. Genetic deficiencies in DNA repair pathway genes are being exploited therapeutically with PARP inhibitors as well as DNA damaging chemotherapeutics. Conclusions: Exome sequencing identified subsets of patients with loss of function events in DNA repair genes that may be associated with benefit from PARP inhibitors and platinum agents. Somatic and germline biomarker testing revealed occurrence of BRCA1/2 as well as other DNA repair gene alterations across multiple cancers. It is imperative to explore the DNA repair pathway beyond BRCA1 and BRCA2 in patient selection for PARP inhibitors and DNA damaging agents and further investigation of this pathway is warranted in ongoing clinical trials. Citation Format: Gargi D. Basu, Tracey White, Janine LoBello, Ahmet Kurdoglu, Jeffrey Trent, Sen Peng, Matthew Halbert, Thomas Royce. Assessing germline and somatic alterations in DNA repair pathway in cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2779. doi:10.1158/1538-7445.AM2017-2779

Management of germline findings revealed throughout the course of tumor-normal whole genome sequencing in oncology.

e13113 Background: The Personalized OncoGenomics (POG) project at the BC Cancer Agency utilizes tumor-normal whole genome sequencing (WGS) to understand key driver pathways and guide personalized treatment decisions. Analysis of the germline data can reveal variants; these may be presumed pathogenic, presumed benign or of unknown significance (VUS). We have developed a process for evaluating and returning presumed pathogenic variants in known cancer susceptibility genes to patients, for counseling and validation in a clinical-accredited laboratory. Methods: Patients receive germline cancer related information as part of the consent process for participation in the POG program. A sub-committee comprised of medical geneticists, bioinformaticians, pathologists, oncologists and an ethicist review the germline results. Any variants suspicious of being an artifact undergo a technical validation step. Presumed pathogenic findings of known cancer susceptibility genes are returned to the patient by their treating oncologist and patients are referred to the Hereditary Cancer Program (HCP), for genetic counseling and clinical confirmation. Results: From June 2012 - January 2017 – 466 patients have consented to the project. To date, 39 cases (8.4%) had at least one variant that was deemed pathogenic, 86 cases had at least one VUS in a known cancer susceptibility gene. 11 out of 23 cases (47.8%) with high penetrance mutations were already known to HCP. All VUS were reviewed by the sub-committee taking in to consideration the VUS and clinical context. 8 of the subjects with pathogenic results and 3 with VUS were known to HCP before POG data was generated. A VUS in 7 cases (1.5%) was returned after review. Conclusions: The number of pathogenic variants in known cancer susceptibility genes is consistent with published oncology results. We created a process to manage clinically relevant germline findings discovered during the course of genomic research to ensure appropriate care for patients. Genetic counseling within HCP and validation of variants in the clinically accredited Cancer Genetics Laboratory enables seamless return of research generated clinically relevant germline results to affected subjects. Clinical trial information: NCT02155621.

Assessing the general population frequency of rare coding variants in the EXT1 and EXT2 genes previously implicated in hereditary multiple exostoses

Hereditary multiple exostoses (HME) is a rare childhood-onset skeletal disease linked to mutations in exostosin glycosyltransferase 1 (EXT1) or 2 (EXT2). Patients are heterozygous for either an EXT1 or EXT2 mutation, and it is widely assumed that exostosis formation and associated defects, such as growth retardation and skeletal deformities, require loss-of-heterozygosity or a second hit in affected cells. However, the relevance and phenotypic impact of many presumed pathogenic EXT variants remain uncertain. We extracted all amino acid-altering (missense) and loss of function (LoF; nonsense, frameshift, or splice-site) variants from the Exome Aggregation Consortium (ExAC), a large population-based repository of exome sequence data from diverse ancestries that has screened out severe pediatric disease, to assess the overall mutation spectrum of predicted protein-damaging variants across these two genes in the general population. We then determined whether clinically-identified, presumably pathogenic variants implicated in HME exist among healthy individuals. We found six EXT1 and four EXT2 missense mutations in ExAC, suggesting that these mutations have either been misclassified as pathogenic or are not fully penetrant. Furthermore, EXT1 is heavily selectively constrained, while EXT2 is more tolerant to protein-damaging variants, especially at its C-terminus, possibly explaining the genotype–phenotype correlation that EXT1 variants usually result in more severe disease. In conclusion, population-based exome data is a useful filter for determining whether clinically detected variants are likely pathogenic, as well as revealing biological insight into rare disease genes such as EXT1 and EXT2.

Abstract 2750: Mutations on the homologous recombination pathway in 13 cancer types

Abstract Homologous recombination (HR) is important in DNA double-strand break repair. HR defects promote carcinogenesis and are associated with selective sensitivity to PARP-inhibitors and DNA-damaging agents. We collected 1029 tumor samples in 13 cancer types and used next-generation sequencing (NGS) to survey genes in the HR pathway. NGS on 591 genes was performed using formalin-fixed paraffin-embedded samples on the Illumina NextSeq platform (Caris Life Sciences, AZ). Mutations in as low as 5% of cells can be detected with &amp;gt; 99% confidence. Deletions larger than 27bp may not be detected by this method. Pathogenic or presumed pathogenic variants are counted as mutations. The table summarizes mutation rates of 7 key genes—ATM, BRCA1, BRCA2, CHEK1, CHEK2, PALB2 and PTEN—included in this pilot study. Another 17 HR genes—ATR, ATRX, BARD1, BLM, BRIP1, FANCA, FANCC, FANCD2, FANCE, FANCF, FANCG, FANCL, MRE11A, NBN, RAD50, RAD51, and RAD51B—were also analyzed. PTEN mutations were seen in 6.3% of tumors, ATM in 5%, BRCA1 in 2%, BRCA2 in 2%, PALB2 in 1%, and CHEK2 in 1%. No CHEK1 mutations were observed. Overall, 15% of tumors carry at least one mutation in any of the 7 key genes. The highest mutation rates were seen in endometrial (43%), glioblastoma (34%), and gastric cancers (23%). The highest rates of ATM (9.7%), BRCA2 (6.5%), and PALB2 (6.5%) were seen in gastric cancer, while the highest CHEK2 (5.6%), BRCA1 (7.3%) and PTEN (44%) mutations were seen in cholangiocarcinoma, ovarian and endometrial tumors, respectively. One 53-year old pt with metastatic poorly-differentiated gastric adenocarcinoma experienced ongoing radiographic partial response and dramatic symptom relief following 4 cycles of FOLFOX without surgery; tumor analysis revealed a nonsense PALB2 (S326*) gene mutation, while the other 23 HR genes were wild type. ERCC1 showed intact expression by IHC. Mutation rates of 7 genes on the HR pathway in 13 cancer typesEndometrialATMBRCA1BRCA2CHEK1CHEK2PALB2PTENAny of the 7 genesEndometrial (N = 35)00002.9%3.0%44.1%42.9%GBM (N = 47)2.1%2.1%000030.4%34.0%Gastric (N = 31)9.7%06.5%006.5%022.6%Bladder (N = 38)2.6%05.4%00010.8%18.4%Kidney (N = 41)2.5%0005.0%010.0%17.1%Ovarian (N = 82)3.7%7.3%1.2%01.2%01.3%14.6%Breast (N = 108)4.6%2.8%1.9%00.9%1.0%3.8%13.9%Cholangiocarcinoma (N = 36)2.8%02.8%05.6%02.9%13.9%CRC(N = 254)6.3%2.0%1.6%00.4%04.0%13.0%Pancreatic (N = 62)4.8%1.6%3.2%001.7%3.3%12.9%NSCLC(N = 234)6.5%00.9%001.4%2.6%11.1%Neuroendocrine(N = 35)2.9%000005.7%8.6%Esophageal (N = 26)3.8%000004.0%7.7%Overall(N = 1029)5.0%1.6%1.6%00.8%0.8%6.3%15.2% Thus, mutation rates of at least 8 to 43% in the HR pathway are reported from 13 cancer types. This method can potentially identify responders to DNA-damaging agents including platinum. Citation Format: Joanne Xiu, Ryan Bender, Brian Abbott, Zoran Gatalica, Sandeep Reddy, Mohamed Salem, Shelly Seward. Mutations on the homologous recombination pathway in 13 cancer types. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2750.

Abstract 2266: Molecular and genomic characterization of SCLC

Abstract Introduction: Small cell lung cancer (SCLC), strongly tobacco-associated, has been described to have a heavy mutation burden, harboring high rates of TP53 and RB1 alterations. While initially responsive to radiation and chemotherapy, SCLC is characterized by eventual progression and resistance to traditional therapy. We retrospectively analyzed a molecular profiling (MP) database to identify potentially actionable alterations using a multi-platform approach which includes massively parallel sequencing. Experimental Procedures: SCLC patient samples were referred to a central CLIA laboratory (Caris Life Sciences, AZ) for MP (immunohistochemistry [IHC] and next generation sequencing [NGS]). Expression of PD1 (MRQ-22, ≥1+) on tumor infiltrating lymphocytes (TILs) and PDL1 (130021, SP142, ≥2+≥5%) in tumor cells was performed by IHC. Additional IHC (ERCC1, TOPO1) and NGS on 591 genes was performed on FFPE samples using the Illumina NextSeq platform in a subset of patient samples. All variants were detected with &amp;gt; 99% confidence. Variants are described as follows: pathogenic, presumed pathogenic, variants of unknown significance and unclassified variants (excluding SNPs). Results: 203 SCLC samples were identified, 48% were females (97) and 52% were males (106). Median age was 65 [range: 29-88]. Cancer cells expressed PDL1 in 2.5% of cases (5/203) and PD1+ TILs were detected in 38% (75/197). For comparison, internal PDL1+ in non-small cell lung carcinomas (NSCLC) was 31% (339/1098). Notable findings from IHC included ERCC1 negative status in 93% (14/15) and TOPO1 + in 70% (14/20). CNV and mutational analysis (NGS) was available for 10 and 22 patients, respectively. Amplifications were found in the following genes: CCND3, CRKL, FGF4, FGFR1 and NFKB1A (n = 1, respectively), and CCND1, CCNE1, CDKN2A and FGF3 (n = 2, respectively). As previously reported, the most frequently altered genes were TP53 (73%) and RB1 (68%). Clinically relevant pathogenic or presumed pathogenic variants included: EGFR (exon 19 deletion), BRAF (G469A), APC (T1556fs), NF1 (A1610fs, D699fs), NOTCH1 (E473fs, C332Y, G546X) and PTCH1 (N1351fs). It was thought the patient with EGFR mutation is a case of NSCLC transformation to SCLC. Variants with unknown significance or unclassified variants detected in genes with clinical relevance and of potential interest for targeted therapy in SCLC include: DDR2, cMET, RET, FGFR1/3, BRCA2, IGF1R, RICTOR and NTRK1. Conclusion: Genomic and molecular characterization of SCLC samples reveals a heterogeneous population. Several potentially actionable targets are identified by NGS. Early reported trial data suggests susceptibility of SCLC to immune checkpoint inhibitors. We observed higher levels of PD1+ TILs, however differences in antibody clones, thresholds or staining localization (tumor cells vs. stromal lymphocytes), may account for the observed overall low PD-L1 expression. Further efforts are needed to identify and validate new therapeutic targets in SCLC. Citation Format: Stephen V. Liu, Edward S. Kim, Rebecca A. Feldman, Zoran Gatalica, Jeffrey Swensen, Hossein Borghaei, Alexander I. Spira, Gerold Bepler, Sandeep Reddy, Afshin Dowlati. Molecular and genomic characterization of SCLC. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2266.

Whole-exome sequencing in undiagnosed genetic diseases: interpreting 119 trios

Purpose:Despite the recognized clinical value of exome-based diagnostics, methods for comprehensive genomic interpretation remain immature. Diagnoses are based on known or presumed pathogenic variants in genes already associated with a similar phenotype. Here, we extend this paradigm by evaluating novel bioinformatics approaches to aid identification of new gene–disease associations.Genet Med 17 10, 774–781.Methods:We analyzed 119 trios to identify both diagnostic genotypes in known genes and candidate genotypes in novel genes. We considered qualifying genotypes based on their population frequency and in silico predicted effects we also characterized the patterns of genotypes enriched among this collection of patients.Genet Med 17 10, 774–781.Results:We obtained a genetic diagnosis for 29 (24%) of our patients. We showed that patients carried an excess of damaging de novo mutations in intolerant genes, particularly those shown to be essential in mice (P = 3.4 × 10−8). This enrichment is only partially explained by mutations found in known disease-causing genes.Genet Med 17 10, 774–781.Conclusion:This work indicates that the application of appropriate bioinformatics analyses to clinical sequence data can also help implicate novel disease genes and suggest expanded phenotypes for known disease genes. These analyses further suggest that some cases resolved by whole-exome sequencing will have direct therapeutic implications.Genet Med 17 10, 774–781.

95 Identification Of Likely Pathogenic Variants In Patients With Bicuspid Aortic Valve: Correlation Of Complex Genotype With A More Severe Aortic Phenotype

IntroductionThe common heritable condition of Bicuspid Aortic Valve (BAV) is phenotypically heterogeneous, with valve dysfunction and aortopathy the major complications. We report overrepresentation of rare, likely pathogenic variants in target...