Clinical evaluation of RB1 genetic testing reveals novel mutations in Vietnamese patients with retinoblastoma.
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SUBMITTER: Hoang CQ
PROVIDER: S-EPMC8278403 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Purpose:Retinoblastoma (Rb) is a rare and unique eye cancer that usually develops in the retinas of children less than 5 years old due to mutations in the RB1 gene. About 40% of affected individuals have the heritable form making genetics testing of the RB1 gene important for disease management. This study aims to identify germline mutations in RB1 in a cohort of patients with Rb from northern Vietnam. Methods:Genomic DNA was extracted from peripheral blood of 34 patients with Rb (nine unilateral and 25 bilateral cases) and their available parents. Twenty-seven exons, flanking sequences, and the promoter region of RB1 gene were screened for mutations with direct PCR sequencing. Multiplex ligation-dependent probe amplification (MLPA) was applied for patients with negative sequencing results. In the mutation-positive patients, their available parental DNA was analyzed to determine the parental origin of the mutation. Results:Germline mutations in RB1 were identified in 25 (73.53%) of 34 patients (four unilateral and 21 bilateral cases). Of these mutations, 19 were detected, including seven nonsense, six frameshift, four splice-site (one was identified in two siblings), and one missense, with Sanger sequencing. Three novel frameshift mutations were discovered in one unilateral and two bilateral patients. MLPA detected mutations in the RB1 gene in six bilateral cases, of whom five had a whole gene deletion (three familial cases) and one had a partial gene deletion (from exon 4 to exon 27) in one allele of the RB1 gene. Parental testing showed five mutations originated from the fathers and one was inherited from a mother who was mosaic for the mutation. Conclusions:This study provides a data set of germline mutations in the RB1 gene in Vietnamese patients with retinoblastoma. Screening of mutations in the RB1 gene can help to identify heritable Rb and contribute to clinical management and genetic counseling for affected families.
| S-EPMC5858657 | biostudies-literature
Project description:PurposeRetinoblastoma (RB) is a rare childhood malignant disorder caused by the biallelic inactivation of the RB1 gene. Early diagnosis and identification of carriers of heritable mutations in RB1 can improve disease outcome and management. In this study, we present the spectrum of mutations in the RB1 gene in Vietnamese patients with RB.MethodsTumor RNA from 50 probands with RB, including 12 bilateral and 38 unilateral cases, was extracted. cDNA, after reverse transcription, was sequenced to identify the RNA mutation of the RB1 gene. At the genomic DNA level, mutational analysis of all RB1 exons, exon-intron boundaries, and the promoter region was conducted using PCR and direct sequencing. Multiplex ligation-dependent probe amplification (MLPA) analysis was performed for patients for whom the first two results were negative. For patients for whom either the sequencing or MLPA results were positive for a tumor mutation, patients' and their parents' blood DNA was analyzed to determine the germline mutation.ResultsForty-one different kinds of RB1 tumor mutations were identified in 41 probands (82.0%), including 11 of 12 bilateral cases (91.7%) and 30 of 38 unilateral cases (78.9%). The majority of the detected mutations were nonsense (15 different kinds), followed by frameshift (11 kinds), and splice site mutations (nine kinds). Each splice site mutation was confirmed to create a deletion of the corresponding exon with RNA sequencing. The single promoter mutation c.-197G>A was reported previously; however, both missense mutations identified in exon 6 (c.601G>C: p.A201P) and exon 22 (c.2264T>C: p.F755S) were novel. Gross deletions were detected with MLPA in three probands. The detection rate of germline mutations in bilateral and unilateral cases with mutations were 81.8% and 30.0%, respectively. Only one father out of the 20 parents tested was positive for a germline mutation.ConclusionsMutations in the RB1 gene in Vietnamese patients were heterogeneous and highly prevalent with pathogenic truncated mutations. With advancement in therapeutics, early detection of RB is important for eye salvation.
| S-EPMC6450663 | biostudies-literature
Project description:The aim of the study was to investigate the frequency and types of mutations on the retinoblastoma gene (RB1 gene) in Turkish population. RB1 gene mutation analysis was performed in a total of 219 individuals (122 probands with retinoblastoma, 14 family members with retinoblastoma and 83 clinically healthy family members). All 27 exons and close intronic regions of the RB1 gene were sequenced for small deletions and insertions using both the Sanger sequencing or NGS methods, and the large deletions and duplications were investigated using the MLPA analysis and CNV algorithm. The bilateral/trilateral retinoblastoma rate was 66% in the study population. The general frequency of RB1 gene mutation in the germline of the patients with retinoblastoma was 41.9%. Approximately 51.5% of the patients were diagnosed earlier than 12 months old, and de novo mutation was found in 32.4% of the patients. Germline small genetic rearrangement mutations were detected in 78.9% of patients and LGRs were detected in 21.1% of patients. An association was detected between the eye color of the RB patients and RB1 mutations. 8 of the mutations detected in the RB1 gene were novel in the study.
| S-EPMC10489529 | biostudies-literature
Project description:PURPOSE:To find correlation between the type of mutations observed and the severity of the disease using multiple techniques like polymerase chain reactions (PCR), quantitative multiplex PCR, sequencing and RNA analysis. METHODS:Prospective, observational study. Patients who had been screened for mutations in the RB1 gene were included in the study. Patient details including demographic data; age and sex, laterality, international classification of intraocular retinoblastoma (ICIOR) staging, modality of management, histopathology high risk factors if the eyes were enucleated and metastasis rate were assessed. RESULTS:Seventy four patients were studied. Fifty three patients had bilateral and 21 unilateral disease. Complete genetic data was analyzed for 74 patients and complete clinical correlation was established for all the 49 patients with mutations. Of the total mutations identified, 11/49 (22.4%) of patients had large deletions, 12/49 (24.5%) had small deletions or insertions, 14/49 (28.6%) had nonsense mutations, 7/49 (14.3%) had splice mutations and 5/49 (10.2%) of patients had missense mutations. Four cases were familial. Group E ICIOR stage at presentation was noted in 40% of patients with large deletions, 33% with small deletions whereas 38.5% with splice mutations and 44.4% of patients with missense mutations presented with Group B ICIOR. Twenty five percentages of eyes with large deletions had high risk features on histopathology and one patient among these developed metastasis. CONCLUSION:Current laboratory testing of RB1 mutations may be feasible in determining the severity of the disease and patient counseling. The study provides a starting point for looking at correlations.
| S-EPMC3729507 | biostudies-literature
Project description:Retinoblastoma (RB) is an inherited childhood ocular cancer caused by mutations in the tumor suppressor RB1 gene. Identification of RB1 mutations is essential to assess the risk of developing retinoblastoma in the patients´ relatives. Retinoblastoma is a potentially curable cancer and an early diagnosis is critical for survival and eye preservation. Unilateral retinoblastoma is mostly non-heritable and results from two somatic mutations whereas bilateral retinoblastoma is heritable and results from one germline and one somatic mutation, both have high penetrance, 90%. The purpose of this study was to identify causative RB1 mutations in RB patients with different clinical presentations. A comprehensive approach was used to study a cohort of 34 patients with unilateral, bilateral and trilateral retinoblastoma. Blood and tumor DNA was analyzed by sequencing and multiplex ligation-dependent probe amplification (MLPA) assay. Validation of an insertion mutation was performed by cloning the PCR product. Most of the patients in our cohort had unilateral RB, eight patients had bilateral RB and one patient had a trilateral tumor with ocular and suprasellar/sellar locations. Other tumors in addition to retinoblastoma were also found in the affected families. One patient had two syndromes, retinoblastoma and schwannomatosis, and another RB patient had a father with a retinoma. Five out of the 25 unilateral RB patients carried germinal mutations (20%), which were mostly missense mutations. The bilateral and trilateral patients carried splice-site, nonsense and frameshift mutations as well as a whole RB1 gene deletion. Missense mutations were associated with mild phenotype: unilateral retinoblastoma, retinoma or no tumor. In this study we identified causative RB1 mutations in most bilateral RB patients and in some unilateral RB patients, including five novel mutations. These data are crucial for genetic counseling and confirm the need to perform complete genetic screening for RB1 mutations in both constitutional and tumor tissues.
| S-EPMC5738096 | biostudies-literature
Project description:Purpose:Retinoblastoma (RB) is a retinal tumor that most commonly occurs in children. Approximately 40% of RB patients carry germline mutations in the RB1 gene. RB survivors with germline mutations are at increased risk of passing on the disease to future offspring and of secondary cancer in adulthood. This highlights the importance of genetic testing in disease management and counseling. This study aimed to identify germline RB1 mutations and to correlate the mutations with clinical phenotypes of RB patients. Methods:Genomic DNA was extracted from peripheral blood mononuclear cells isolated from 52 RB patients (27 unilaterally and 25 bilaterally affected probands). Mutations in the RB1 gene, including the promoter and exons 1-27 with flanking intronic sequences, were identified by direct sequencing. The samples with negative test results were subjected to multiplex ligation-dependent probe amplification (MLPA) to detect any gross mutations. A correlation of germline RB1 mutations with tumor laterality or age at diagnosis was determined for RB patients. Age at diagnosis was examined in regard to genetic test results and the presence of extraocular tumor extension. Results:Germline RB1 mutations were detected in 60% (31/52) of patients. RB1 mutations were identified in 92% (22/25) of bilateral RB patients, and a high rate of germline RB1 mutations was found in unilateral RB cases (33% or 9/27). Whole gene and exon deletions were reported in five patients. Twenty-three distinct mutations as a result of base substitutions and small deletions were identified in 26 patients; seven mutations were novel. Nonsense and splicing mutations were commonly identified in RB patients. Furthermore, a synonymous mutation was detected in a patient with familial RB; affected mutation carriers in this family exhibited differences in disease severity. The types of germline RB1 mutations were not associated with age at diagnosis or laterality. In addition, patients with positive and negative test results for germline RB1 mutations were similar in age at diagnosis. The incidence of extraocular tumors was high in patients with heritable RB (83% or 5/6), particularly in unilateral cases (33% or 3/9); the mean age at diagnosis of these patients was not different from that of patients with intraocular tumors. Conclusions:This study provides a data set of an RB1 genotypic spectrum of germline mutations and clinical phenotypes and reports the distribution of disease-associated germline mutations in Thai RB patients who attended our center. Our data and the detection methods could assist in identifying a patient with heritable RB, establishing management plans, and informing proper counseling for patients and their families.
| S-EPMC6300611 | biostudies-literature
Project description:PurposeTo identify the spectrum of RB1 gene mutations in 114 Chinese patients with retinoblastoma.MethodsGenomic DNA was extracted from the peripheral blood of 114 Rb patients. Polymerase chain reactions (PCRs) followed by direct Sanger sequencing were used to screen for mutations in the RB1 gene, which contains 26 exons with flanking intronic sequences, except exon 15. Clinical data, including gender, age at diagnosis, laterality of ocular lesions, and associated symptoms, were recorded and compared.ResultsWe identified five novel mutations in the RB1 gene. Twenty-five other mutations found in this study have been previously reported. A higher rate of RB1 mutations, with 47.3% of mutations among bilaterally affected patients vs. 6.8% within unilaterally affected patients, was also observed (p < 0.0001). Bilaterally affected patients were diagnosed earlier when compared to unilaterally affected patients (11 ± 7 months versus 20 ± 14 months, p = 0.0002). Furthermore, nonsense mutations were abundant (n = 14), followed by frameshift mutations (n = 8), splicing site mutations (n = 5), while missense mutations were few (n = 3).ConclusionsWe found five novel mutations in RB1 genes, which expands the mutational spectrum of the gene. Children with bilateral Rb exhibited higher mutation rates and were diagnosed earlier than those with unilateral Rb. These findings will inform clinical diagnosis and genetic therapeutic targeting in Rb patients.
| S-EPMC9587959 | biostudies-literature
Project description:Survivors of hereditary retinoblastoma have a high risk of second primary malignancies, but it has not been investigated whether specific RB1 germline mutations are associated with greater risk of second primary malignancies in a large cohort. We conducted a retrospective cohort study of 199 survivors of hereditary retinoblastoma with a documented RB1 germline mutation diagnosed between 1905 and 2005. In total, 44 hereditary retinoblastoma survivors developed a second primary malignancy after a median follow-up of 30.2 years (range 1.33-76.0). A significantly increased risk of second primary malignancy was observed among carriers of one of the 11 recurrent CGA>TGA nonsense RB1 mutations (hazard ratio (HR) = 3.53; [95% confidence interval (CI) = 1.82-6.84]; P = .000), and there was a significantly lower risk for subjects with a low penetrance mutation (HR = .19; [95% CI = .05-.81]; P = .025). Our findings suggest a genotype-phenotype correlation for second primary cancers of retinoblastoma survivors and may impact on long-term surveillance protocols of patients with hereditary retinoblastoma, if confirmed by future studies.
| S-EPMC3365233 | biostudies-literature
Project description:Retinoblastoma (Rb) is a primary intraocular malignant tumor that occurs primarily in children, and results from loss-of-function mutations in the RB transcriptional corepressor 1 (RB1) gene. Genetic testing forms the basis of genetic counseling for affected families, as well as for clinical management of this disease. The aim of this study was to identify germline RB1 mutations and correlate the identified mutations with the clinical features of Rb patients. Genomic DNA was isolated from peripheral blood of 180 unrelated Rb patients and their parents (118 unilaterally and 62 bilaterally affected probands). Mutations in the RB1 gene, including the promoter region and exons 1-27 with flanking intronic sequences, were identified by Sanger sequencing. The samples with negative sequencing results were further subjected to methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) to detect gross deletions or duplications. Sixty-three distinct mutations were identified in 75 of the 180 (41.7%) probands. Of the 75 patients carrying RB1 mutations, 56 developed bilateral Rb, while 19 developed unilateral Rb. The total detection rates for bilateral and unilateral Rb were 90.3% (56/62) and 16.1% (19/118), respectively. Among the 75 patients, the spectrum of mutation types comprised 29.3% (22/75) nonsense mutations, 22.7% (17/75) splicing mutations, 17.3% (13/75) small insertions/deletions, 16.0% (12/75) large deletions/duplications, and 13.3% (10/75) missense mutations, while only 1% (1/75) of the mutations were in the promoter region of the RB1 gene. Age at diagnosis was significantly different (p < 0.01) between patients with positive and negative test results for germline RB1 mutations. A c.2359C > T mutation (p.R787X) was identified in identical twins, but one child was affected bilaterally and the other unilaterally. Of the five patients with deletion of the entire RB1 gene, the deletion of two patients was inherited from unaffected parents. In conclusion, in this study, we provide a comprehensive spectrum of RB1 germline mutations in Chinese Rb patients, and describe the correlations among RB1 mutations, age at diagnosis, and laterality; moreover, we report that the clinical features of individuals carrying an identical mutation in the RB1 gene were highly variable, indicating that the pathogenesis of Rb is more complicated than currently believed.
| S-EPMC7080181 | biostudies-literature
Project description:Retinoblastoma (RB) is a rare childhood malignant disorder caused by the biallelic inactivation of RB1 gene. Early diagnosis and identification of carriers of heritable RB1 mutations can improve disease outcome and management. In this study, mutational analysis was conducted on fifty-nine matched tumor and peripheral blood samples from 18 bilateral and 41 unilateral unrelated RB cases by a combinatorial approach of Multiplex Ligation-dependent Probe Amplification (MLPA) assay, deletion screening, direct sequencing, copy number gene dosage analysis and methylation assays. Screening of both blood and tumor samples yielded a mutation detection rate of 94.9% (56/59) while only 42.4% (25/59) of mutations were detected if blood samples alone were analyzed. Biallelic mutations were observed in 43/59 (72.9%) of tumors screened. There were 3 cases (5.1%) in which no mutations could be detected and germline mutations were detected in 19.5% (8/41) of unilateral cases. A total of 61 point mutations were identified, of which 10 were novel. There was a high incidence of previously reported recurrent mutations, occurring at 38.98% (23/59) of all cases. Of interest were three cases of mosaic RB1 mutations detected in the blood from patients with unilateral retinoblastoma. Additionally, two germline mutations previously reported to be associated with low-penetrance phenotypes: missense-c.1981C>T and splice variant-c.607+1G>T, were observed in a bilateral and a unilateral proband, respectively. These findings have implications for genetic counselling and risk prediction for the affected families. This is the first published report on the spectrum of mutations in RB patients from Singapore and shows that further improved mutation screening strategies are required in order to provide a definitive molecular diagnosis for every case of RB. Our findings also underscore the importance of genetic testing in supporting individualized disease management plans for patients and asymptomatic family members carrying low-penetrance, germline mosaicism or heritable unilateral mutational phenotypes.
| S-EPMC5456385 | biostudies-literature