Ki-O Lee

UNC13D is the predominant causative gene with recurrent splicing mutations in Korean patients with familial hemophagocytic lymphohistiocytosis

Background Familial hemophagocytic lymphohistiocytosis is a fatal disease characterized by immune dysregulation from defective function of cytotoxic lymphocytes. Three causative genes have been identified for this autosomal recessive disorder (PRF1, UNC13D, and STX11). We investigated the molecular genetics of familial hemophagocytic lymphohistiocytosis in Korea. Design and Methods Pediatric patients who fulfilled the HLH-2004 criteria were recruited from the Korean Registry for Histiocytosis. Molecular genetic studies were performed on the patients’ DNA samples by direct sequencing of all coding exons and flanking sequences of PRF1, UNC13D, and STX11. Results Forty patients were studied and familial hemophagocytic lymphohistiocytosis mutations were identified in nine; eight patients had UNC13D mutations (89%) and one had a mutation in PRF1. No patient had a STX11 mutation. Notably, four patients had only one UNC13D mutant allele, suggesting that the other mutation was missed by conventional direct sequencing. All UNC13D mutations were deleterious in nature. One known splicing mutation, c.754-1G>C, was recurrent, accounting for 58% of all the mutant alleles (7/12). Five UNC13D mutations were novel (p.Gln98X, p.Glu565SerfsX7, c.1993-2A>G, c.2367+1G>A, and c.2954+5G>A). The one patient with PRF1 mutation was homozygous for a frameshift mutation (p.Leu364GlufsX93), which was previously reported to be the most frequent PRF1 mutation in Japan. Conclusions This is the first investigation on the molecular genetics of familial hemophagocytic lymphohistiocytosis in Korea. The data showed that UNC13D is the predominant causative gene in the Korean population. The identification of mutations missed by conventional sequencing would better delineate the mutation spectrum and help to establish the optimal molecular diagnostic strategy for familial hemophagocytic lymphohistiocytosis in Korea, which might need an RNA-based screening strategy.

Capecitabine monotherapy in patients with anthracycline-and taxane-pretreated metastatic breast cancer

The selection of chemotherapeutic regimens is challenging for metastatic breast cancer (MBC) patients whose diseases have failed to respond to anthracyline and taxane. Capecitabine has advantages of oral administration and favorable toxicity profiles. This study was conducted to evaluate the efficacy of capecitabine and to identify the subgroup of patients who would potentially have benefit from capecitabine monotherapy in patients with anthracycline- and taxane-pretreated MBC. Female patients with MBC who had been previously treated with anthracycline and taxane received oral capecitabine 2500 mg/m2 divided in two doses daily for 2 wk with 1-wk rest period. Between September, 1999, and December, 2002, a total of 38 patients were enrolled. Among the 36 evaluable patients, one patient achieved a complete response (CR), 9 patients had partial responses (PRs), and 13 patients had stable diseases (SDs). Response rate was 26% [95% confidence interval (CI), 12–40%] and the tumor control rate (TCR, CR+PR+SD) was 61% (95% CI, 45–77%). The median follow-up duration was 27.8 mo. The median duration of response was 8.9 mo, the median time to progression was 4.6 mo, and the median overall survival was 18.1 mo. The major toxicities were hand-foot syndrome, diarrhea, and emesis. There was no treatment-related death. The predictors of better overall survival were positivity of hormone receptor, disease-free survival longer than 1 yr, non-refractoriness to anthracycline, and fewer number (≤3) of involved organs. Capecitabine monotherapy is effective and well tolerated for MBC patients who had previously been treated with anthracycline and taxane. The TCR could predict overall survival as well as the objective respose in this study, suggesting a possible role of TCR as a surrogate marker for survival in MBC patients on salvage chemotherapy. The patients who have relatively slow growing tumor and less tumor burden could have benefit from capecitabine monotherapy following anthracycline- and taxane-based chemotherapy.

Distinct frequencies and mutation spectrums of genetic thrombophilia in Korea in comparison with other Asian countries both in patients with thromboembolism and in the general population

Hereditary natural anticoagulant deficiencies are the major cause of genetic thrombophilia in Asia. Given the growing acknowledgment of the risk of venous thromboembolism in Asian populations, we investigated the frequency and mutation spectrums of natural anticoagulant deficiency in Korea. The group of patients consisted of consecutive patients with venous thromboembolism screened for thrombophilia. Genetic tests were performed on suspicion of natural anticoagulant deficiency. For the population group, >3,000 individuals were screened from routine check-ups, and those with a low level (<1st percentile) of natural anticoagulant underwent genetic tests. Mutations were detected by direct sequencing of PROC, PROS1, and SERPINC1, followed by additional multiplex ligation-dependent probe amplification for PROS1 and SERPINC1 for dosage mutations. Among 500 patients screened, 127 were suspected of having a natural anticoagulant deficiency, and this was genetically confirmed in 71: protein C deficiency in 36 (50.7%), antithrombin deficiency in 21 (29.6%), and protein S deficiency in 14 (19.7%). Among 3,129 individuals from the population who were screened, the frequency of natural anticoagulant deficiency was ~1.0%: antithrombin deficiency 0.49%, protein C deficiency 0.35%, and protein S deficiency 0.16%. Two PROC mutations causing type I protein C deficiency were prevalent (Arg211Trp and Met406Ile in patients and Arg211Trp in the population). Two SERPINC1 mutations causing type II antithrombin deficiency, Arg79Cys and Ser158Pro, were prevalent in the population group. This is the first study on the genetic epidemiology of natural anticoagulant deficiencies in Korea. The results demonstrated that the frequencies and spectrum of mutations underlying genetic thrombophilia in Korea are different not only from those in Caucasians but also those in other Asian populations.

A novel mutation Gly603Arg of TMPRSS6 in a Korean female with iron-refractory iron deficiency anemia†‡

Iron‐refractory iron deficiency anemia (IRIDA) is a rare hereditary form of IDA with autosomal recessive inheritance. IRIDA is characterized by hypochromic microcytic anemia unresponsive to oral iron treatment, low transferrin saturation, and a high level of iron‐regulated hormone hepcidin. The genetic background of IRIDA is mutations in the TMPRSS6 gene encoding matriptase‐2 (TMPRSS6) that prevent inactivation of hemojuvelin, an activator of hepcidin transcription. We herein report a Korean female with IRIDA who was compound heterozygous for two mutations in TMPRSS6: a novel missense mutation c.1807G>C (p.Gly603Arg) in the serine protease domain and a known splicing mutation c.863+1G>T (IVS6+1G>T). Pediatr Blood Cancer 2012; 58: 640–642.

A novel initiation codon mutation in the ribosomal protein S17 gene (RPS17) in a patient with Diamond‐Blackfan anemia

Diamond‐Blackfan anemia (DBA) is an inherited bone marrow failure syndrome characterized by pure red cell aplasia, various congenital anomalies, and cancer predisposition. We report a novel mutation in the RPS17 gene in a Korean patient with DBA. The mutation occurred in the translation initiation codon, changing Atg to Gtg (c.1A>G), thus disrupting the natural start of the RPS17 protein biosynthesis. This is the third case of DBA from a RPS17 mutation in the literature and is the second case of a RPS17 mutation in the translation initiation codon, following c.2T>G. Pediatr Blood Cancer 2010;54:629–631.

Heterogeneous lengths of copy number mutations in human coagulopathy revealed by genome-wide high-density SNP array

Background The recent advent of genome-wide molecular platforms has facilitated our understanding of the human genome and disease, particularly copy number aberrations. We performed genome-wide single nucleotide polymorphism-array in hereditary coagulopathy to delineate the extent of copy number mutations and to assess its diagnostic utility. Design and Methods The study subjects were 17 patients with hereditary coagulopathy from copy number mutations in coagulation genes detected by multiple ligation-dependent probe amplification. Eleven had hemophilia (7 hemophilia A and 4 hemophilia B) and 6 had thrombophilia (4 protein S deficiency and 2 antithrombin deficiency). Single nucleotide polymorphism-array experiments were performed using Affymetrix Genome-Wide Human SNP arrays 6.0. Results Copy number mutations were identified by single nucleotide polymorphism-array in 9 patients, which ranged in length from 51 Kb to 6,288 Kb harboring 2 to ~160 genes. Single nucleotide polymorphism-array showed a neutral copy number status in 8 patients including 7 with either a single-exon copy number mutation or duplication mutations of PROS1. Conclusions This study revealed unexpectedly heterogeneous lengths of copy number mutations underlying human coagulopathy. Single nucleotide polymorphism-array had limitations in detecting copy number mutations involving a single exon or those of a gene with homologous sequences such as a pseudogene.

Molecular genetic analysis of Korean patients with coagulation factor XII deficiency.

Coagulation factor XII (FXII) participates in the initiation of blood coagulation, fibrinolysis, complement systems, and bradykinin generation. Hereditary FXII deficiency is caused by mutations in the F12 gene. In this report, we describe three Korean patients with FXII deficiency. In all three patients, prolonged activated partial thromboplastin time (aPTT) was detected on preoperative coagulation screening, without a history of bleeding tendency. Patient 1 was a 4-year-old girl with congenital heart anomaly. Her aPTT was prolonged at 67.5 s (reference range, 29–42 s) and the FXII activity was 5% (48–177%). Patient 2 was a 50-year-old man with anal fistula. His aPTT was 95.2 s and FXII activity was less than 0.5% (below the detection limit). Patient 3 was a 53-year-old woman with pituitary macroadenoma. Her aPTT was 238.8 s and FXII activity was less than 0.5%. Direct sequencing analyses of F12 revealed two mutations in all three patients. Patient 1 was compound heterozygous for two missense mutations, c.1583C>T (p.S528F) and c.1744G>A (p.G582S). Patient 2 was homozygous for a nonsense mutation, c.405C>A (p.C135X). Patient 3 was compound heterozygous for a frameshift mutation and a missense mutation, c.1093_1094insC (p.K365QfsX69) and c.1744G>A (p.G582S). All four mutations were novel. To the best of our knowledge, these are the first genetically confirmed cases of FXII deficiency in Korea. FXII deficiency should be included in differential diagnoses of prolonged aPTT, particularly in those without bleeding tendency.

Sequence variation data of F8 and F9 genes in functionally validated control individuals: implications on the molecular diagnosis of hemophilia

Background The F8 and F9 genes encode for coagulation factor VIII (FVIII) and FIX, respectively, and mutations in these genes are the genetic basis of hemophilia A/B. To determine whether a sequence variation in F8/F9 is a disease-causing mutation, frequency data from a control population is needed. This study aimed to obtain data on sequence variation in F8/F9 in a set of functionally validated control chromosomes of Korean descent. Methods We re-sequenced F8 and F9 from DNA samples of 100 Korean male control individuals with normal PT, aPTT, and FVIII activity. PCR and direct sequencing analyses were performed using primer pairs to cover all coding regions and the flanking intronic sequences. Results Thirteen individuals (13%) were hemizygous for sequence variations in the coding region of F8. Six (6%) had c.3780C>G (p.Asp1260Glu), five (5%) had c.3864A>C (p.Ser1288=). One each individual (1%) had c.4794G>T (p.Glu1598Asp) and c.5069 A>G (p.Glu1690Gly). Asp1260Glu and Ser1288= were known SNPs (rs1800291 and rs1800292, respectively). Glu1598Asp was assigned as a missense mutation in public databases (HGMD and HAMSTeRS), and Glu1690Gly was a novel variation. Based on the normal FVIII activities in control individuals carrying these variations (109% and 148%, respectively), they were considered to be rare SNPs. No variation was observed in F9 of control individuals. Conclusion A significant proportion of control individuals carried sequence variations in F8, but not in F9. These results can be used as a reference dataset for molecular diagnosis of hemophilia A and B, particularly in Korea.

Combined congenital dysfibrinogenemia and factor VII deficiency from mutations in the FGB and F7 genes.

Dysfibrinogenemia and factor VII (FVII) deficiency are rare congenital coagulopathies. In this report, the authors describe a man with both defects confirmed by molecular genetic tests. The patient was a 51-year-old man referred for prolonged prothrombin time (PT) that had been accidentally detected on preoperative screening. He had no history of bleeding tendency even on occasions of surgery. Routine coagulation studies revealed prolonged PT (1.53 INR) and thrombin time (42.2 s), and decreased fibrinogen level (57 mg/dl) and FVII activity (44%). Direct sequencing analyses were performed on FGA, FGB, and FGG genes to confirm dysfibrinogenemia and on the F7 gene to confirm FVII deficiency. As a result, the patient was shown to be heterozygous for a point mutation in exon 8 of the FGB gene (c.1475A > G, p.*492Trpext*12; Fibrinogen Magdeburg II) and for a missense mutation in exon 6 of the F7 gene (c.466G > A, p.Gly156Ser). To our knowledge, this is the first report on a case of combined dysfibrinogenemia and FVII deficiency confirmed by molecular genetic tests.

A novel nonsense mutation in the MPL gene in congenital amegakaryocytic thrombocytopenia

Congenital amegakaryocytic thrombocytopenia (CAMT) is a rare autosomal recessive disorder characterized by thrombocytopenia from failure of megakaryopoiesis. CAMT is one of the bone marrow failure syndromes, and the disease progression may involve other lineages leading to pancytopenia. The genetic background of CAMT is mutations in the MPL gene encoding the thrombopoietin receptor. Here, we describe a Korean male with CAMT. Molecular genetic analyses by direct sequencing revealed that he was compound heterozygous for two nonsense mutations in MPL, Tyr63X (c.189C>A), and Arg357X (c.1069C>T), the latter being a novel mutation. Pediatr Blood Cancer 2011;56:??–??.