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Previous clinical trials indicate that 10%-25% of patients received genomically matched therapy after comprehensive genomic profiling (CGP) tests. However, the clinical utility of CGP tests has not been assessed in clinical practice. We assessed the clinical utility of CGP tests for advanced or metastatic solid tumor and determined the proportion of patients receiving genomically matched therapy among those with common and non-common cancers. From August 2019 to July 2020, a total of 418 patients had undergone CGP tests, and the results were discussed through the molecular tumor board at our site. The median age of patients was 57 (range: 3-86) years. Colorectal cancer was the most common, with 47 (11%) patients. Actionable genomic alterations (median 3, range: 1-17) were identified in 368 (88.0%) of 418 patients. Druggable genomic alterations were determined in 196 (46.9%) of 418 patients through the molecular tumor board. Genomically matched therapy was administered as the subsequent line of therapy in 51 (12.2%) patients, which is comparable to the proportion we previously reported in a clinical trial (13.4%) (p = 0.6919). The proportion of patients receiving genomically matched therapy was significantly higher among those with common cancers (16.2%) than non-common cancers (9.4%) (p = 0.0365). Genomically matched therapy after the CGP tests was administered to 12.2% of patients, which is similar to the proportion reported in the previous clinical trials. The clinical utility of CGP tests in patients with common cancers greatly exceeded that in patients with non-common cancers.

Background Clinical trials have reported the efficacy of tropomyosin receptor kinase (TRK) inhibitors against neurotrophic receptor tyrosine kinase (NTRK) fusion gene-positive advanced solid tumors. The accumulated evidence of tumor-agnostic agent has made since TRK inhibitors were approved and used in clinical practice. Therefore, we have revised the ‘Japan Society of Clinical Oncology (JSCO)/Japanese Society of Medical Oncology (JSMO)-led clinical recommendations on the diagnosis and use of tropomyosin receptor kinase inhibitors in adult and pediatric patients with neurotrophic receptor tyrosine kinase fusion-positive advanced solid tumors, cooperated by the Japanese Society of Pediatric Hematology/Oncology (JSPHO)’. Methods Clinical questions regarding medical care were formulated for patients with NTRK fusion-positive advanced solid tumors. Relevant publications were searched by PubMed and Cochrane Database. Critical publications and conference reports were added manually. Systematic reviews were performed for each clinical question for the purpose of developing clinical recommendations. The committee members identified by JSCO, JSMO, and JSPHO voted to determine the level of each recommendation considering the strength of evidence, expected risks and benefits to patients, and other related factors. Thereafter, a peer review by experts nominated from JSCO, JSMO, and JSPHO, and the public comments among all societies' members was done. Results The current guideline describes 3 clinical questions and 14 recommendations for whom, when, and how NTRK fusion should be tested, and what is recommended for patients with NTRK fusion-positive advanced solid tumors. Conclusion The committee proposed 14 recommendations for performing NTRK testing properly to select patients who are likely to benefit from TRK inhibitors.

PDF file 65K, Primers used in assessment of RINT1 transcripts

PDF file 199K, RINT1 c.1334-5delA, c.1334-1_1335delGTT minigene assay

PDF file 69K, Distribution of cases and controls included in the analysis, by study center and ethnic group

BRCA1 is a high-risk susceptibility gene for breast and ovarian cancer. Pathogenic protein-truncating variants are scattered across the open reading frame, but all known missense substitutions that are pathogenic because of missense dysfunction are located in either the amino-terminal RING domain or the carboxy-terminal BRCT domain. Heterodimerization of the BRCA1 and BARD1 RING domains is a molecularly defined obligate activity. Hence, we tested every BRCA1 RING domain missense substitution that can be created by a single nucleotide change for heterodimerization with BARD1 in a mammalian two-hybrid assay. Downstream of the laboratory assay, we addressed three additional challenges: assay calibration, validation thereof, and integration of the calibrated results with other available data, such as computational evidence and patient/population observational data to achieve clinically applicable classification. Overall, we found that 15%-20% of BRCA1 RING domain missense substitutions are pathogenic. Using a Bayesian point system for data integration and variant classification, we achieved clinical classification of 89% of observed missense substitutions. Moreover, among missense substitutions not present in the human observational data used here, we find an additional 45 with concordant computational and functional assay evidence in favor of pathogenicity plus 223 with concordant evidence in favor of benignity; these are particularly likely to be classified as likely pathogenic and likely benign, respectively, once human observational data become available.

PURPOSE: Several professional societies have published guidelines for the clinical interpretation of somatic variants, which specifically address diagnostic, prognostic, and therapeutic implications. While these guidelines for the clinical interpretation of variants include data types that may be used to determine the oncogenicity of a variant (e.g., population frequency, functional and in silico data or somatic frequency), they do not provide a direct, systematic, and comprehensive set of standards and rules to classify the oncogenicity of a somatic variant. This insufficient guidance leads to inconsistent classification of rare somatic variants in cancer, generates variability in their clinical interpretation and importantly impacts patient care. Therefore, it’s essential to address this unmet need. METHODS: Clinical Genome Resource (ClinGen) Somatic Cancer Clinical Domain Working Group and ClinGen Germline/Somatic Variant Subcommittee, the Cancer Genomics Consortium (CGC), and the Variant Interpretation for Cancer Consortium (VICC) used a consensus approach to develop a Standard Operating Procedure (SOP) for the classification of oncogenicity of somatic variants. RESULTS: This comprehensive SOP has been developed to improve consistency in somatic variant classification, and has been validated on 94 somatic variants in ten common cancer-related genes. CONCLUSION: The comprehensive SOP is now available for classification of oncogenicity of a somatic variant.

Background and Purpose In Japan, cancer-gene profiling (CGP) tests, such as the OncoGuide NCC Oncopanel System, FoundationOne CDx Cancer Genome Profile (F-one), and FoundationOne Liquid CDx Cancer Genome Profile, are covered by insurance and clinically applied to promote cancer genomic medicine. Information from CGP tests is managed at the Center for Cancer Genomics and Advanced Therapeutics (C-CAT) and shared with clinicians after discussions at the Expert Panel (EP). We provide an overview of the registered pediatric CNS tumors. Methods We extracted and analyzed CNS/brain data from the C-CAT registry from June 2019 to June 2022 for ages 0–19 years. Variables such as age, gender, histopathological diagnosis, CGP test type, ECOG Performance Status (PS), registration status, genetic abnormalities, and treatments were analyzed. Results Among 1133 patients of all ages with CNS tumors, 361 (31.9%) were aged 0–19 years; Of the 918 patients aged 0-19 years with any type of cancer, those with CNS tumors accounted for 34.2%; 188 patients were aged under 10 years, 173 patients were teenagers, 174 patients were males, 187 patients were females; PS was 0 in 47.9% of patients. The most common histopathological diagnoses were medulloblastoma, diffuse midline glioma and glioblastoma. Genetic abnormalities included TP53 mutations in 85 cases, H3F3A mutations in 38 cases, STK11 mutations in 36 cases, BRAF mutations in 33 cases, BRAF-KIAA1549 fusions in 24 cases, and TMB high in 16 cases. Seventy-six patients (21.1%) were offered treatment options in the EP, and 26 patients (7.2%) received the recommended treatment, including two in physician-initiated trials, four in company trials, eleven in treatments within insurance coverage, and eleven in others. Discussion This is the first report of genetic analysis of pediatric CNS tumors on a nationwide scale. Pediatric CNS tumors have a large number of registered cases; thus, drug development for them is urgently needed.

The main aim of this study was to screen epigenetic modifier genes and known breast cancer driver genes for germline mutations in non-BRCA1/2 (BRCAx) breast cancer families in order to identify novel susceptibility genes of moderate–high penetrance. We screened 264 candidate susceptibility genes in 656 index cases from non-BRCA1/2 families. Potentially pathogenic candidate mutations were then genotyped in all available family members for the assessment of co-segregation of the variant with disease in the family in order to estimate the breast cancer risks associated with these mutations. For 11 of the candidate susceptibility genes, we screened an additional 800 non-BRCA1/2 breast cancer cases and 787 controls. Only two genes, CHD8 and USH2A showed any evidence of an increased risk of breast cancer (RR = 2.40 (95% CI 1.0–7.32) and 2.48 (95% CI 1.11–6.67), respectively). We found no convincing evidence that epigenetic modifier and known breast cancer driver genes carry germline mutations that increase breast cancer risk. USH2A is no longer regarded as a breast cancer driver gene and seems an implausible candidate given its association with Usher syndrome. However, somatic mutations in CHD8 have been recently reported, making it an even more promising candidate, but further analysis of CHD8 in very large cohorts of families or case–control studies would be required to determine if it is a moderate-risk breast cancer susceptibility gene.

Background: Systemic lupus erythematosus (SLE) is characterised by dysregulation of autoreactive lymphocytes and antigen-presenting cells. Signalling through Toll-like receptor 9 (TLR9), a mediator of innate immune responses, has a role in activation of dendritic cells and autoreactive B cells. Objective: To investigate whether TLR9 polymorphisms are associated with an increased risk of SLE. Methods: DNA samples were obtained from 220 Japanese patients with SLE (with >4 American College of Rheumatology criteria for SLE) and 203 controls. The genetic variations of TLR9 were detected by PCR, followed by DNA sequencing. The promoter and enhancer activities of TLR9 were measured by luciferase reporter gene assay. The titres of anti-dsDNA antibodies in sera from control or TLR9-deficient mice were analysed by ELISA. Results: The G allele at position +1174 (located in intron 1 of TLR9) is closely associated with an increased risk of SLE (p = 0.029). Furthermore, patients with SLE tend to have C allele at position −1486 (p = 0.11). Both alleles down regulate TLR9 expression by reporter gene assay. TLR9-deficient mice under a C57BL/6 background possess higher titres of anti-dsDNA serum antibodies than control C57BL/6 mice. Conclusions: These results indicate that the presence of the G allele at position +1174 of TLR9 predisposes humans to an increased risk of SLE. It is speculated that TLR9 normally prevents the development of human SLE.

Gitelman syndrome (GS) is a renal tubular disorder characterized by hypokalemia, hypomagnesemia, metabolic alkalosis and hypocalciuria due to defective tubular reabsorption of magnesium and potassium. This disease is caused by mutations of the thiazide-sensitive Na-Cl cotransporter (NCCT) gene, SLC12A3. Manifestations of GS are heterogeneous, from asymptomatic to mild symptoms of cramps and easy fatigue, to tetany and paralysis. Polydipsia, polyuria, and nocturia are also frequent in GS patients. Here we describe two Japanese patients with GS followed as nocturnal enuresis. In the first patient, occasional muscle cramps, easy fatigue and headache led to the diagnosis of GS. The parents of this patient reported that he had been affected by polydipsia and polyuria, especially nocturnal enuresis from early childhood. The second patient was referred to our clinic because of muscular weakness and cramps. He had a past history of transient muscle weakness and muscle cramps. He had also suffered from nocturnal enuresis since 3 yr of age. Laboratory findings of these patients were consistent with those of GS. Sequencing analysis of the SLC12A3 gene from two patients showed four mutations, which were previously reported. In our two patients, their manifestations had been underestimated and the correct diagnosis was delayed. GS is generally likely to be benign, however signs of GS are found in early childhood. Especially, we must recognize that nocturnal enuresis is frequent in symptoms of GS.

Approximately half of the familial aggregation of breast cancer remains unexplained. A multiple-case breast cancer family exome-sequencing study identified three likely pathogenic mutations in RINT1 (NM_021930.4) not present in public sequencing databases: RINT1 c.343C>T (p.Q115X), c.1132_1134del (p.M378del), and c.1207G>T (p.D403Y). On the basis of this finding, a population-based case–control mutation-screening study was conducted that identified 29 carriers of rare (minor allele frequency < 0.5%), likely pathogenic variants: 23 in 1,313 early-onset breast cancer cases and six in 1,123 frequency-matched controls [OR, 3.24; 95% confidence interval (CI), 1.29–8.17; P = 0.013]. RINT1 mutation screening of probands from 798 multiple-case breast cancer families identified four additional carriers of rare genetic variants. Analysis of the incidence of first primary cancers in families of women carrying RINT1 mutations estimated that carriers were at increased risk of Lynch syndrome–spectrum cancers [standardized incidence ratio (SIR), 3.35; 95% CI, 1.7–6.0; P = 0.005], particularly for relatives diagnosed with cancer under the age of 60 years (SIR, 10.9; 95% CI, 4.7–21; P = 0.0003). Significance: The work described in this study adds RINT1 to the growing list of genes in which rare sequence variants are associated with intermediate levels of breast cancer risk. Given that RINT1 is also associated with a spectrum of cancers with mismatch repair defects, these findings have clinical applications and raise interesting biological questions. Cancer Discov; 4(7); 804–15. ©2014 AACR. See related commentary by Ngeow and Eng, p. 762 This article is highlighted in the In This Issue feature, p. 745

Background Characterising genetic diversity through the analysis of massively parallel sequencing (MPS) data offers enormous potential to significantly improve our understanding of the genetic basis for observed phenotypes, including predisposition to and progression of complex human disease. Great challenges remain in resolving genetic variants that are genuine from the millions of artefactual signals. Results FAVR is a suite of new methods designed to work with commonly used MPS analysis pipelines to assist in the resolution of some of the issues related to the analysis of the vast amount of resulting data, with a focus on relatively rare genetic variants. To the best of our knowledge, no equivalent method has previously been described. The most important and novel aspect of FAVR is the use of signatures in comparator sequence alignment files during variant filtering, and annotation of variants potentially shared between individuals. The FAVR methods use these signatures to facilitate filtering of (i) platform and/or mapping-specific artefacts, (ii) common genetic variants, and, where relevant, (iii) artefacts derived from imbalanced paired-end sequencing, as well as annotation of genetic variants based on evidence of co-occurrence in individuals. We applied conventional variant calling applied to whole-exome sequencing datasets, produced using both SOLiD and TruSeq chemistries, with or without downstream processing by FAVR methods. We demonstrate a 3-fold smaller rare single nucleotide variant shortlist with no detected reduction in sensitivity. This analysis included Sanger sequencing of rare variant signals not evident in dbSNP131, assessment of known variant signal preservation, and comparison of observed and expected rare variant numbers across a range of first cousin pairs. The principles described herein were applied in our recent publication identifying XRCC2 as a new breast cancer risk gene and have been made publically available as a suite of software tools. Conclusions FAVR is a platform-agnostic suite of methods that significantly enhances the analysis of large volumes of sequencing data for the study of rare genetic variants and their influence on phenotypes.

Objective:The primary aim of this study was to investigate the association of polymorphisms of TRAF1-C5, a newly identified rheumatoid arthritis (RA) risk locus in Caucasians, with susceptibility to RA and systemic lupus erythematosus (SLE) in Japanese populations. Gene expression levels of TRAF1 and C5 to assess the functional significance of genotypes were also analysed.Methods:A multicentre association study consisting of 4 RA case-control series (4397 cases and 2857 controls) and 3 SLE case-control series (591 cases and 2199 shared controls) was conducted. Genotyping was performed using TaqMan genotyping assay for two single nucleotide polymorphisms (SNPs) that showed the best evidence of association in the previous Caucasian studies. Quantifications of TRAF1 and C5 expression were performed with TaqMan expression assay.Results:Significant differences in allele frequency for both SNPs were observed between RA and control subjects (combined odds ratio = 1.09), while no significant difference was detected between patients with SLE and controls. Interestingly, alleles rs3761847 A and rs10818488 G had increased the risk for RA in the present study, while they decreased the risk in the original studies. A significant difference was found between risk allele carriers and non-carriers of rs10818488 for the expression level of TRAF1 in phorbol myristate acetate-stimulated lymphoblastoid cell lines (p = 0.04).Conclusion:Association of TRAF1-C5 locus with RA susceptibility was detected in the Japanese populations with modest magnitude, while no significant association was observed for SLE. Significant positive effect of genotype on the expression of TRAF1 might support the genetic association between TRAF1 and RA.

Objective STAT4 encodes a transcriptional factor that transmits signals induced by several key cytokines, and it might be a key molecule in the development of autoimmune diseases. Recently, a STAT4 haplotype was reported to be associated with rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE) in Caucasian populations. This was replicated in a Korean RA population. Interestingly, the degree of risk of RA susceptibility with the STAT4 haplotype was similar in the Caucasian and Korean populations. The present study was undertaken to investigate the effect of STAT4 on susceptibility to RA and SLE in the Japanese. Methods We performed an association study using 3 independent Japanese RA case–control populations (total 3,567 cases and 2,199 controls) and 3 independent Japanese SLE populations (total 591 cases). All samples were genotyped using the TaqMan fluorogenic 5′ nuclease assay for single-nucleotide polymorphism (SNP) rs7574865, which tags the susceptibility haplotype. The association of the SNP with disease susceptibility in each case–control study was calculated using Fisher's exact test, and the results were combined, using the Mantel-Haenszel method, to obtain combined odds ratios (ORs). Results We observed a significant association of the STAT4 polymorphism with susceptibility to both RA and SLE. The combined ORs for RA and SLE, respectively, were 1.27 (P = 8.4 × 10−9) and 1.61 (P = 2.1 × 10−11) for allele frequency distribution; these ORs were quite similar to those previously observed in the Caucasian population. Conclusion We conclude that STAT4 is associated with RA and SLE in the Japanese. Our results indicate that STAT4 is a common genetic risk factor for autoimmune diseases, with similar strength across major racial groups.