RESUMO
Loss of Heterozygosity (LOH) due to mitotic recombination is frequently associated with the development of various cancers (e.g. retinoblastoma). LOH is also an important source of genetic diversity, especially in organisms where meiosis is infrequent. Irc20 is a putative helicase, and E3 ubiquitin ligase involved in DNA double-strand break repair pathway. We analyzed genome-wide LOH events, gross chromosomal changes, small insertion-deletions and single nucleotide mutations in eleven S. cerevisiae mutation accumulation lines of irc20∆, which underwent 50 mitotic bottlenecks. LOH enhancement in irc20∆ was small (1.6 fold), but statistically significant as compared to the wild type. Short (≤ 1â¯kb) and long (> 10â¯kb) LOH tracts were significantly enhanced in irc20∆. Both interstitial and terminal LOH events were also significantly enhanced in irc20∆ compared to the wild type. LOH events in irc20∆ were more telomere proximal and away from centromeres compared to the wild type. Gross chromosomal changes, single nucleotide mutations and in-dels were comparable between irc20∆ and wild type. Locus based and genome-wide analysis of meiotic recombination showed that meiotic crossover frequencies are not altered in irc20∆. These results suggest Irc20 primarily regulates mitotic recombination and does not affect meiotic crossovers. Our results suggest that the IRC20 gene is important for regulating LOH frequency and distribution.
Assuntos
Perda de Heterozigosidade , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , DNA Helicases/metabolismo , DNA Helicases/genética , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitina-Proteína Ligases/genética , Mitose , Mutação , Reparo do DNA , Meiose , Quebras de DNA de Cadeia DuplaRESUMO
According to Mendel's second law, chromosomes segregate randomly in meiosis. Non-random segregation is primarily known for cases of selfish meiotic drive in females, in which particular alleles bias their own transmission into the oocyte. Here we report a rare example of unselfish meiotic drive for crossover inheritance in the clonal raider ant, Ooceraea biroi, in which both alleles are co-inherited at all loci across the entire genome. This species produces diploid offspring parthenogenetically via fusion of two haploid nuclei from the same meiosis. This process should cause rapid genotypic degeneration due to loss of heterozygosity, which results if crossover recombination is followed by random (Mendelian) segregation of chromosomes. However, by comparing whole genomes of mothers and daughters, we show that loss of heterozygosity is exceedingly rare, raising the possibility that crossovers are infrequent or absent in O. biroi meiosis. Using a combination of cytology and whole-genome sequencing, we show that crossover recombination is, in fact, common but that loss of heterozygosity is avoided because crossover products are faithfully co-inherited. This results from a programmed violation of Mendel's law of segregation, such that crossover products segregate together rather than randomly. This discovery highlights an extreme example of cellular 'memory' of crossovers, which could be a common yet cryptic feature of chromosomal segregation.
Assuntos
Formigas , Cromátides , Heterozigoto , Partenogênese , Animais , Partenogênese/genética , Formigas/genética , Formigas/fisiologia , Cromátides/genética , Feminino , Meiose/genética , Recombinação Genética , Troca Genética , Perda de HeterozigosidadeRESUMO
BACKGROUND: While soft tissue sarcomas affect younger patients, few studies have assessed the distribution of underlying pathogenic germline variants. PATIENTS AND METHODS: We retrospectively identified all pediatric and young adult patients (0-22 years) at Haukeland University Hospital, Norway (1981-2019), through clinical and pathological records. We identified n = 46 eligible patients. From these 46 patients, adequate material representing normal tissue was available for n = 41 cases (n = 24 diagnosed with rhabdomyosarcoma, 9 with synovial sarcomas, 2 with Ewing sarcomas, and 6 without further classification), with matching tumor tissue for n = 40. Normal tissue samples were analyzed for germline pathogenic variants (PVs) by targeted sequencing of 360 cancer genes. RESULTS: Out of the 41 analyzed cases, we found PVs or likely PVs in 7 (17%). These variants were found in TP53, MUTYH, FANCC, DICER1, FANCA, MYO3A, and MYO5B. Supporting the causality of these PVs, four cases revealed loss of heterozygosity (LOH) of the wild-type allele in the tumor tissue, one patient with a PV in DICER1 had a second somatic variant in DICER1, and a patient with a PV in TP53 had the altered allele amplified in the tumor. For three out of five with available family history, a history of other cancers in relatives was recorded. Among genes with variants of uncertain significance, CHD1L was of particular interest, revealing a stop-gain and a missense variant. INTERPRETATION: A high fraction of young patients with soft tissue sarcoma harbor PVs. Among the genes affected, we substantiate a potential role of MYO5B and propose a potential role for MYO3A.
Assuntos
Mutação em Linhagem Germinativa , Humanos , Masculino , Criança , Adolescente , Feminino , Adulto Jovem , Estudos Retrospectivos , Pré-Escolar , Lactente , Sarcoma/genética , Sarcoma/patologia , Recém-Nascido , Adulto , Noruega , Predisposição Genética para Doença , Sarcoma Sinovial/genética , Sarcoma Sinovial/patologia , Rabdomiossarcoma/genética , Rabdomiossarcoma/patologia , Perda de Heterozigosidade , Neoplasias de Tecidos Moles/genética , Neoplasias de Tecidos Moles/patologia , Sarcoma de Ewing/genética , Sarcoma de Ewing/patologiaRESUMO
Osteosarcoma is a primary bone tumor that exhibits a complex genomic landscape characterized by gross chromosomal abnormalities. Osteosarcoma patients often develop metastatic disease, resulting in limited therapeutic options and poor survival rates. To gain knowledge on the mechanisms underlying osteosarcoma heterogeneity and metastatic process, it is important to obtain a detailed profile of the genomic alterations that accompany osteosarcoma progression. We performed WGS on multiple tissue samples from six patients with osteosarcoma, including the treatment naïve biopsy of the primary tumor, resection of the primary tumor after neoadjuvant chemotherapy, local recurrence, and distant metastases. A comprehensive analysis of single-nucleotide variants (SNVs), structural variants, copy number alterations (CNAs), and chromothripsis events revealed the genomic heterogeneity during osteosarcoma progression. SNVs and structural variants were found to accumulate over time, contributing to an increased complexity of the genome of osteosarcoma during disease progression. Phylogenetic trees based on SNVs and structural variants reveal distinct evolutionary patterns between patients, including linear, neutral, and branched patterns. The majority of osteosarcomas showed variable copy number profiles or gained whole-genome doubling in later occurrences. Large proportions of the genome were affected by loss of heterozygosity (LOH), although these regions remain stable during progression. Additionally, chromothripsis is not confined to a single early event, as multiple other chromothripsis events may appear in later occurrences. Together, we provide a detailed analysis of the complex genome of osteosarcomas and show that five of six osteosarcoma genomes are highly dynamic and variable during progression.
Assuntos
Neoplasias Ósseas , Variações do Número de Cópias de DNA , Progressão da Doença , Osteossarcoma , Humanos , Osteossarcoma/genética , Osteossarcoma/patologia , Neoplasias Ósseas/genética , Neoplasias Ósseas/patologia , Masculino , Feminino , Adulto , Polimorfismo de Nucleotídeo Único , Perda de Heterozigosidade , Sequenciamento Completo do Genoma , Cromotripsia , Adolescente , Genoma HumanoRESUMO
The adaptation of gene deletion methods based on the CRISPR-Cas9 system has facilitated the genetic manipulation of the pathogenic yeast Candida albicans, because homozygous mutants of this diploid fungus can now be generated in a single step, allowing the rapid screening of candidate genes for their involvement in a phenotype of interest. However, the Cas9-mediated double-strand breaks at the target site may result in an undesired loss of heterozygosity (LOH) on the affected chromosome and cause phenotypic alterations that are not related to the function of the investigated gene. In our present study, we harnessed Cas9-facilitated gene deletion to probe a set of genes that are constitutively overexpressed in strains containing hyperactive forms of the transcription factor Mrr1 for a possible contribution to the fluconazole resistance of such strains. To this aim, we used gene deletion cassettes containing two different dominant selection markers, caSAT1 and HygB, which confer resistance to nourseothricin and hygromycin, respectively, for simultaneous genomic integration in a single step, hypothesizing that this would minimize undesired LOH events at the target locus. We found that selection for resistance to both nourseothricin and hygromycin strongly increased the proportion of homozygous deletion mutants among the transformants compared with selection on media containing only one of the antibiotics, but it did not avoid undesired LOH events. Our results demonstrate that LOH on the target chromosome is a significant problem when using Cas9 for the generation of C. albicans gene deletion mutants, which demands a thorough examination of recombination events at the target site. IMPORTANCE: Candida albicans is one of the medically most important fungi and a model organism to study fungal pathogenicity. Investigating gene function in this diploid yeast has been facilitated by the adaptation of gene deletion methods based on the bacterial CRISPR-Cas9 system, because they enable the generation of homozygous mutants in a single step. We found that, in addition to increasing the efficiency of gene replacement by selection markers, the Cas9-mediated double-strand breaks also result in frequent loss of heterozygosity on the same chromosome, even when two different selection markers were independently integrated into the two alleles of the target gene. Since loss of heterozygosity for other genes can result in phenotypic alterations that are not caused by the absence of the target gene, these findings show that it is important to thoroughly analyze recombination events at the target locus when using Cas9 to generate gene deletion mutants in C. albicans.
Assuntos
Sistemas CRISPR-Cas , Candida albicans , Perda de Heterozigosidade , Recombinação Genética , Candida albicans/genética , Candida albicans/efeitos dos fármacos , Deleção de Genes , Farmacorresistência Fúngica/genética , Antifúngicos/farmacologia , Fluconazol/farmacologia , Higromicina B/farmacologia , Proteína 9 Associada à CRISPR/genética , Edição de Genes/métodos , Estreptotricinas/farmacologia , Marcadores GenéticosRESUMO
BACKGROUND: Talazoparib monotherapy in patients with germline BRCA-mutated, early-stage triple-negative breast cancer (TNBC) showed activity in the neoadjuvant setting in the phase II NEOTALA study (NCT03499353). These biomarker analyses further assessed the mutational landscape of the patients enrolled in the NEOTALA study. METHODS: Baseline tumor tissue from the NEOTALA study was tested retrospectively using FoundationOne®CDx. To further hypothesis-driven correlative analyses, agnostic heat-map visualizations of the FoundationOne®CDx tumor dataset were used to assess overall mutational landscape and identify additional candidate predictive biomarkers of response. RESULTS: All patients enrolled (N = 61) had TNBC. In the biomarker analysis population, 75.0% (39/52) and 25.0% (13/52) of patients exhibited BRCA1 and BRCA2 mutations, respectively. Strong concordance (97.8%) was observed between tumor BRCA and germline BRCA mutations, and 90.5% (38/42) of patients with tumor BRCA mutations evaluable for somatic-germline-zygosity were predicted to exhibit BRCA loss of heterozygosity (LOH). No patients had non-BRCA germline DNA damage response (DDR) gene variants with known/likely pathogenicity, based on a panel of 14 non-BRCA DDR genes. Ninety-eight percent of patients had TP53 mutations. Genomic LOH, assessed continuously or categorically, was not associated with response. CONCLUSION: The results from this exploratory biomarker analysis support the central role of BRCA and TP53 mutations in tumor pathobiology. Furthermore, these data support assessing germline BRCA mutational status for molecular eligibility for talazoparib in patients with TNBC.
Assuntos
Proteína BRCA1 , Proteína BRCA2 , Mutação em Linhagem Germinativa , Terapia Neoadjuvante , Ftalazinas , Neoplasias de Mama Triplo Negativas , Humanos , Feminino , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/patologia , Ftalazinas/uso terapêutico , Ftalazinas/administração & dosagem , Proteína BRCA2/genética , Proteína BRCA1/genética , Terapia Neoadjuvante/métodos , Pessoa de Meia-Idade , Estudos Retrospectivos , Adulto , Biomarcadores Tumorais/genética , Idoso , Perda de HeterozigosidadeRESUMO
Genetic alterations in the retinoblastoma susceptibility gene (RB1) are present in up to 40% of triple-negative breast cancers (BCs) and frequent in tumors with neuroendocrine differentiation, including small cell neuroendocrine carcinoma. Data on RB1 genetic alterations in estrogen receptor (ER)-positive BCs are scarce. In this study, we sought to define the morphologic, immunohistochemical, and genetic features of ER-positive BCs harboring somatic alterations in RB1, with emphasis on neuroendocrine differentiation. ER-positive BCs with pathogenic RB1 genetic alterations were identified in <1% of cases (N = 55) from a cohort of 6026 BCs previously subjected to targeted next-generation sequencing, including 23 primary BCs (pBCs) and 32 recurrent/metastatic BCs (mBCs). In cases where loss of heterozygosity of the wild-type RB1 allele could be assessed (93%, 51/55), most pBCs (82%, 18/22) and mBCs (90%, 26/29) exhibited biallelic RB1 inactivation, primarily through loss-of-function mutation and loss of heterozygosity (98%, 43/44). Upon histologic review, a subset of RB1-altered tumors exhibited neuroendocrine morphology (13%, 7/55), which correlated with expression of neuroendocrine markers (39%, 9/23) in both pBCs (27%, 3/11) and mBCs (50%, 6/12). Loss of Rb protein expression was observed in BCs with biallelic RB1 loss only, with similar frequency in pBCs (82%, 9/11) and mBCs (75%, 9/12). All cases with neuroendocrine marker expression (n = 9) and/or neuroendocrine morphology (n = 7) harbored biallelic genetic inactivation of RB1 and exhibited Rb loss of expression. TP53 (53%, 29/55) and PIK3CA (45%, 25/55) were the most frequently comutated genes across the cohort. Overall, these findings suggest that ER-positive BCs with biallelic RB1 genetic alterations frequently exhibit Rb protein loss, which correlates with neuroendocrine differentiation in select BCs. This study provides insights into the molecular and phenotypic heterogeneity of BCs with RB1 genetic inactivation, underscoring the need for further research into the potential clinical implications associated with these tumors.
Assuntos
Neoplasias da Mama , Carcinoma Neuroendócrino , Receptores de Estrogênio , Proteínas de Ligação a Retinoblastoma , Ubiquitina-Proteína Ligases , Humanos , Feminino , Proteínas de Ligação a Retinoblastoma/genética , Receptores de Estrogênio/metabolismo , Receptores de Estrogênio/genética , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Neoplasias da Mama/metabolismo , Carcinoma Neuroendócrino/genética , Carcinoma Neuroendócrino/patologia , Carcinoma Neuroendócrino/metabolismo , Ubiquitina-Proteína Ligases/genética , Mutação , Diferenciação Celular , Pessoa de Meia-Idade , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/análise , Idoso , Adulto , Perda de HeterozigosidadeRESUMO
BACKGROUND: Copy number variation (CNV) is a key genetic characteristic for cancer diagnostics and can be used as a biomarker for the selection of therapeutic treatments. Using data sets established in our previous study, we benchmark the performance of cancer CNV calling by six most recent and commonly used software tools on their detection accuracy, sensitivity, and reproducibility. In comparison to other orthogonal methods, such as microarray and Bionano, we also explore the consistency of CNV calling across different technologies on a challenging genome. RESULTS: While consistent results are observed for copy gain, loss, and loss of heterozygosity (LOH) calls across sequencing centers, CNV callers, and different technologies, variation of CNV calls are mostly affected by the determination of genome ploidy. Using consensus results from six CNV callers and confirmation from three orthogonal methods, we establish a high confident CNV call set for the reference cancer cell line (HCC1395). CONCLUSIONS: NGS technologies and current bioinformatics tools can offer reliable results for detection of copy gain, loss, and LOH. However, when working with a hyper-diploid genome, some software tools can call excessive copy gain or loss due to inaccurate assessment of genome ploidy. With performance matrices on various experimental conditions, this study raises awareness within the cancer research community for the selection of sequencing platforms, sample preparation, sequencing coverage, and the choice of CNV detection tools.
Assuntos
Biologia Computacional , Variações do Número de Cópias de DNA , Sequenciamento de Nucleotídeos em Larga Escala , Perda de Heterozigosidade , Neoplasias , Software , Humanos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Neoplasias/genética , Biologia Computacional/métodos , Diploide , Genoma Humano , Linhagem Celular Tumoral , Reprodutibilidade dos Testes , Análise de Sequência de DNA/métodosRESUMO
Chromosomal microarray, including single-nucleotide polymorphism (SNP) array and array comparative genomic hybridization (aCGH), enables the detection of DNA copy-number loss and/or gain associated with unbalanced chromosomal aberrations. In addition, SNP array and aCGH with SNP component also detect copy-neutral loss of heterozygosity (CN-LOH). Here we describe the chromosomal microarray procedure from the sample preparation using extracted DNA to the scanning of the array chip.
Assuntos
Hibridização Genômica Comparativa , Neoplasias , Análise de Sequência com Séries de Oligonucleotídeos , Polimorfismo de Nucleotídeo Único , Humanos , Hibridização Genômica Comparativa/métodos , Neoplasias/genética , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Perda de Heterozigosidade , Variações do Número de Cópias de DNA , Aberrações CromossômicasRESUMO
Loss of heterozygosity (LOH) has been reported to occur in HLA regions in cervical intraepithelial neoplasia (CIN) and cervical cancer. However, the details of how this is related to the progression of CIN have been unclear. In this study, we examined the human papillomavirus (HPV) antigen-presenting capacity of people with CIN and the significance of LOH of HLA class I in the progression of CIN. It was shown that differences in antigen-presenting capacity among each case depended on HLA types, not HPV genotypes. Focusing on the HLA type, there was a positive correlation between antigen-presenting capacity against HPV and the frequency of allelic loss. Furthermore, the lost HLA-B alleles had a higher HPV antigen-presenting capacity than intact alleles. In addition, frequency of LOH of HLA class I was significantly higher in advanced CIN (CIN2-3) than in cervicitis or early-stage CIN (CIN1): around half of CIN2-3 had LOH of any HLA class I. Moreover, the antigen-presenting capacity against E5, which is the HPV proteins that facilitate viral escape from this immune surveillance by suppressing HLA class I expression, had the most significant impact on the LOH in HLA-B. This study suggests that HPV evades immune surveillance mechanisms when host cells lose the capacity for antigen presentation by HLA class I molecules, resulting in long-term infection and progression to advanced lesions.
Assuntos
Antígenos de Histocompatibilidade Classe I , Perda de Heterozigosidade , Infecções por Papillomavirus , Displasia do Colo do Útero , Neoplasias do Colo do Útero , Humanos , Displasia do Colo do Útero/imunologia , Displasia do Colo do Útero/genética , Displasia do Colo do Útero/virologia , Displasia do Colo do Útero/patologia , Feminino , Neoplasias do Colo do Útero/imunologia , Neoplasias do Colo do Útero/genética , Antígenos de Histocompatibilidade Classe I/imunologia , Antígenos de Histocompatibilidade Classe I/genética , Infecções por Papillomavirus/imunologia , Infecções por Papillomavirus/genética , Apresentação de Antígeno/imunologia , Adulto , Alelos , Papillomaviridae/imunologia , Vigilância Imunológica , Pessoa de Meia-Idade , GenótipoRESUMO
PURPOSE: Ovarian cancer (OC) is characterized by a high recurrence rate, and homologous recombination deficiency (HRD) is an important biomarker in the clinical management of OC. We investigated the differences in clinical genomic profiles between the primary and platinum-sensitive recurrent OC (PSROC), focusing on HRD status. MATERIALS AND METHODS: A total of 40 formalin-fixed paraffin-embedded (FFPE) tissues of primary tumors and their first platinum-sensitive recurrence from 20 OC patients were collected, and comprehensive genomic profiling (CGP) analysis of FoundationOne®CDx (F1CDx) was applied to explore the genetic (dis)similarities of the primary and recurrent tumors. RESULTS: By comparing between paired samples, we found that genomic loss of heterozygosity (gLOH) score had a high intra-patient correlation (r2 = 0.79) and that short variants (including TP53, BRCA1/2 and NOTCH1 mutations), tumor mutational burden (TMB) and microsatellite stability status remained stable. The frequency of (likely) pathological BRCA1/2 mutations was 30% (12/40) in all samples positively correlated with gLOH scores, but the proportion of gLOH-high status (score > 16%) was 50% (10/20) and 55% (11/20) in the primary and recurrent samples, respectively. An additional 20% (4/20) of patients needed attention, a quarter of which carried the pathological BRCA1 mutation but had a gLOH-low status (gLOH < 16%), and three-quarters had different gLOH status in primary-recurrent pairs. Furthermore, we observed the PSROC samples had higher gLOH scores (16.1 ± 9.24 vs. 19.4 ± 11.1, p = 0.007), more CNVs (36.1% vs. 15.1% of discordant genomic alternations), and significant enrichment of altered genes in TGF-beta signaling and Hippo signaling pathways (p < 0.05 for all) than their paired primaries. Lastly, mutational signature and oncodrive gene analyses showed that the computed mutational signature similarity in the primary and recurrent tumors were best matched the COSMI 3 signature (Aetiology of HRD) and had consistent candidate cancer driver genes of MSH2, NOTCH1 and MSH6. CONCLUSION: The high genetic concordance of the short variants remains stable along OC recurrence. However, the results reveal significantly higher gLOH scores in the recurrent setting than in paired primaries, supporting further clinically instantaneity HRD assay strategy.
Assuntos
Genômica , Recidiva Local de Neoplasia , Neoplasias Ovarianas , Humanos , Feminino , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/patologia , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/genética , Genômica/métodos , Idoso , Mutação , Perda de Heterozigosidade , Adulto , Biomarcadores Tumorais/genética , Perfilação da Expressão Gênica/métodosRESUMO
The p53 tumor suppressor, encoded by the TP53 gene, serves as a major barrier against malignant transformation. Patients with Li-Fraumeni syndrome (LFS) inherit a mutated TP53 allele from one parent and a wild-type TP53 allele from the other. Subsequently, the wild-type allele is lost and only the mutant TP53 allele remains. This process, which is termed loss of heterozygosity (LOH), results in only mutant p53 protein expression. We used primary dermal fibroblasts from LFS patients carrying the hotspot p53 gain-of-function pathogenic variant, R248Q to study the LOH process and characterize alterations in various pathways before and after LOH. We previously described the derivation of mutant p53 reactivating peptides, designated pCAPs (p53 Conformation Activating Peptides). In this study, we tested the effect of lead peptide pCAP-250 on LOH and on its associated cellular changes. We report that treatment of LFS fibroblasts with pCAP-250 prevents the accumulation of mutant p53 protein, inhibits LOH, and alleviates its cellular consequences. Furthermore, prolonged treatment with pCAP-250 significantly reduces DNA damage and restores long-term genomic stability. pCAPs may thus be contemplated as a potential preventive treatment to prevent or delay early onset cancer in carriers of mutant p53.
Assuntos
Fibroblastos , Síndrome de Li-Fraumeni , Perda de Heterozigosidade , Proteína Supressora de Tumor p53 , Síndrome de Li-Fraumeni/genética , Síndrome de Li-Fraumeni/patologia , Humanos , Proteína Supressora de Tumor p53/metabolismo , Proteína Supressora de Tumor p53/genética , Fibroblastos/metabolismo , Fibroblastos/efeitos dos fármacos , Fibroblastos/patologia , Dano ao DNA , MutaçãoRESUMO
We analyzed variations in the epidermal growth factor receptor (EGFR) gene and 5'-upstream region to identify potential molecular predictors of treatment response in primary epithelial ovarian cancer. Tumor tissues collected during debulking surgery from the prospective multicenter OVCAD study were investigated. Copy number variations in the human endogenous retrovirus sequence human endogenous retrovirus K9 (HERVK9) and EGFR Exons 7 and 9, as well as repeat length and loss of heterozygosity of polymorphic CA-SSR I and relative EGFR mRNA expression were determined quantitatively. At least one EGFR variation was observed in 94% of the patients. Among the 30 combinations of variations discovered, enhanced platinum sensitivity (n = 151) was found dominantly with HERVK9 haploidy and Exon 7 tetraploidy, overrepresented among patients with survival ≥120 months (24/29, p = .0212). EGFR overexpression (≥80 percentile) was significantly less likely in the responders (17% vs. 32%, p = .044). Multivariate Cox regression analysis, including age, FIGO stage, and grade, indicated that the patients' subgroup was prognostically significant for CA-SSR I repeat length <18 CA for both alleles (HR 0.276, 95% confidence interval 0.109-0.655, p = .001). Although EGFR variations occur in ovarian cancer, the mRNA levels remain low compared to other EGFR-mutated cancers. Notably, the inherited length of the CA-SSR I repeat, HERVK9 haploidy, and Exon 7 tetraploidy conferred three times higher odds ratio to survive for more than 10 years under therapy. This may add value in guiding therapies if determined during follow-up in circulating tumor cells or circulating tumor DNA and offers HERVK9 as a potential therapeutic target.
Assuntos
Cromossomos Humanos Par 7 , Variações do Número de Cópias de DNA , Receptores ErbB , Neoplasias Ovarianas , Humanos , Feminino , Receptores ErbB/genética , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/mortalidade , Neoplasias Ovarianas/patologia , Neoplasias Ovarianas/tratamento farmacológico , Pessoa de Meia-Idade , Cromossomos Humanos Par 7/genética , Estudos Prospectivos , Idoso , Carcinoma Epitelial do Ovário/genética , Carcinoma Epitelial do Ovário/mortalidade , Carcinoma Epitelial do Ovário/patologia , Adulto , Retroelementos/genética , Fenótipo , Resistencia a Medicamentos Antineoplásicos/genética , Retrovirus Endógenos/genética , Perda de HeterozigosidadeRESUMO
Pituitary neuroendocrine tumors (PitNETs) exhibiting aggressive, treatment-refractory behavior are the rare subset that progress after surgery, conventional medical therapies, and an initial course of radiation and are characterized by unrelenting growth and/or metastatic dissemination. Two groups of patients with PitNETs were sequenced: a prospective group of patients (n = 66) who consented to sequencing prior to surgery and a retrospective group (n = 26) comprised of aggressive/higher risk PitNETs. A higher mutational burden and fraction of loss of heterozygosity (LOH) was found in the aggressive, treatment-refractory PitNETs compared to the benign tumors (p = 1.3 × 10-10 and p = 8.5 × 10-9, respectively). Within the corticotroph lineage, a characteristic pattern of recurrent chromosomal LOH in 12 specific chromosomes was associated with treatment-refractoriness (occurring in 11 of 14 treatment-refractory versus 1 of 14 benign corticotroph PitNETs, p = 1.7 × 10-4). Across the cohort, a higher fraction of LOH was identified in tumors with TP53 mutations (p = 3.3 × 10-8). A machine learning approach identified loss of heterozygosity as the most predictive variable for aggressive, treatment-refractory behavior, outperforming the most common gene-level alteration, TP53, with an accuracy of 0.88 (95% CI: 0.70-0.96). Aggressive, treatment-refractory PitNETs are characterized by significant aneuploidy due to widespread chromosomal LOH, most prominently in the corticotroph tumors. This LOH predicts treatment-refractoriness with high accuracy and represents a novel biomarker for this poorly defined PitNET category.
Assuntos
Perda de Heterozigosidade , Tumores Neuroendócrinos , Neoplasias Hipofisárias , Humanos , Perda de Heterozigosidade/genética , Neoplasias Hipofisárias/genética , Neoplasias Hipofisárias/patologia , Tumores Neuroendócrinos/genética , Tumores Neuroendócrinos/patologia , Tumores Neuroendócrinos/terapia , Masculino , Feminino , Pessoa de Meia-Idade , Adulto , Idoso , Estudos Retrospectivos , Mutação/genética , Estudos ProspectivosRESUMO
BACKGROUND: MUTYH germline monoallelic variants have been detected in a number of patients affected by breast/ovarian cancer or endometrial cancer, suggesting a potential susceptibility role, though their significance remains elusive since the disease mechanism is normally recessive. Hence, the aim of this research was to explore the hypothesis that a second hit could have arisen in the other allele in the tumor tissue. METHODS: we used Sanger sequencing and immunohistochemistry to search for a second MUTYH variant in the tumoral DNA and to assess protein expression, respectively. RESULTS: we detected one variant of unknown significance, one variant with conflicting interpretation of pathogenicity and three benign/likely benign variants; the MUTYH protein was not detected in the tumor tissue of half of the patients, and in others, its expression was reduced. CONCLUSIONS: our results fail to demonstrate that germinal monoallelic MUTYH variants increase cancer risk through a LOH (loss of heterozygosity) mechanism in the somatic tissue; however, the absence or partial loss of the MUTYH protein in many tumors suggests its dysregulation regardless of MUTYH genetic status.
Assuntos
Neoplasias da Mama , DNA Glicosilases , Neoplasias do Endométrio , Neoplasias Ovarianas , Humanos , DNA Glicosilases/genética , Feminino , Neoplasias do Endométrio/genética , Neoplasias do Endométrio/patologia , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/patologia , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Pessoa de Meia-Idade , Perda de Heterozigosidade , Predisposição Genética para Doença , Idoso , AdultoRESUMO
Loss-of-function variants in CCM1/KRIT1, CCM2/MGC4607, and CCM3/PDCD10 genes are identified in the vast majority of familial cases with multiple cerebral cavernous malformations. However, genomic DNA sequencing combined with large rearrangement screening fails to detect a pathogenic variant in 5% of the patients. We report a family with two affected members harboring multiple CCM lesions, one with severe hemorrhages and one asymptomatic. No causative variant was detected using DNA sequencing of the three CCM genes, CNV detection analysis, and RNA sequencing. However, a loss of heterozygosity in CCM2 was observed on cDNA sequences in one of the two affected members, which strongly suggested that this locus might be involved. Whole genome sequencing (WGS) identified a balanced structural variant on chromosome 7 with a breakpoint interrupting the CCM2 gene, preventing normal mRNA synthesis. These data underline the importance of WGS in undiagnosed patients with typical multiple CCM.
Assuntos
Hemangioma Cavernoso do Sistema Nervoso Central , Perda de Heterozigosidade , Linhagem , Humanos , Hemangioma Cavernoso do Sistema Nervoso Central/genética , Hemangioma Cavernoso do Sistema Nervoso Central/diagnóstico , Hemangioma Cavernoso do Sistema Nervoso Central/patologia , Feminino , Masculino , Adulto , Proteínas de Transporte/genética , Cromossomos Humanos Par 7/genética , DNA Complementar/genética , Pessoa de Meia-IdadeRESUMO
Clonal hematopoiesis (CH) is characterized by the acquisition of a somatic mutation in a hematopoietic stem cell that results in a clonal expansion. These driver mutations can be single nucleotide variants in cancer driver genes or larger structural rearrangements called mosaic chromosomal alterations (mCAs). The factors that influence the variations in mCA fitness and ultimately result in different clonal expansion rates are not well understood. We used the Passenger-Approximated Clonal Expansion Rate (PACER) method to estimate clonal expansion rate as PACER scores for 6,381 individuals in the NHLBI TOPMed cohort with gain, loss, and copy-neutral loss of heterozygosity mCAs. Our mCA fitness estimates, derived by aggregating per-individual PACER scores, were correlated (R2 = 0.49) with an alternative approach that estimated fitness of mCAs in the UK Biobank using population-level distributions of clonal fraction. Among individuals with JAK2 V617F clonal hematopoiesis of indeterminate potential or mCAs affecting the JAK2 gene on chromosome 9, PACER score was strongly correlated with erythrocyte count. In a cross-sectional analysis, genome-wide association study of estimates of mCA expansion rate identified a TCL1A locus variant associated with mCA clonal expansion rate, with suggestive variants in NRIP1 and TERT.
Assuntos
Aberrações Cromossômicas , Hematopoiese Clonal , Mosaicismo , Humanos , Hematopoiese Clonal/genética , Masculino , Feminino , Estudo de Associação Genômica Ampla , Janus Quinase 2/genética , Telomerase/genética , Telomerase/metabolismo , Perda de Heterozigosidade , Estudos Transversais , Mutação , Pessoa de Meia-Idade , Células-Tronco Hematopoéticas/metabolismo , Polimorfismo de Nucleotídeo Único , IdosoRESUMO
BACKGROUND: The absence of heterozygosity (AOH) is a kind of genomic change characterized by a long contiguous region of homozygous alleles in a chromosome, which may cause human genetic disorders. However, no method of low-pass whole genome sequencing (LP-WGS) has been reported for the detection of AOH in a low-pass setting of less than onefold. We developed a method, termed CNVseq-AOH, for predicting the absence of heterozygosity using LP-WGS with ultra-low sequencing data, which overcomes the sparse nature of typical LP-WGS data by combing population-based haplotype information, adjustable sliding windows, and recurrent neural network (RNN). We tested the feasibility of CNVseq-AOH for the detection of AOH in 409 cases (11 AOH regions for model training and 863 AOH regions for validation) from the 1000 Genomes Project (1KGP). AOH detection using CNVseq-AOH was also performed on 6 clinical cases with previously ascertained AOHs by whole exome sequencing (WES). RESULTS: Using SNP-based microarray results as reference (AOHs detected by CNVseq-AOH with at least a 50% overlap with the AOHs detected by chromosomal microarray analysis), 409 samples (863 AOH regions) in the 1KGP were used for concordant analysis. For 784 AOHs on autosomes and 79 AOHs on the X chromosome, CNVseq-AOH can predict AOHs with a concordant rate of 96.23% and 59.49% respectively based on the analysis of 0.1-fold LP-WGS data, which is far lower than the current standard in the field. Using 0.1-fold LP-WGS data, CNVseq-AOH revealed 5 additional AOHs (larger than 10 Mb in size) in the 409 samples. We further analyzed AOHs larger than 10 Mb, which is recommended for reporting the possibility of UPD. For the 291 AOH regions larger than 10 Mb, CNVseq-AOH can predict AOHs with a concordant rate of 99.66% with only 0.1-fold LP-WGS data. In the 6 clinical cases, CNVseq-AOH revealed all 15 known AOH regions. CONCLUSIONS: Here we reported a method for analyzing LP-WGS data to accurately identify regions of AOH, which possesses great potential to improve genetic testing of AOH.
Assuntos
Perda de Heterozigosidade , Redes Neurais de Computação , Sequenciamento Completo do Genoma , Humanos , Sequenciamento Completo do Genoma/métodos , Polimorfismo de Nucleotídeo Único , Genoma HumanoRESUMO
PURPOSE: Diffuse pleural mesotheliomas (DPM) with genomic near-haploidization (GNH) represent a novel subtype first recognized by The Cancer Genome Atlas project; however, its clinicopathologic and molecular features remain poorly defined. EXPERIMENTAL DESIGN: We analyzed clinical genomic profiling data from 290 patients with DPM using the Memorial Sloan Kettering-Integrated Mutation Profiling of Actionable Cancer Targets (MSK-IMPACT) assay. Allele-specific copy number analysis was performed using the Fraction and Allele-Specific Copy Number Estimates from Tumor Sequencing (FACETS) algorithm. RESULTS: A total of 210 patients were evaluable for loss of heterozygosity (LOH) analysis using FACETS from MSK-IMPACT tumor:normal sequencing data. In this cohort, GNH, defined as LOH across >80% of the genome, was detected in 10 cases (4.8%). Compared with non-GNH tumors, GNH DPMs were associated with younger age and less frequent self-reported history of occupational asbestos exposure. Histologically, GNH DPMs were enriched in biphasic subtype (80% vs. 14.5%) and showed abundant tumor-infiltrating lymphocytes (TILs). Genomic analysis revealed a higher frequency of TP53 alterations, whereas SETDB1 mutations were present in nearly all and only in this subset. The clinicopathologic and molecular findings were further validated in a separate cohort. Despite the younger age, patients with GNH DPMs had a shorter overall survival (10.9 vs. 25.4 months, P = 0.004); the poor prognostic impact of GNH remained significant after controlling for biphasic histology. Of three patients with GNH DPMs who received immune checkpoint blockade, two achieved a clinician-assessed partial response. CONCLUSIONS: GNH defines an aggressive subtype of mainly biphasic DPMs in younger patients with recurrent alterations in SETDB1 and TP53. The enrichment in biphasic histology and TILs, together with our preliminary immune checkpoint blockade response data and anecdotal clinical trial data, suggests that further evaluation of immunotherapy may be warranted in this subset.