Genetic study of the CDKN2A and CDKN2B genes in renal cell carcinoma patients.

Objectives: While recent studies have demonstrated several genetic alterations are associated with pathogenesis of RCC, the significance of cyclin-dependent kinase inhibitor 2A (CDKN2A) and cyclin-dependent kinase inhibitor 2B (CDKN2B) in tumorigenesis of RCC is less clear. We investigate the distribution of CDKN2A and CDKN2B mutations in patients with RCC and analyze the impact of CDKN2A and CDKN2B mutations on RCC. Methods: A pathological examination was conducted using thirty fresh renal tissue samples with renal masses that had undergone partial or radical nephrectomy. Multiplex ligation-dependent probe amplification (MLPA) was used to detect genetic aberrations of CDKN2A and CDKN2B in genomic DNA isolated from samples. Subsequently, CDKN2A and CDKN2B mutations were confirmed using chromosomal microarray technique. Results: Twenty-one patients were diagnosed with RCC, eight with benign diseases, including angiomyolipoma (AML) and oncocytoma, and one with mucinous adenocarcinoma of renal pelvis. Two of twenty-one patients (9.5 %) with clear-cell RCC were positive for CDKN2A and CDKN2B gene deletions. Interestingly, patients with CDKN2A and CDKN2B mutations were associated with sarcomatoid patterns of RCC (2 out of 4, 50 %). In contrast, no CDKN2A or CDKN2B deletions were detected in samples from benign renal tumors, papillary RCC, or other kidney cancers. Conclusions: This study demonstrated the potential use of CDKN2A and CDKN2B as biomarkers for the prognostic and molecular classification of renal cancer. CDKN2A and CDKN2B mutations may be associated with RCC development and sarcomatoid changes. Further research is needed to understand the underlying molecular mechanisms of CDKN2A and CDKN2B in the pathogenesis of RCC.

Chromosomal abnormalities study for anembryonic pregnancy by BACs-on-Beads technique.

OBJECTIVE: This study evaluated the BACs-on-Beads™ (BoBs) efficiency assay in detecting chromosomal anomalies in products of conception (POC) specimens associated with anembryonic pregnancy (AP) among Thai pregnant women. METHOD: Retrospective analysis applied the BoBs™ assay to examine AP samples from 2010 to 2022. The incidences of AP with chromosomal abnormalities were reported. RESULT: Assessment of villi from anembryonic pregnancy samples found normal chromosome complement in 50% of the cases, while the remainder showed chromosomal abnormalities. Trisomy 16 was found in 15% of the cases and trisomies 22, 15, and 19 in 9.6%, 3.8%, and 3.8%, respectively. Advanced maternal age was associated with a higher incidence of aneuploidy. CONCLUSION: The BoBs™ assay effectively detected diverse chromosomal abnormalities in villi samples from POC. The diagnostic utility of the BoBs™ assay was highlighted in identifying chromosomal irregularities in AP cases. Trisomy 16 possessed the most chromosomal abnormalities in the AP samples.

Therapeutic potential of third-generation chimeric antigen receptor T cells targeting B cell maturation antigen for treating multiple myeloma.

Multiple myeloma (MM) is an incurable hematologic malignancy characterized by the rapid proliferation of malignant plasma cells within the bone marrow. Standard therapies often fail due to patient resistance. The US FDA has approved second-generation chimeric antigen receptor (CAR) T cells targeting B-cell maturation antigen (anti-BCMA-CAR2 T cells) for MM treatment. However, achieving enduring clinical responses remains a challenge in CAR T cell therapy. This study developed third-generation T cells with an anti-BCMA CAR (anti-BCMA-CAR3). The CAR incorporated a fully human scFv specific to BCMA, linked to the CD8 hinge region. The design included the CD28 transmembrane domain, two co-stimulatory domains (CD28 and 4-1BB), and the CD3ζ signaling domain (28BBζ). Lentiviral technology generated these modified T cells, which were compared against anti-BCMA-CAR2 T cells for efficacy against cancer. Anti-BCMA-CAR3 T cells exhibited significantly higher cytotoxic activity against BCMA-expressing cells (KMS-12-PE and NCI-H929) compared to anti-BCMA-CAR2 T cells. At an effector-to-target ratio of 10:1, anti-BCMA-CAR3 T cells induced lysis in 75.5 ± 3.8% of NCI-H929 cells, whereas anti-BCMA-CAR2 T cells achieved 56.7 ± 3.4% (p = 0.0023). Notably, after twelve days of cultivation, anti-BCMA-CAR3 T cells nearly eradicated BCMA-positive cells (4.1 ± 2.1%), while anti-BCMA-CAR2 T cells allowed 36.8 ± 20.1% to survive. This study highlights the superior efficacy of anti-BCMA-CAR3 T cells against both low and high BCMA-expressing MM cells, surpassing anti-BCMA-CAR2 T cells. These findings suggest potential for advancing anti-BCMA-CAR3 T cells in chimeric antigen receptor T (CAR-T) therapy for relapsed/refractory MM.

Committed change of real-time quantitative PCR to droplet digital PCR for monitoring BCR::ABL1 transcripts in tyrosine kinase inhibitor treated CML.

OBJECTIVES: We performed a feasibility study of an FDA-approved commercial ddPCR assay to measure BCR::ABL1 in CML patients treated using TKI therapy. METHODS: Assay performance of standard RQ-PCR and commercially available FDA-approved ddPCR were compared to measure BCR::ABL1 p210 transcripts in RNA samples from 100 CML patients who received TKI therapy. RESULTS: %BCR::ABL1/ABL1IS levels obtained from both methods were not statistically significant difference after normalization with batch-specific conversion factor (p = 0.0651). The correlation and agreement of %BCR::ABL1/ABL1IS between the two assays were high. Molecular response stratification data showed 56% concordance between RQ-PCR and ddPCR, and 37% higher residual disease detection using ddPCR. Furthermore, 21.21% (7/33) of RQ-PCR undetectable samples were detected by ddPCR, representing high sensitivity to quantify the low abundance of BCR::ABL1 transcripts. CONCLUSION: ddPCR was proven to be a highly sensitive method with the potential to overcome some limitations of traditional RQ-PCR, and has the potential of being a valuable tool for monitoring BCR::ABL1 transcripts in CML during TKI therapy. (163 words).

A customized mass array panel for BCR::ABL1 tyrosine kinase domain mutation screening in chronic myeloid leukemia.

Introduction: The therapeutic strategy and management of chronic myeloid leukemia (CML) have rapidly improved with the discovery of effective tyrosine kinase inhibitors (TKIs) to target BCR::ABL1 oncoprotein. However, nearly 30% of patients develop TKI resistance due to acquired mutations on the tyrosine kinase domain (TKD) of BCR::ABL1. Methods: We customized a mass array panel initially intended to detect and monitor the mutational burden of hotspot BCR::ABL1 TKD mutations accumulated in our database, including key mutations recently recommended by European LeukemiaNet. Additionally, we extended the feasibility of using the assay panel for the molecular classification of myeloproliferative neoplasms (MPNs) by incorporating primer sets specific for analyzing JAK2 V617F, MPL 515 K/L, and CALR types 1 and 2. Results: We found that the developed mass array panel was superior for detecting and monitoring clinically significant BCR::ABL1 TKD mutations, especially in cases with low mutational burden and harboring compound/polyclonal mutations, compared with direct sequencing. Moreover, our customized mass array panel detected common genetic alterations in MPNs, and the findings were consistent with those of other comparable assays available in our laboratory. Conclusions: Our customized mass array panel was practicably used as a routine robust assay for screening and monitoring BCR::ABL1 TKD mutations in patients with CML undergoing TKI treatment and feasible for analyzing common genetic mutations in MPNs.

The Utilization of MS-MLPA as the First-Line Test for the Diagnosis of Prader-Willi Syndrome in Thai Patients.

Prader-Willi syndrome (PWS) is a genetic disorder caused by the expression disruption of genes on the paternally inherited allele of chromosome 15q11.2-q13. Apart from clinical diagnostic criteria, PWS is confirmed by genetic testing. Methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) is one of the molecular techniques used to analyze this syndrome. This study aimed to evaluate the concordance of the test results of MS-MLPA with conventional techniques in the diagnosis of PWS in Thai patients. Forty leftover specimens from routine genetic testing (MS-PCR and FISH) were tested to obtain MS-MLPA results. By comparison, perfect concordance was shown between the result of MS-MLPA and those of conventional techniques. In conclusion, MS-MLPA is an accurate and cost-effective assay that can be used to confirm PWS diagnosis with explicit deletion of affected genes.

Incidence and Types of Fetal Chromosomal Abnormalities in First Trimester of Thai Pregnant Women between Miscarriages and Intrauterine Survivals.

Abortion is a common pregnancy complication. Fetuses with several types of chromosomal abnormalities are aborted during the first trimester, while others have a better chance of surviving. This research aims to study and compare the incidence and types of fetal chromosomal abnormalities during the first trimester of Thai pregnant women between miscarriages and intrauterine survivals. Cytogenetic and BACs-on-Beads™ assays were assessed from 2010 to 2020 in Ramathibodi Hospital using first trimester samples of 265 chorionic villi as a retrospective study. Chromosomal abnormalities were observed in 135 cases (50.94%) including 38.11% miscarriages and 12.83% intrauterine survivals. In total, 75.56% single autosomal trisomies, 18.52% sex chromosome aneuploidies, 5.19% double aneuploidies, and 0.74% structural abnormalities were detected. In miscarriages, all chromosomes were involved in abnormalities except chromosomes 1, 5, 8, 9, 11, and 17, while survivals had only trisomy 13, 18, 21, and sex chromosome aneuploidy. Trisomy 16 and 18 were the most common abnormalities in miscarriages and intrauterine survivals, respectively. The highest rate of chromosomal aberrations was demonstrated in 8-9+6 and 12-13+6 weeks of gestation in miscarriages and intrauterine survivals, respectively. Correlation between chromosomal abnormalities and maternal age <35 years and ≥35 years was significant (p < 0.05) in intrauterine survival and first trimester groups.

TP53 mutation in newly diagnosed acute myeloid leukemia and myelodysplastic syndrome.

OBJECTIVES: TP53 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. METHODS: Patients 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). RESULTS: A 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. DISCUSSION: In 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.

Targeting UCHL1 Induces Cell Cycle Arrest in High-Risk Multiple Myeloma with t(4;14).

Multiple myeloma (MM) patients considered to be at high cytogenetic risk commonly fail to respond to standard treatment. A thorough understanding of the molecular mechanism of MM development is, therefore, needed. We endeavored to explore the transcriptional signature among different subgroups of newly diagnosed MM using gene chip-based expression microarray. Bone marrow samples of 15 newly diagnosed Thai MM patients were included. The chromosomal translocation t(4;14) was the most frequently identified genetic alteration in the high-risk subgroup. Cluster analysis from expression profiling demonstrated that high-risk MM have a distinctly different expression pattern compared to standard-risk patients. The most significant differentially expressed gene was UCHL1. Functional enrichment analysis by Gene Set Enrichment Analysis, FUNRICH, and Gene Ontology Panther pathway revealed the gene sets involved in cell cycle control to be enriched in the t(4;14) high-risk group. Interestingly, among the well-established downstream targets of UCHL1, only CCND2 was significantly expressed in the t(4;14) high-risk group. Suppression of UCHL1 protein level by LDN-5744 inhibitor could arrest the cell cycle in G1 phase in cell lines. These findings shed light on the molecular mechanism of UCHL1 in t(4;14) high-risk MM and support the evidence that alteration of the UCHL1 pathway may play a role in the pathogenesis of high-risk MM.

High Expression of miR-483-5p Predicts Chemotherapy Resistance in Epithelial Ovarian Cancer.

BACKGROUND: Ovarian cancer is the most deadly cancer that requires novel diagnostics and therapeutics. MicroRNAs are viewed as essential gene regulatory elements involved in different pathobiological mechanisms of many cancers, including ovarian cancer. OBJECTIVE: This study examined the relationship between microRNA (miRNA) expression and response to platinum-based chemotherapy. METHODS: Genome-wide miRNA expression analysis was conducted using Epithelial Ovarian Cancer (EOC) tissues from 25 patients with 17 malignant tumors and eight benign ovarian tumors. Candidate miRNAs that respond to platinum-based chemotherapy were selected for validation by quantitative RT-PCR. RESULTS: Among 2,578 mature human miRNAs, high expression of miR-483-5p correlated with poor responses to platinum-based chemotherapy in EOC patients. Furthermore, high levels of miR-483-5p in the resistant group suppressed expression of the apoptotic regulator TAOK-1. CONCLUSION: A possible marker for the prediction of chemotherapy response and resistance in patients may be miR-483-5p. Choosing the right treatment for each patient with EOC can avoid the risk of developing chemotherapy resistance.

Practical Laboratory Tools for Monitoring of BCR-ABL1 Transcripts and Tyrosine Kinase (TK) Domain Mutations in Chronic Myeloid Leukemia Patients Undergoing TK Inhibitor Therapy: A Single-Center Experience in Thailand.

OBJECTIVE: The genetic hallmark of CML is known as the appearance of t(9;22)(q34.1;q11.2) (BCR-ABL1) which is present in more than 95% of cases. Here, we demonstrated practical laboratory tools for monitoring of BCR-ABL1 transcripts in chronic myeloid leukemia patients undergoing TK inhibitor therapy. METHODS: Real time quantitative PCR and direct sequencing were performed for monitoring of BCR-ABL1 transcripts in 245 treated CML. RESULTS: At month 3 after first time point of monitoring, we found that 89% (218/245), 2% (5/245), and 9% (22/245) of patients are determined as optimal, warning, and failure response, respectively. The responses to TKI were slightly decreased at months 6 as following 73% optimal (180/245), 18% warning (43/245), and 9% failure response (22/245). Additionally, responses to TKI were gradually decreased at month 12 after first time point of monitoring as following 65% optimal (160/245), 13% warning (31/245), and 22% failure (54/245). We could detect 20% (49/245) of patients positive for BCR-ABL1 TKD mutations. Interestingly, one third (17 of 49) of TKD mutated cases were positive for compound/polyclonal mutation patterns. While major molecular response were observed in the majority of patients without TKD mutation, resistant to TKI were detected in patients with T315I mutation (n = 9; % mean IS = 8.1510, % median IS = 9.7000), compound/polyclonal mutations with T315I (n = 9; % mean IS = 13.0779, % median IS = 5.404), and other TKD mutations (n = 14; % mean IS = 8.1416, % median IS = 1.060), respectively. Conlusion: These practical laboratory techniques provided a more comprehensive understanding of CML progression during drug therapy and could be of benefit in earlier prognosis.

MicroRNA Expression Profiling of Epithelial Ovarian Cancer Identifies New Markers of Tumor Subtype.

BACKGROUND: Epithelial Ovarian Cancer (EOC) is often challenging to diagnose, even though histological examination. MicroRNA (miRNA or miRNA) is bound to the target messenger RNA (mRNA) due to which the mRNA molecules are silenced. The identification of miRNA expression- based EOC subtypes is considered a critical means of prognostication. So far, the studies on EOC subtypes have not been well characterized. OBJECTIVE: This study aimed to confirm the existence of miRNAs in EOC and to assess their potential as clinical biomarkers for EOC. METHODS: We sampled 25 ovarian tumor tissues from patients with human ovarian tumors (17 malignant; 12 serous EOC, five non-serous EOC, and eight benign ovarian tumors). miRNA microarray detection was performed to identify EOC miRNAs. Real-time PCR was adapted for the validation of differentially expressed miRNAs detected by microarray analysis related to hybridization intensity. RESULTS: The results confirmed that miRNAs exist in EOC, relative expression of EOC miRNAs demonstrated that the upregulation of miR-483-5p, and downregulation of miR-127-3p, and miR- 532-5p were significantly expressed in the malignant group than in the benign group at p < 0.05. Besides, miR-483-5p could also distinguish EOC subtypes between serous EOC and non-serous EOC, with a p < 0.05. CONCLUSION: A comprehensive miRNA expression profiling can help to refine subtype classification in EOC, opening new opportunities for identifying clinically applicable markers for improved stratification and diagnostics of ovarian tumors.

Performance of the MLPA technique for detecting common mutations in Leber hereditary optic neuropathy.

Leber hereditary optic neuropathy (LHON) causes painless vision loss resulting from mitochondrial DNA (mtDNA) mutation. Over 95% of LHON cases result from one of three mtDNA point mutations (m.3460G>A, m.11778G>A, and m.14484T>C). There is no established cure for LHON; early and accurate diagnosis would enable patients to be given appropriate treatments leading to a reduction of the disease progression. To increase the accessibility to molecular genetic testing for LHON, an accurate and cost-effective technique is required. The purpose of this study was to evaluate the accuracy of multiplex ligation-dependent probe amplification (MLPA) for detecting the three common mutations in 18 LHON blood specimens. Validation of the results using direct DNA sequencing technology proved that the MLPA technique had 100% accuracy, with no false-positive results. This study demonstrates that MLPA could provide a highly accurate, economical, and widely accessible technique for routine molecular genetic testing for mitochondrial disorders.

Direct STR typing from fired and unfired bullet casings.

Cartridges, bullets, and casings (CBCs) are commonly recovered after shooting incidents and could provide valuable DNA information from touch DNA that has been left behind during handling of bullets and loading of guns. Direct PCR, in which the DNA extraction and quantification steps are bypassed, has been shown to provide comparable and sometimes better results from touch DNA and trace DNA samples. Here, direct PCR was applied to touch DNA retrieved from bullet casings from three ammunition types and guns. The results were evaluated to determine whether the technique should be recommended as a standard operating protocol for forensic DNA analysts. Three experiments were carried out to investigate the following: the effect of firing bullets on DNA deposited on bullet casings; the effect of gun and ammunition types on short tandem repeat (STR) profile quality; and the feasibility of using direct PCR in real-world cases via typing of mock casework samples. DNA extraction resulted in a loss of about 40% of DNA originally deposited, and firing a bullet reduced the amount of DNA recovered by 27%. Using the direct PCR protocol, conventional extraction protocol, and dilution protocol on touch DNA from fired bullet casings, we recovered means (and 95% credible intervals), respectively, of 11.1 (7.9-13.9), 5.6 (3.0-7.7), and 2.3 (0.2-4.0) alleles. No statistical difference in alleles recovered was observed between different calibers of ammunition fired from three guns (9mm, 7.62mm, and 5.56mm from Glock Model 19, AK47, and Tavor T-21, respectively). As expected, mixed DNA profiles were observed in 40% of the mock casework samples, in which guns were shared between volunteers. This study showed that direct amplification of touch DNA from bullet casings improved STR profiles. As direct PCR is quicker, cheaper, and resulted in more alleles recovered, forensic DNA analysts may benefit from using direct PCR.

Cytogenetics and FLT3-ITD mutation predict clinical outcomes in non transplant patients with acute myeloid leukemia.

BACKGROUND: Cytogenetic abnormalities and mutated genes indicate the role of consolidation therapy with hematopoietic stem cell transplantation (HSCT) or chemotherapy in acute myeloid leukemia (AML). In this study, we conducted a retrospective study in adult AML patients with newly diagnosed with de novo AML who did not undergo HSCT, to study long term relapse free survival (RFS) and overall survival (OS) after consolidation chemotherapy. METHODS: We recruited 141 consecutive AML patients during January 2010-June 2017, the patients received induction chemotherapy with standard dose Ara-C and Idarubicin (7 + 3 or 5 + 2 regimen) followed by intermediate (IDAC) or high dose Ara-c (HiDAC) consolidation therapy. RESULTS: Normal karyotype, complex, favorable, intermediate and adverse chromosomal aberrations were found in 59%, 16%, 5%, 14% and 6%, respectively. Mutated NPM1, FLT3-ITD and CEBPA genes in CN-AML were seen in 33%, 18% and 19%, respectively. A 5 year follow up, 5y-RFS was 16% and 5y-OS was 14% in the whole study population. 5y-RFS and 5y-OS in patients completed 4 cycles of consolidation therapy were 25% and 40%, respectively. Adverse cytogenetic risk and mutated FLT3-ITD were significantly associated with poor RFS (9 and 15 months, respectively) and OS (14 and 16 months, respectively), whereas patients with mutant NPM1 had favorable outcomes (RFS/OS = 51/63 months). Patients receiving 4 cycles of consolidation therapy had significantly impacts on median RFS and OS compared with those treated with 1 or 2 cycles; 15 versus 11 months (p = 0.006) and 31 versus 15 months (p < 0.001), respectively. CONCLUSIONS: Cytogenetic and mutation tests for FLT3-ITD, NPM1 and CEBPA genes were meaningful for predicting outcomes in adult AML patients. Adverse cytogenetic abnormalities and FLT3-ITD mutation showed dismal RFS and OS.

Characterization of five complete Cyrtodactylus mitogenome structures reveals low structural diversity and conservation of repeated sequences in the lineage.

Mitochondrial genomes (mitogenomes) of five Cyrtodactylus were determined. Their compositions and structures were similar to most of the available gecko lizard mitogenomes as 13 protein-coding, two rRNA and 22 tRNA genes. The non-coding control region (CR) of almost all Cyrtodactylus mitogenome structures contained a repeated sequence named the 75-bp box family, except for C. auribalteatus which contained the 225-bp box. Sequence similarities indicated that the 225-bp box resulted from the duplication event of 75-bp boxes, followed by homogenization and fixation in C. auribalteatus. The 75-bp box family was found in most gecko lizards with high conservation (55-75% similarities) and could form secondary structures, suggesting that this repeated sequence family played an important role under selective pressure and might involve mitogenome replication and the likelihood of rearrangements in CR. The 75-bp box family was acquired in the common ancestral genome of the gecko lizard, evolving gradually through each lineage by independent nucleotide mutation. Comparison of gecko lizard mitogenomes revealed low structural diversity with at least six types of mitochondrial gene rearrangements. Cyrtodactylus mitogenome structure showed the same gene rearrangement as found in most gecko lizards. Advanced mitogenome information will enable a better understanding of structure evolution mechanisms.

The Frequency of SF3B1 Mutations in Thai Patients with Myelodysplastic Syndrome

Genetic mutations in genes encoding critical component of RNA splicing machinery including SF3B1 are frequently identified and recognized as the pathogenesis in the development of myelodysplatic syndrome (MDS). In this study, PCR sequencings specific for SF3B1 exon 13, 14, 15, and 16 were performed to analyse genomic DNA isolated from bone marrow samples of 72 newly diagnosed MDS patients. We found that 10 of 72 (14%) patients harbor SF3B1 missense mutations including E622D (1/72), R625C/G (2/72), H662Q (1/72), K666T (1/72), K700E (4/72) and G740E (1/72), respectively. Mutations were predominantly located on exon 14 and 15 of SF3B1 coding sequence. Interestingly, patients with SF3B1 mutations exhibited higher platelet counts (195×109/L VS. 140×109/L, p-value = 0.025) as well as lower hemoglobin levels (81 g/L VS. 92 g/L, p-value = 0.009) and associated with ring sideroblast phenotype (p-value < 0.001) when compared with patients without the SF3B1 mutation. In summary, we reported the frequency of SF3B1 mutations in Thai patients with different subtypes of MDS. SF3B1 mutations were predominantly occurred in MDS-RS and considered as favourable prognosis value. This study further highlighted the clinical important of SF3B1 mutations analysis for the classification of MDS.

EGFL7 and RASSF1 promoter hypermethylation in epithelial ovarian cancer.

DNA methylation is one of the epigenetic mechanisms associated with gene expression and plays a key role as in activation and deactivation of oncogenes and tumor suppressor genes, respectively. This study employed DNA methylation array to identify methylated genes which are highly correlated with various phenotypes of epithelial ovarian cancer (EOC) in Thai patients and to quantify promoter CpG-island methylation of candidate genes. Tissues from patients with serous and non-serous EOC showed significantly higher promoter methylation of EGFL7 and RASSF1 compared to benign cases. These results indicate the potential of investigating promoter CpG-island methylation of cancer-associated genes as biomarkers of disease progression and even possibly of early detection.

The Utilization of Karyotyping, iFISH, and MLPA for the Detection of Recurrence Genetic Aberrations in Multiple Myeloma

Multiple myeloma (MM) is a hematological malignancy characterized by abnormal accumulation of clonal plasma cells in the bone marrow. Recently, multiplex ligation-dependent probe amplification (MLPA) has emerged as an effective and robust method for detection of common genetic alterations in MM patients. Here, we aimed to confirm MLPA utility for this purpose and furthermore to test the feasibility of a combination of karyotyping, interphase fluorescence in situ hybridization (iFISH) and MLPA methods for diagnosis, prognostic assessment and risk stratification of MM. Thirty-five genomic DNA samples isolated from CD138-enriched plasma cells from bone marrow of MM patients were analyzed using the MLPA method. We found that amp (1q) was the most frequent genetic alteration (48.6%) in the tested samples, followed by del (1p) and del (13q) (34.3%). Moreover, concordant results between sensitivity and specificity of iFISH and MLPA for the detection of del (13q) (p-value >0.05) and del (17p) (p-value >0.05) were obtained. In summary, we could provide evidence of MLPA assay utility for the detection of common genetic alterations in MM. The combination of karyotyping, iFISH, and MLPA proved very helpful for clinical risk stratification.

Mutation Analysis of Isocitrate Dehydrogenase (IDH1/2) and DNA Methyltransferase 3A (DNMT3A) in Thai Patients with Newly Diagnosed Acute Myeloid Leukemia

Acute myeloid leukemia (AML) is a clonal hematopoietic stem/progenitor cell disorder which features several genetic mutations. Recurrent genetic alterations identified in AML are recognized as causes of the disease, finding application as diagnostic, prognostic and monitoring markers, with potential use as targets for cancer therapy. Here, we performed a pyrosequencing technique to investigate common mutations of IDH1, IDH2 and DNMT3A in 81 newly diagnosed AML patients. The prevalences of IDH1, IDH2 and DNMT3A mutations were 6.2%, 18.5%, and 7.4%, respectively. In addition, exclusive mutations in IDH1 codon 132 (R132H, R132C, R132G and R132S) were identified in all IDH1-mutated cases indicating that these are strongly associated with AML. Interestingly, higher median blast cell counts were significantly associated with IDH1/2 and DNMT3A mutations. In summary, we could establish a routine robust pyrosequencing method to detect common mutations in IDH1/2 and DNMT3A and demonstrate the frequency of those mutations in adult Thai AML patients.