Double Heterozygous Mutations Research Articles

Patients with Non-Transfusion-Dependent Thalassemia May Evolve to Transfusion-Dependent Thalassemia in Adulthood

Introduction: Non-transfusion-dependent thalassemia (NTDT) describes a prevalent and clinically heterogeneous group of patients in the United States. Adults with significant forms of NTDT display progressive age-related complications due to chronic anemia and ineffective erythropoiesis. While the importance of early diagnosis and close monitoring in pediatric patients with NTDT is well-established, the characteristics and complications of adults with NTDT in the United States is poorly described. Many adult patients with NTDT are managed by community providers who often have limited experience with this condition.This single-center observational study describes the clinical characteristics of patients with NTDT during their initial presentation to an adult thalassemia treatment center and identifies intervenable complications related to NTDT. Methods: We evaluated patients referred to the Penn Comprehensive Adult Thalassemia Program between 2013 and 2023 who carried a diagnosis of NTDT (defined as alpha or beta thalassemia and requiring fewer than 8 units of red cells in the 12 months preceding their initial visit). Data was collected via manual chart review. Clinical complications were defined as symptoms at presentation, history of splenectomy, hepatosplenomegaly, paraspinal masses, pulmonary hypertension, cardiac arrhythmia, endocrine complications (hypothyroidism, hypogonadism, diabetes, decreased bone mineral density), thrombotic events, and iron overload requiring chelation. Laboratory markers of disease severity included hemoglobin (Hgb), markers of hemolysis, nucleated red blood cells, serum ferritin and MRI quantitation of liver and cardiac iron. Results: Of the 94 patients with thalassemia referred to the Penn Comprehensive Adult Thalassemia Program, 34 carried a diagnosis of NTDT. Of these, 13 had α- thalassemia (deletional and non-deletional HbH), 19 had β-thalassemia variants (homozygous or double heterozygous β-globin gene mutations), and 2 had hemoglobin E/β-thalassemia. The median age at referral was 37.5 years (range 20-74); 71% were female. In general, patients with β-thalassemia had a larger number of significant complications than those with α-thalassemia. Overall, 11 patients (32%, all with β-thalassemia) had a history of splenectomy, 21 (62%) had evidence of extramedullary hematopoiesis with hepatosplenomegaly, and 7 (21%) had paraspinal masses. Pulmonary hypertension was seen in 5 of 21 patients with β-thalassemia and hemoglobin E/β-thalassemia (24%) while 4 patients overall (11%) had a history of thrombosis. The most common endocrinopathy was decreased bone mineral density (32%). Mean Hgb was 9.1 g/dL, however, 53% had a Hgb < 9 g/dL. Seven patients had an increase in nucleated red blood cells (range 161-1522 %) as a marker of ineffective erythropoiesis. Serum ferritin exceeded international NTDT guidelines for tissue evaluation (> 800 ng/mL) in 32% of patients; the mean ferritin for all patients was 897 ng/mL (range 45-7601 ng/mL). A liver MRI was obtained in 56% of patients with mean iron content of 25 mg/g dry weight (range 1.5 - 41.2 mg/g dry weight). No patients had evidence of cardiac iron overload. Of the 34 patients referred for NTDT, 14 (41%) met criteria to initiate a chronic transfusion regimen to maintain a pre-transfusion Hgb > 9.5 g/dL per established guidelines. Criteria included fatigue (all patients), extramedullary hematopoiesis (32%), Hgb < 7 g/dL (8%), and pulmonary hypertension (15%). Several patients had more than one indication for chronic transfusion. Conclusions: This is the first report of patients with NTDT in the United States that highlights clinically significant complications that accumulate in adults. Our results demonstrate a previously unknown high prevalence of conditions that warrant close monitoring, screening and intervention. Clinically, 41% of adult NTDT patients referred to our comprehensive adult thalassemia program met criteria to initiate a chronic transfusion program, a number that is significantly higher than previously published rates outside of the USA. These findings indicate a critical need for thalassemia education of adult providers that emphasizes the importance of monitoring for early complications of NTDT, as well as criteria for initiating a chronic transfusion program to mitigate complications and provide meaningful impacts in clinical outcomes.

S3832 Double Heterozygous HFE Mutation With Iron Overload Presenting as Fatty Liver – A Rare Case Report

S3832 Double Heterozygous HFE Mutation With Iron Overload Presenting as Fatty Liver – A Rare Case Report

Double heterozygous pathogenic mutations in KIF3C and ZNF513 cause hereditary gingival fibromatosis

Hereditary gingival fibromatosis (HGF) is a rare inherited condition with fibromatoid hyperplasia of the gingival tissue that exhibits great genetic heterogeneity. Five distinct loci related to non-syndromic HGF have been identified; however, only two disease-causing genes, SOS1 and REST, inducing HGF have been identified at two loci, GINGF1 and GINGF5, respectively. Here, based on a family pedigree with 26 members, including nine patients with HGF, we identified double heterozygous pathogenic mutations in the ZNF513 (c.C748T, p.R250W) and KIF3C (c.G1229A, p.R410H) genes within the GINGF3 locus related to HGF. Functional studies demonstrated that the ZNF513 p.R250W and KIF3C p.R410H variants significantly increased the expression of ZNF513 and KIF3C in vitro and in vivo. ZNF513, a transcription factor, binds to KIF3C exon 1 and participates in the positive regulation of KIF3C expression in gingival fibroblasts. Furthermore, a knock-in mouse model confirmed that heterozygous or homozygous mutations within Zfp513 (p.R250W) or Kif3c (p.R412H) alone do not led to clear phenotypes with gingival fibromatosis, whereas the double mutations led to gingival hyperplasia phenotypes. In addition, we found that ZNF513 binds to the SOS1 promoter and plays an important positive role in regulating the expression of SOS1. Moreover, the KIF3C p.R410H mutation could activate the PI3K and KCNQ1 potassium channels. ZNF513 combined with KIF3C regulates gingival fibroblast proliferation, migration, and fibrosis response via the PI3K/AKT/mTOR and Ras/Raf/MEK/ERK pathways. In summary, these results demonstrate ZNF513 + KIF3C as an important genetic combination in HGF manifestation and suggest that ZNF513 mutation may be a major risk factor for HGF.

Outcomes after HSCT for mucolipidosis II (I-cell disease) caused by novel compound heterozygous GNPTAB mutations.

Mucolipidosis type II (MLII), or I-cell disease, is a rare lysosomal storage disease (LSD) caused by variants in the GNPTAB gene. MLII patients exhibit clinical phenotypes in the prenatal or neonatal stage, such as marked dysmorphic features, cardiac involvement, respiratory symptoms, dysostosis multiplex, severe growth abnormalities, and mental and motor developmental abnormalities. The median age at diagnosis for MLII is 0.7 years, the median survival is 5.0 years, and the median age at death is 1.8 years. No cure for MLII exists. Sanger sequencing of the GNPTAB gene identified the compound heterozygous mutations c.673C > T in exon 7 and c.1090C > T in exon 9, which were novel double heterozygous mutations first reported in China. For the first time, we describe our experience in the use of HSCT for MLII. Our patient underwent HSCT with cells from a 9/10 human leukocyte antigen (HLA)-matched unrelated donor at 12 months of age. Myeloid neutrophil and platelet engraftment occurred on Days 10 and 11, respectively. The patient's limb muscle tension was significantly reduced, and his gross and fine motor skills were improved four months after transplantation. DST(Developmental Screen Test) results showed that the patient's fine motor skills and mental development were improved compared with before HSCT. MLII is a very severe lysosomal storage disease, to date, only 3 cases have been reported on the use of HSCT to treat MLII. Our data show that HSCT is a potential way to prolong the life of patients and improve their quality of life. Due to the lack of comparable data and time, the exact benefit remains unclear in MLII patients. Longer-term follow-up and in-depth prospective studies are indispensable.

Analysis of an Inherited FXII Deficiency Pedigree Associated with Double Heterozygous Mutations in the F12 Gene.

This article is intended to identify the potential mutations of the FXII gene (F12) in an inherited FXII deficiency pedigree and illuminate the pathogenesis of the disease. The coagulation FXII activity (FXII:C) and FXII antigen (FXII:Ag) were inspected by one-stage clotting assay and enzyme-linked immunosorbent assay(ELISA), respectively. Polymerase chain reaction amplification (PCR) was performed and the F12 gene was sequenced directly. A molecular model of FXIIprotein was established for further analysis. ClustalX-2.1-win and online bioinformatic software were used to estimate the conservatism and possible impact of the protein change. The proband presented prolonged APTT(180s) and extreme low FXII:C and FXII:Ag (both < 1%, reference range:72-113%). A compound heterozygous were found by the direct sequencing of the F12 gene. One was a deletion mutation c.1792_1796delGTCTA, which is a novel mutation; the other was an insertion mutation, c.1092_1093insC. Bioinformatic and modeling analyses indicated that the the two frameshift mutations may be deleterious and possibly alter the structure and the function of the protein. The mutations c.1792_1796delGTCTA and c.1092_1093insC could be the main causes of reducing FXII in this pedigree, and c.1792_1796delGTCTA mutation was the first report in the world.

Atypical Overlapping Features Complicating Diagnoses of Dysferlinopathy and Immune-Mediated Necrotizing Myopathy. (P6-8.006)

<h3>Objective:</h3> To present overlapping features and atypical characteristics of dysferlinopathy and immune-mediated necrotizing myopathy (IMNM). <h3>Background:</h3> Dysferlinopathy typically presents with early onset progressive muscle weakness that can be proximal or distal. Definitive diagnosis is made with DYSF gene testing. Treatment is supportive. IMNM is an autoimmune inflammatory myopathy that affects older individuals and is often associated with occult malignancy. Aggressive immunotherapy improves outcomes. Muscle biopsy (MBx) reveals inflammatory infiltrates in both conditions. <h3>Design/Methods:</h3> N/A <h3>Results:</h3> <h3>Case 1:</h3> A 35-year-old man presented with subacute progressive limb weakness and elevated creatine phosphokinase (CPK) &gt;8,000 units/L. MBx showed necrotizing myopathy and sarcolemmal C5b-9 staining; dystrophy stains were omitted due to the patient’s age and subacute symptoms. Anti-signal recognition peptide (SRP) and anti-3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR) antibodies and malignancy screen were negative. The patient was initially diagnosed with seronegative IMNM and treated with steroids, methotrexate and IVIG without improvement. He was subsequently diagnosed with dysferlinopathy based on genetic testing showing double heterozygous pathogenic DYSF gene mutations. <h3>Case 2:</h3> An 18-year-old man presented with 3-years of progressive proximal limb weakness associated with Gower’s sign and elevated CPK &gt;10,000 units/L. Broad muscular dystrophy genetic testing was negative. MBx showed necrotizing myopathy and sarcolemmal C5b-9 staining suggestive of IMNM. Anti-SRP and anti-HMGCR antibodies, serum paraneoplastic panel, and malignancy screening with tumor markers, contrasted CT imaging, and scrotal ultrasound were negative. The patient was diagnosed with early-onset seronegative IMNM. CPK improved and symptoms stabilized with prednisone, methotrexate and IVIG therapy. <h3>Conclusions:</h3> Dysferlinopathy and IMNM can have overlapping clinical features and MBx findings. Heightened awareness of atypical features aids in timely diagnosis. Accurate diagnosis of dysferlinopathy based on genetic testing limits further invasive workup and the side effects of ineffective empiric therapy, whereas early diagnosis of IMNM allows for prompt immunotherapy which is shown to improve prognosis. <b>Disclosure:</b> Dr. Stout has nothing to disclose. Dr. Su has nothing to disclose.

Severe Microcytic Anemia Caused by Complex Hereditary Spherocytosis and X-Linked Sideroblastic Anemia with Mutations in SPTB and ALAS2 Genes

We report a case of severe anemia caused by complex hereditary spherocytosis (HS) and X-linked sideroblastic anemia (XLSA) with two mutations in the spectrin beta (SPTB) and 5-aminolevulinic acid synthase (ALAS2) genes. The proband was a 16-year-old male with severe jaundice and microcytic hypochromic anemia since his childhood. He had more severe anemia requiring erythrocyte transfusion, and had no response to vitamin B6 treatment. Next-generation sequencing (NGS) revealed double heterozygous mutations, one in exon 19 (c.3936G > A:p.W1312X) of the SPTB gene and another in exon 2 (c.37A > G:p.K13E) of the ALAS2 gene, and confirmed again by Sanger sequencing. The mutation of ALAS2 (c.37A > G) is inherited from his asymptomatic heterozygous mother, causing amino acid p.K13E, and the mutation has not yet been reported. The mutation of SPTB (c.3936G > A) is a nonsense mutation, leading to a premature termination codon in exon 19, and the mutation in the SPTB gene is not found in any of his relatives, which indicates a de novo monoallelic mutation. Conclusions: The double heterozygous mutations in the SPTB and ALAS2 genes lead to the joint occurrence of HS and XLSA in this patient, and are implicated in the more severe clinical phenotypes.

Genetic analysis of a Fanconi anemia case revealed the presence of FANCF mutation (exon 1;469>C-T) with implications to develop acute myeloid leukemia.

Fanconi anemia (FA) is a rare genetic disorder and one of the most common inherited forms of aplastic anemia. FA is an autosomal recessive or X-linked genetic disorder that is characterized by typical physical malformations and haematopoietic anomalies. In most cases of FA, patients harbor homozygous or double heterozygous mutations in the FANCA (60-65%), FANCC (10-15%), FANCG (~ 10%), FANCD2 (3-6%) or FANCF (2%) genes in different ethnic populations, which leads to inherited bone marrow failure (IBMF). Hence, it is important to screen such mutations in correlation with clinical manifestations of FA in various ethnic populations. An 11 year old female pediatric patient of an East India family was presented with febrile illness, having thrombocytopenia with positive dengue IgM (Immunoglobulin M) and treated as a case of dengue hemorrhagic fever at the initial stage of diagnosis. Chromosomal breakage study was performed based on the abnormal physical examination, which showed 100% breaks, triradials, and quadrilaterals in mitomycin (MMC)-induced peripheral blood lymphocyte culture. Importantly, conventional cytogenetic assay in most of the bone marrow cells revealed an additional gain in chromosome 3q+ [46,XX,add(3)(q25)] and terminal loss in chr8p- [46,XX,del(8)(p23)], which might have a prognostic relevance in the outcomes of the FA patient. The bone marrow aspiration and biopsy were repeated and the results showed acute leukemia with 39% blast cells. Whole-genome sequencing analysis of the patient confirmed the presence of (exon 1; 496 > C-T) non-sense mutation leading to a truncated FANCF protein attributed to a stop codon at the amino acid position 166. The study reported the presence of a homozygous C-T exon 1 mutation in FANCF gene in the female pediatric patient from Odisha, India associated with FA. Furthermore, both parents were found to be carriers of FANCF gene mutation, as this allele was found to be in heterozygous state upon genome sequencing. The pathogenicity of the agent was robustly supported by the clinical phenotype and biochemical observations, wherein the patient eventually developed acute myeloid leukemia. The findings of the study infer the importance of early detection of FA and the associated mutations, which might lead to the development of acute myeloid leukemia.

Phenotypic and genotypic analysis of patients with congenital factor VII deficiency in a multicenter study in Thailand

ObjectiveThe phenotypic and genotypic analysis of patients with congenital factor VII deficiency were retrospectively conducted. MethodsThe study included 26 patients defined as severe (n = 25) and moderate (n = 1) degree by FVII <10% and 10–20%, respectively. ResultsThe diagnosis of factor VII deficiency was based on an isolated prolonged prothrombin time with subsequent factor assay. The median age of first bleed was 7 days (IQR 2–11.2 days). Central nervous system and gastrointestinal tract bleeding were found among all patients with severe degree. Patients with FVII <1–3% exhibited serious bleeding during the first week to the first month of life while patients with FVII >3% did not exhibit serious spontaneous bleeding. The initial bleeding episodes were controlled by administering fresh frozen plasma (FFP) and switched to factor concentrates among a few patients upon definite diagnosis. Subsequent prophylaxis was provided to patients with initial severe bleeding manifestation using FFP (15 ml/kg) 2–3 times weekly or recombinant factor VIIa (90 μg/kg) twice weekly. Genotypic analysis revealed homozygous or double heterozygous mutations among all patients except one patient with heterozygous mutation combined with homozygous polymorphism at codon 413 of G to A substitution (AA) at exon 8 of the FVII gene. The FVII gene mutation was commonly found at IVS6+1G > T (38.3%), followed by p.K376 X (19.2%) and IVS2+2T > C (17.0%). The case fatality rate was 19.2% (5/26) among patients with severe degree. ConclusionEarly diagnosis and appropriate management of congenital factor VII deficiency is essential for favorable outcomes.

The Genetic Spectrum of Familial Hypertriglyceridemia in Oman.

Familial hypertriglyceridemia (F-HTG) is an autosomal disorder that causes severe elevation of serum triglyceride levels. It is caused by genetic alterations in LPL, APOC2, APOA5, LMF1, and GPIHBP1 genes. The mutation spectrum of F-HTG in Arabic populations is limited. Here, we report the genetic spectrum of six families of F-HTG of Arab ancestry in Oman. Methods: six Omani families affected with triglyceride levels >11.2 mmol/L were included in this study. Ampli-Seq sequencing of the selected gene panels was performed. Whole-exome sequencing and copy number variant analysis were also performed in cases with negative exome results. Three novel pathogenic missense variants in the LPL gene were identified, p.M328T, p.H229L, and p.S286G, along with a novel splice variant c.1322+15T > G. The LPL p.H229L variant existed in double heterozygous mutation with the APOA5 gene p.V153M variant. One family had a homozygous mutation in the LMF1 gene (c.G107A; p.G36D) and a heterozygous mutation in the LPL gene (c.G106A; p.D36N). All affected subjects did not have a serum deficiency of LPL protein. Genetic analysis in one family did not show any pathogenic variants even after whole-exome sequencing. These novel LPL and APOA5 mutations are not reported in other ethnic groups. This suggests that patients with F-HTG in Oman have a founder effect and are genetically unique. This warrants further analysis of patients of F-HTG in the Middle East for preventative and counseling purposes to limit the spread of the disease in a population of high consanguinity.

Thrombotic Thrombocytopenic Purpura (ADAMTS13 [a Disintegrin and Metalloproteinase With a Thrombospondin Type 1 Motif, Member 13] Deficiency) as Cause of Recurrent Multiterritory Ischemic Strokes.

Thrombotic Thrombocytopenic Purpura (ADAMTS13 [a Disintegrin and Metalloproteinase With a Thrombospondin Type 1 Motif, Member 13] Deficiency) as Cause of Recurrent Multiterritory Ischemic Strokes.

Characteristics and Clinical Implication of UGT1A1 Heterozygous Mutation in Tumor.

BackgroundThe literature recommends that reduced dosage of CPT-11 should be applied in patients with UGT1A1 homozygous mutations, but the impact of UGT1A1 heterozygous mutations on the adverse reactions of CPT-11 is still not fully clear.MethodsA total of 107 patients with UGT1A1 heterozygous mutation or wild-type, who were treated with CPT-11 from January 2018 to September 2021 in Peking University Third Hospital, were retrospectively enrolled. The adverse reaction spectra of patients with UGT1A1*6 and UGT1A1*28 mutations were analyzed. Adverse reactions were evaluated according to National Cancer Institute Common Terminology Criteria for Adverse Events (NCI-CTCAE) 5.0. The efficacy was evaluated according to Response Evaluation Criteria in Solid Tumors (RECIST) 1.1. The genotypes of UGT1A1*6 and UGT1A1*28 were detected by digital fluorescence molecular hybridization.ResultsThere were 43 patients with UGT1A1*6 heterozygous mutation, 26 patients with UGT1A1*28 heterozygous mutation, 8 patients with UGT1A1*6 and UGT1A1*28 double heterozygous mutations, 61 patients with heterozygous mutation at any gene locus of UGT1A1*6 and UGT1A1*28. Logistic regression analysis showed that the presence or absence of vomiting (P=0.013) and mucositis (P=0.005) was significantly correlated with heterozygous mutation of UGT1A1*28, and the severity of vomiting (P < 0.001) and neutropenia (P=0.021) were significantly correlated with heterozygous mutation of UGT1A1*6. In colorectal cancer, UGT1A1*6 was significantly correlated to diarrhea (P=0.005), and the other adverse reactions spectrum was similar to that of the whole patient cohort, and efficacy and prognosis were similar between patients with different genotypes and patients treated with reduced CPT-11 dosage or not.ConclusionIn clinical use, heterozygous mutations of UGT1A1*6 and UGT1A1*28 are related to the risk and severity of vomiting, diarrhea, neutropenia and mucositis in patients with Pan-tumor and colorectal cancer post CPT-11 therpy. In colorectal cancer, UGT1A1*6 is significantly related to diarrhea post CPT-11 use, efficacy and prognosis is not affected by various genotypes or CPT-11 dosage reduction.

TP53 mutation in newly diagnosed acute myeloid leukemia and myelodysplastic syndrome

ObjectivesTP53 mutation is found frequently in therapy related acute myeloid leukemia (AML)/ myelodysplastic syndrome (MDS), AML and MDS patients with monosomy or complex karyotype. However, the prevalence and treatment outcome in TP53 mutated AML/MDS patients in Asian population are scarce. We therefore conducted this study to analyze the prevalence and the treatment outcomes of TP53 mutation in AML and MDS-EB patients.MethodsPatients with newly diagnosed AML and MDS-EB were recruited, extraction of deoxyribonucleic acid from bone marrow samples were done and then performing TP53 mutation analysis, using MassArray® System (Agena Bioscience, CA, USA).ResultsA total of 132 AML/MDS patients were recruited, patients with de novo AML, secondary AML, MDS-EB1, MDS-EB2 and T-AML/MDS were seen in 66, 13, 9, 9 and 3%, respectively. TP53 mutation was found in 14 patients (10.6%), and prevalence of TP53 mutation in T-AML/MDS, secondary AML, de novo AML and MDS-EB patients were 50, 17.6, 9.2 and 8%, respectively. Three patients had double heterozygous TP53 mutation. Mutated TP53 was significantly detected in patients with monosomy and complex chromosome. Common TP53 mutation were R290C, T220C, A249S and V31I which V31I mutation was reported only in Taiwanese patients. Most variant allele frequency (VAF) of TP53 mutation in the study were greater than 40%. Three year-overall survival (OS) in the whole population was 22%, 3y-OS in AML and MDS-EB patients were 22 and 27%, respectively. The 1y-OS in patients with TP53-mutant AML/MDS were shorter than that in TP53 wild-type patients, 14% versus 50%, P = 0.001. In multivariate analysis, factors affecting OS in 132 AML/MDS patients was mutant TP53 (P = 0.023, HR = 1.20–7.02), whereas, WBC count> 100,000/μL (P = 0.004, HR = 1.32–4.16) and complex karyotype (P = 0.038, HR = 1.07–9.78) were associated with shorter OS in AML patients.DiscussionIn this study, the prevalence of TP53 mutation in de novo AML and MDS-EB patients were low but it had impact on survival. Patients with monosomy or complex karyotype had more frequent TP53 mutation.

Identification of BRCA2 Cis Double Heterozygous Breast Cancer Cases Using Whole Exome Sequencing: Phenotypic Expression and Impact on Personalized Oncology.

BRCA1 and BRCA2 are the most commonly mutated breast cancer susceptibility genes that convey a high risk of breast and ovarian cancer. Most BRCA1 or BRCA2 mutation carriers have inherited a single heterozygous mutation. In recent years, very rare cases with biallelic or trans double heterozygous mutations on BRCA1 and or BRCA2 have been identified and seem to be associated with distinctive phenotypes. Given that this genotype-phenotype correlation in cancer predisposing hereditary conditions is of relevance for oncological prevention and genetic testing, it is important to investigate these rare BRCA genotypes for better clinical management of BRCA mutation carriers. Here we present the first report on Cis double heterozygosity (Cis DH) on BRCA2 gene identified using Whole exome sequencing (WES) in a Tunisian family with two BRCA2 mutations namely: c.632-1G>A and c.1310_1313DelAAGA that are both reported as pathogenic in ClinVar database. Subsequent analysis in 300 high-risk Tunisian breast cancer families detected this Cis double heterozygous genotype in 8 additional individuals belonging to 5 families from the same geographic origin suggesting a founder effect. Moreover, the observed Cis DH seems to be associated with an early age of onset (mean age = 35.33 years) and severe phenotype of the disease with high breast cancer grade and multiple cancer cases in the family. The identification of unusual BRCA genotypes in this Tunisian cohort highlights the importance of performing genetic studies in under-investigated populations. This will also potentially help avoiding erroneous classifications of genetic variants in African population and therefore avoiding clinical misdiagnosis of BRCA related cancers. Our findings will also have an impact on the genetic testing and the clinical management of North African breast cancer patients as well as patients from different other ethnic groups in regard to several emerging target therapies such as PARP inhibitors.

Frequency of mutations in PRKN, PINK1, and DJ1 in Patients With Early-Onset Parkinson Disease from neighboring countries in Central Europe

IntroductionApproximately 10% of patients with Parkinson disease (PD) present with early-onset disease (EOPD), defined as diagnosis before 50 years of age. Genetic factors are known to contribute to EOPD, with most commonly observed mutations in PRKN, PINK1, and DJ1 genes. The aim of our study was to analyze the frequency of PRKN, PINK1, and DJ1 mutations in an EOPD series from 4 neighboring European countries: Czech Republic, Germany, Poland, and Ukraine. MethodsDiagnosis of PD was made based on UK Brain Bank diagnostic criteria in departments experienced in movement disorders (1 from Czech Republic, 1 from Germany, 9 from Poland, and 3 from Ukraine). EOPD was defined as onset at or before 50 years of age. Of the 541 patients recruited to the study, 11 were Czech, 38 German, 476 Polish, and 16 Ukrainian. All cohorts were fully screened with Sanger sequencing for PRKN, PINK1, and DJ1 and multiplex ligation-dependent probe amplification for exon dosage. ResultsPRKN homozygous or double heterozygous mutations were identified in 17 patients: 1 Czech (9.1%), 1 German (2.6%), 14 Polish (2.9%), and 1 Ukrainian (6.3%). PINK1 homozygous mutations were only identified in 3 Polish patients (0.6%). There were no homozygous or compound heterozygous DJ1 mutations in analyzed subpopulations. One novel variant in PRKN was identified in the Ukrainian series. ConclusionIn the analyzed cohorts, mutations in the genes PRKN, PINK1, and DJ1 are not frequently observed.

Analysis of Gene Mutation Types in 141 Cases of α-Thalassemia in Honghe Prefecture, Yunnan Province

To detecte the carrying rate, the type and distribution of α-Thalassemia gene mutation in Honghe Prefecture, Yunnan Province, and analyze the differences in average erythrocyte volume (MCV), mean erythrocyte hemoglobin content (MCH) and hemoglobin among different types of α-Thalassemia. The DNA samples from small cell hypochromic carriers or anemia patients and women of childbearing age who underwent hematological screening in The First People's Hospital of Honghe State was from 2015 to 2019 were enrolled and analyzed, and the mutation types and frequency of alpha-thalassemia positive rate were diagnosed by PCR reverse dot blot or PCR fluorescence dissolution curve. Among the 1 016 samples, 141(13.88%) of the patients were diagnosed as α-thalassemia. The α-thalassemia was subdivided into 3 types, silent (36.17%), minor (51.77%), and HbH disease (12.06%), and the MCV, MCH and HB levels were detected and showed a obvious decrease trend with significant difference (P < 0.05). The gene mutation types were 9 kinds, the deletion type gene was mainly --SEA (51.06%), followed by -α3.7/αα deletion (29.79%), the α- mutation type gene was mainly αcsα(3.55%). The absence of complex heterozygote was most common, which was 17 cases, accounting for 12.77%, mainly --SEA/-α3.7 (10.64%). The areas were mainly distributed in Mengzi (47.52%), followed by Jinping (17.02%). Ethnic groups were mainly distributed in Han nationality (49.65%), followed by Zhuang (15.60%), Yi (3.48%) and Dai (7.09%). The patients with double heterozygous mutation was slightly higher in Mengzi than that in Jinping County (4.26% vs 3.55%), the Dai (2.13%) showed Zhuang ethnic group (2.13%) had the relatively high incidence except the Hans, and showed the most serious anemia. Alpha-thalassemia in Honghe prefecture of Yunnan Province shows complex genetic diversity and significant genetic heterogeneity, and the mainly type of gene mutation is --SEA and --SEA/-α3.7, which is mainly distributed in Han, Zhuang and Dai ethnic groups in Mengzi, Jinping. The anemia index of HbH group is the most obvious, and it is significantly different from other groups.

Abstract PS16-12: Double heterozygous pathogenic variants prevalence in a cohort of hereditary breast cancer patients

Abstract Significant progress has been made in the identification of inherited genetic factors underlying hereditary cancers. Inherited pathogenic variants in cancer predisposition genes that confer moderate to high risk of breast cancer (BC) are found in 10% of BC cases. Pathogenic variants in BRCA1 and BRCA2 genes are the underlying alterations in hereditary BC. Women with a pathogenic variant in BRCA1 or BRCA2 are at substantially higher risk of developing breast, ovarian and pancreatic cancers and represent 25-28% of hereditary BC cases. The gold standard for molecular diagnosis of a hereditary cancer syndrome is aNext Generation Sequencing (NGS) panel that includes all genes known to confer moderate to high-risk for BC (TP53, PTEN, CHEK2, ATM, PALB2 STK11, RAD51C and BRIP1). Identifying who is eligible to perform genetic testing remains a challenge but presently age at the time of tumor diagnosis, family history and disease stage are still the best predictors. The identification of a germline pathogenic variant enables appropriate genetic counseling, risk-reducing interventions. The detection of variants in BRCA1, BRCA2 and PALB2 may also determine treatment options in patients with advanced or metastatic disease. The known non-BRCA1/2-associated hereditary BC comprise a heterogeneous group of tumors, for most of which the prevalence of histologic and immunohistochemical (IHQ) phenotypes are yet to be identified while triple-negative BC is the most prevalent in BRCA1 carriers. The co-occurence of more than one germline pathogenic variant in BC genes is a rare event and it may affect cancer risk and penetrance is still unknown. Only a few reports on cases and series of double mutation carriers have been described in BRCA1, BRCA2, PALB2, CHEK2, BLM or NBN. In this series case we report patients with double heterozygous (DH) mutation in BC, their age at diagnosis, presence of second neoplasia and cancer phenotype in moderate and high penetrance genes on germline panels. Considering this scenario, we performed an active search among the 2651 women from the Hereditary Cancer Registry at Hospital Sirio-Libanes, São Paulo, Brazil, between January 2013 and June 2020. We have included 229 patients with a histopathological confirmation of a BC who have tested positive for pathogenic/likely pathogenic variant. Of these, 4,3% (10/229) women were identified with DH mutation in high and moderate penetrance genes in BC (table 1). All patients carried a germline pathogenic variant in at least one high-penetrance gene. A total of 7 cases (7/10) carried at least one BRCA1/2 pathogenic variant. Two women (2/10) included had developed bilateral BC. The occurrence of a second primary cancer was described in three patients (3/10). The median age at first BC diagnosis was 36,7 years old. Two BC were found to be triple negative, five cases were of Luminal subtype and one was HER2+. The age at diagnosis is similar to most hereditary BC related to those developed by BRCA1 and TP53 germline variant carriers. In this case series there is no evidence of additive effect on the risk of tumor development. Larger series of DH mutation carriers are needed to estimate the risk in double heterozygous populations and to determine whether the presence of DH have a synergistic interaction or additive effect on cancer risk. Table1 - Breast Cancer double variantsBreast Cancer double VariantsGeneVariantAge of Breast Cancer onset IHQ statusBilateral BCOther prymary neoplasia (age)BRCA1c.5266dupC36 yoER- PR- HER2 +YesPancreas (40), Lung (53)TP53c.1010G&amp;gt;ABRCA1c.4165.4166delAG32yoN/ANoBreast (37), Skin (50)PALB2c.1240C&amp;gt;TBRCA1c.5266dupC33yoER - RP- HER2 -NoNoATMc.6729_6730delAABRCA1c.3748G&amp;gt;T33yoER- PR -HER2-NoNoRECQL4c.1568_1573delinsCCCCCBRCA2c.2516dupA49yoER+ PR+ HER2 -NoNoPMS2c.2182_2184delinsGBRCA2c.8960del32yoER+ PR+ HER2-NoNoATMc.901+1G&amp;gt;ABRCA1c.1687C&amp;gt;T38yoER+NoBreast Cancer (58)NBNc.1142delC PR +MUTYHc.536A&amp;gt;G Her2-TP53c.1010G&amp;gt;A35yoN/ANoNoMUTYHc.1187G&amp;gt;AATMc.4906C&amp;gt;T42yoER+NoNoTP53c.1010G&amp;gt;APR+HER2-PALB2c.2711G&amp;gt;A37yoER+yesNoCHEK2c.319+2T&amp;gt;APR+HER2-BRCA2c.8960del32yoER+ PR+ HER2-NoNo Citation Format: Thais Megid, Maria Isabel Achatz, Janina Pontes Pisani, Mariana Cartaxo. Double heterozygous pathogenic variants prevalence in a cohort of hereditary breast cancer patients [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS16-12.

Severely affected familial hypercholesterolaemia patients with double and triple heterozygous mutations of candidate genes

Background and Aims: Familial hypercholesterolemia (FH) is one of the most frequent diseases with monogenic inheritance caused by various mutations in the genes encoding the low-density lipoprotein (LDL) receptor (LDLR), apolipoprotein (Apo) B100, the proprotein convertase subtilisin/kexin type 9 (PCSK9) and signal-transducing adaptor family member 1 (STAP1). We present three cases of a severely affected FH probands with double and triple heterozygous mutations of these candidate genes.

Application Value of Capillary Electrophoresis in Screeningβ-Thalassemia of Children

to explore the value of capillary electrophoresis in screening β- thalassemia of children, and to establish the cutoff values of HbA2 and HbF in our laboratory. The data of hemoglobin capillary electrophoresis and genetic diagnosis of β- thalassemia from 886 examined children were retrospectively analyzed. The cutoff values of HbA2 and HbF were determined by ROC curve. The cutoff value of HbA2 screening minor β- thalassemia was 3.65%, the specificity was 0.996, and the sensitivity was 0.995. The cut-off value of HbF for screening minor β- thalassemia was 1.45%, specificity was 0.751 and sensitivity was 0.675. Thus, 1 case with codon5 (CCT→C) mutation, 1 case with SEA -HPFH β deletion, 1 case with - 28 (A→G) merger IVS-Ι-128 (T→G) double heterozygous mutations yet were found out, 1 case with 47 bp β gene missing has not yet been reported in literature. Capillary electrophoresis has more high sensitivity and specificity in the screening of β- thalassemia in children, especially for the detection of rare β- thalassemia.

Application Value of Capillary Electrophoresis in Screeningβ-Thalassemia of Children

to explore the value of capillary electrophoresis in screening β- thalassemia of children, and to establish the cutoff values of HbA2 and HbF in our laboratory. The data of hemoglobin capillary electrophoresis and genetic diagnosis of β- thalassemia from 886 examined children were retrospectively analyzed. The cutoff values of HbA2 and HbF were determined by ROC curve. The cutoff value of HbA2 screening minor β- thalassemia was 3.65%, the specificity was 0.996, and the sensitivity was 0.995. The cut-off value of HbF for screening minor β- thalassemia was 1.45%, specificity was 0.751 and sensitivity was 0.675. Thus, 1 case with codon5 (CCT→C) mutation, 1 case with SEA -HPFH β deletion, 1 case with - 28 (A→G) merger IVS-Ι-128 (T→G) double heterozygous mutations yet were found out, 1 case with 47 bp β gene missing has not yet been reported in literature. Capillary electrophoresis has more high sensitivity and specificity in the screening of β- thalassemia in children, especially for the detection of rare β- thalassemia.