Evaluating the clinical significance of FLT3 mutation status in Syrian newly diagnosed acute myeloid leukemia patients with normal karyotype.

The FMS-like tyrosine kinase-3 internal tandem duplication (FLT3-ITD) is one of the most prevalent mutations, affecting between 20 and 30 percent of cases in patients with acute myeloid leukemia (AML). The Patients with a FLT3-ITD mutation have a poor prognosis. In the present study, we investigated the FLT3 (ITD-TKD) mutations in 100 newly adult Syrian patients with AML-Normal karyotype (NK). Our results revealed that prevalence of FLT3-ITD mutation was 24%. Interestingly, 20 patients had a typical duplication mutation and four patients had different mutations. From those four mentioned patients, two of them carried a 39 base pair (bp) duplication in different location: (c.1838_1877dup39, p.591-603dup) and (c.1836_1874 dup 39, p.591-603dup), the third patient, showed FLT3-ITD duplication and a newly insertion together, this insertion was not demonstrated before: (c.1842_1865dup24, c.1865_1866insGAA). Finally, the fourth patient exhibited a duplication of 21bp (c.1855_1875dup21, p.597-603dup). In addition, statistically significant differences were observed for the relation between the presence of FLT3-ITD mutation and lactate dehydrogenase (LDH) level, overall survival (OS), relapse, and event free survival (EFS). We demonstrated that our patients with FLT3-ITD mutation had a poor prognosis. Also, the frequency of FLT3-TKD mutation was low 2% and no compound between the two mutations was found, as individuals showed to carry the two mutations were not detected. These findings are likely useful for a better understanding of molecular leukemogenetic steps in AML-NK patients and may be beneficial for clinical relevance for risk grouping, study design and choice of therapy in Syrian population.

Prognostic Relevance of DNMT3A, FLT3 and NPM1 Mutations in Syrian Acute Myeloid Leukemia Patients.

OBJECTIVE: Among all types of hematological neoplasms, acute myeloid leukemia (AML) has the highest death rate. Recently, cytogenetic and molecular genetics are crucial in the management, as a consequence of their effect on AML pathogenesis, classification, risk-stratification, prognosis and treatment. METHODS: 100 Syrian adults with Normal Karyotype (NK) newly diagnosed  AML patients were included in this study, all cases confirmed histologically and immunohistochemically. Patients were divided into six subgroups using flow cytometry and cytological results. Polymerase chain reaction (PCR) was performed on exon 11-12 for FMS-like tyrosine kinase-3 internal tandem duplication (FLT3-ITD), exon 12 for Nucleophosmin1 (NPM1), and exon 23 for DNA methyltransferase 3A (DNMT3A) using target primers, the electropherograms were analyzed for gene mutations by comparing with the reference DNA sequence. Data were compared and aligned with different sequences using the NCBI BLAST Assembled Genomes tool. RESULTS: FLT3-ITD, NPM1 and DNMT3A were detected in 24%, 22 % and 4%  patients respectively. M2 subtype had the most frequent incidence of diagnosis in AML. FLT3-ITD mutation patients had the highest mean of death cases, while the DNMT3A mutation patients had the lowest. On the other hand, the highest mean of remission was in patients with NPM1 mutation and the lowest in the carriers of the FLT3-ITD mutation. It was observed that the mean relapsed patients with FLT3-ITD and DNMT3A mutation was 3.4 and 2 months respectively, with no significant differences between (FLT3-ITD and DNMT3A) carriers and non-carriers relapsed. On the contrary,  the mean relapsed for NPM1 mutation carriers was 2.4  months with significant statistical differences. The mean survival time for patients with FLT3-ITD and NPM1  mutation was 5.9 months and 5.85 months respectively, with significant correlation. Between it was 5.88 months in DNMT3A patients with no significant differences. Finally, It was noted that the mean event free survival (EFS) of FLT3-ITD mutation patients was 4.818 months and the mean EFS of NPM1 mutation patients was 4.805 months, with significant statistical differences (p<0.05) between the mutation patients and non-mutated patients regarding to EFS, While this mean was not statistically significant in patients carrying DNMT3A mutation. CONCLUSION: Patients with FLT3-ITD and NPM1 mutations have the worst prognosis, where the presence of those mutations was significantly related to overall survival (OS) and EFS. Our study reflects that DNMT3A was not an extremely bad prognostic effect as an independent factor. We can declare according to this study that genetic mutation and variants detection could easily be incorporated into the regimen evaluation of AML patients.

Acute myeloid leukemia due to germline CEBPA mutation in a Syrian family.

BACKGROUND: Familial cases of adult acute myeloid leukemia (AML) with germline-mutated CCAAT/enhancer-binding protein-α (CEBPA) gene are a rare entity classified in World Health Organization (WHO) classification 2016. Most families reported in the literature show an autosomal dominant inheritance pattern consistent with a single-gene mutation. METHODS: Here we studied a Syrian family with four individuals suffering from AML for CEBPA gene mutations by Sanger sequencing. RESULTS: The father, his three affected, and one yet unaffected child had the same mutation in the N-terminal region of CEBPA (c.198dupC), resulting in termination at Tyr67Leufs*41. All affected family members had a good primary response to chemotherapy and achieved complete remission. CONCLUSION: Overall, another AML family with CEBPA gene mutation is added to the literature, presenting with yet unreported FAB subtype M5 and absence of CD7 expression in some family members.

Frequency of FLT3 Internal Tandem Duplications in Adult Syrian Patients with Acute Myeloid Leukemia and Normal Karyotype.

OBJECTIVE: Activating mutations of the fms-like tyrosine kinase 3 gene (FLT3) by internal tandem duplications (ITDs) in the juxtamembrane domain (JMD) have been reported in ~30% of adult acute myeloid leukemia (AML) patients with cytogenetically normal karyotype (CN). However, FLT3/ITD mutations are frequently accompanied with leukocytosis, high percentage of blasts in bone marrow (BM), and increased the risk of treatment failure in AML patients. FLT3-ITD mutated AML patients mainly with normal karyotype have higher relapse probability and shorter duration of complete remission (CR) after chemotherapy, so FLT3-ITD mutation is considered as an independent poor prognostic factor in AML. METHODS: FLT3-ITD and FLT3-KTD were studied by polymerase chain reaction (PCR) and restriction fragment length polymorphism- PCR (RFLP-PCR) in 44 adults AML patients with cytogenetically normal karyotype (AML-CN) at diagnosis to characterize FLT3 status. The results were correlated with the prognostic factors. RESULTS: In this study, FLT3-ITD mutations were identified in 7 (15.9%) of the 44 AML-CN patients. Among the 7 patients with FLT3/ITD mutations, 6 patients revealed a typical ITDs mutation (fragment size was 329 bp) and one patient showed untypical ITD mutation (fragment size was ~400 bp). Whereas 37 patients (61.7%) were FLT3-ITD. None of all AML-CN patients examined showed FLT3-KTD mutations. CONCLUSIONS: Our results support that FLT3-ITD are independent adverse prognostic factors for elderly AML-CN patients and are associated with low overall survival (OS), low rate of CR, high relapse rate (RR), and high percentage of BM blast at diagnosis. We concluded, FLT3 mutation analysis should be performed as a routine test in AML-CN patients.

Two Novel Mutations of the NPM1 Gene in Syrian Adult Patients with Acute Myeloid Leukemia and Normal Karyotype.

OBJECTIVE: Somatic mutations in exon 12 of the NPM1 gene is one of the most common genetic abnormalities in adult acute myeloid leukemia (AML), which is observed in 25-35% of AML patients and in 50-60% of patients with cytogenetically normal AML (CN-AML). METHODS: We performed Sanger sequencing of exon 12 of the NPM1 gene, on 44 CN-AML patients to characterize NPM1 status. RESULTS: In this study, NPM1 mutations were identified in 10 (22.7%) of the 44 CN-AML patients. Among the 10 patients with NPM1 mutations, type A NPM1 mutations were identified in 8 (80%) patients, whereas non-A type NPM1 mutations were observed in 2 (20%) patients. Two non-A type NPM1 mutations were not previously reported: c.867-868InsCGGA and c.861-862InsTGCA. These two novel mutant proteins display a nuclear export signal (NES) motif (L-xxx-L-xx-V-x-L) less frequently and L-x-Lx-V-xx-V-x-L it has been never seen before, yet. However, both novel mutations show a tryptophan loss at codon 288 and 290 at the mutant C-terminus which are crucial for aberrant nuclear export of NPM into the cytoplasm. CONCLUSIONS: This study suggests previously unreported NPM1 mutations may be non-rare and thus additional sequence analysis is needed along with conventional targeted mutational analysis to detect non type-A NPM1 mutations.

De novo adult acute myeloid leukemia with two new mutations in juxtatransmembrane domain of the FLT3 gene: a case report.

BACKGROUND: Approximately 30% of adult acute myeloid leukemia (AML) acquire within fms-like tyrosine kinase 3 gene (FLT3) internal tandem duplications (FLT3/ITDs) in their juxtamembrane domain (JMD). FLT3/ITDs range in size from three to hundreds of nucleotides, and confer an adverse prognosis. Studies on a possible relationship between of FLT3/ITDs length and clinical outcomes in those AML patients were inconclusive, yet. CASE PRESENTATION: Here we report a 54-year-old Arab male diagnosed with AML who had two FLT3-ITD mutations in addition to NPM1 mutation. Cytogenetic approaches (banding cytogenetics) and fluorescence in situ hybridization (FISH) using specific probes to detect translocations t(8;21), t(15;17), t(16;16), t(12;21), and deletion del(13q)) were applied to exclude chromosomal abnormalities. Molecular genetic approaches (polymerase chain reaction (PCR) and the Sanger sequencing) identified a yet unreported combination of two new mutations in FLT3-ITDs. The first mutation induced a frameshift in JMD, and the second led to a homozygous substitution of c.1836T>A (p.F612L) also in JMD. Additionally a NPM1 type A mutation was detected. The first chemotherapeutic treatment was successful, but 1 month after the initial diagnosis, the patient experienced a relapse and unfortunately died. CONCLUSIONS: To the best of our knowledge, a combination of two FLT3-ITD mutations in JMD together with an NPM1 type A mutation were not previously reported in adult AML. Further studies are necessary to prove or rule out whether the size of these FLT3-ITDs mutations and potential other double mutations in FLT3-ITD are correlated with the observed adverse outcome.

Investigation of the mtDNA mutations in Syrian families with non-syndromic sensorineural hearing loss.

OBJECTIVE: Hearing loss is a common sensory disorder, and at least 50% of cases are due to a genetic etiology. Several mitochondrial DNA mutations (mtDNA) have been reported to be associated with nonsyndromic hearing loss (NSHL) in different population. However, There is no previous available data about the frequency of mtDNA mutations as etiology for deafness in Syrian. The aim of present study is to investigate the incidence of common mt DNA mutations in our families with congenital hearing loss and not related to the ototoxicity or aminoglycosides. METHODS: A total of 50 deaf families were enrolled in the present study. Direct sequencing and PCR-RFLP methods were employed to detect seven mt DNA mutations, including A1555G, A3243G, C1494T, G3316A, T7510C, A7445G, and 7472insC. RESULTS: Our results revealed a high prevalence of mt DNA mutation (10%) in deaf families (5/50). In surprising, the unexpected mutations were observed. The G3316A mutation was found in 2 families as homoplasmic genotype. Also, we found the homoplasmic and heteroplasmic genotype for the C1494T mutation in two families. In one family the heteroplasmic genotype for T7510C mutation was observed; this family harbor 35delG mutation in GJB2 gene. None of the common mtDNA mutations (A1555G, A3243G) and other mutations (A7445G, 7472insC) were detected here. CONCLUSION: Our findings indicate to significant contribution of the mt DNA mutations in our families with NSHL. The presented data is the first report about mt DNA and it will improve the genetic counseling of hearing impaired in Syrian families.

Down syndrome associated childhood myeloid leukemia with yet unreported acquired chromosomal abnormalities and a new potential adverse marker: dup(1)(q25q44).

BACKGROUND: Children with constitutional trisomy 21, i.e. Down syndrome (DS, OMIM #190685) have a 10 to 20-fold increased risk for a hematopoietic malignancy. They may suffer from acute lymphoblastic leukemia or acute myeloid leukemia (AML). AML referred to as myeloid leukemia of Down syndrome (ML-DS) is observed especially after birth at an early gestational age and characterized by enhanced white blood cell count, failure of spontaneous remission, liver fibrosis or liver dysfunction, and is significantly associated with early death. There are only few studies yet focusing on the clonal cytogenetic changes during evolution of ML-DS. CASE PRESENTATION: In a 1.4-year-old boy with DS an immunophenotype consistent with AML-M1 according to French-American-British (FAB) classification was diagnoses. Cytogenetic and molecular cytogenetic analyses revealed, besides constitutional free trisomy 21, an unbalanced translocation as der(16)t(1;16)(q25.3;q24), plus a balanced translocation t(3;20)(q25;q13.1). A poor clinical outcome was observed here. CONCLUSIONS: To the best of our knowledge, an ML-DS case associated with identical acquired chromosomal abnormalities was not previously reported. Our findings suggest that especially partial trisomy 1q25 to 1q44 may be indicative for a poor prognosis in ML-DS.

Association of Methylenetetrahydrofolate Reductase C677T and A1298C Gene Polymorphisms With Recurrent Pregnancy Loss in Syrian Women.

C677T polymorphism of the methylenetetrahydrofolate reductase ( MTHFR) gene was a risk factor for recurrent pregnancy loss (RPL), but few studies have confirmed a possible role of MTHFR A1298C polymorphism in RPL risk. This study was carried out to determine the influence of the MTHFR gene polymorphisms in RPL Syrian women. A case-control study was performed on 2 groups (106 healthy and 100 RPL women). The frequency of the MTHFR gene polymorphisms was determined by polymerase chain reaction based on restriction fragment length gene polymorphism. In the RPL group, the genotype frequencies of MTHFR C677T were CC (41%), CT (41%), and TT (18%), and in the control group, the frequencies were CC (62.2%), CT (36.7%), and TT (1%). Statistical analysis showed a homozygous TT genotype and T allele were significantly different in the RPL group ( P = .000003 and P = .000019, respectively). The genotype frequencies of MTHFR A1298C were AA (53%), AC (44%), and CC (8%) in the RPL group, whereas in the control group, these were AA (61.3%), AC (37.8%), and CC (1%). A significant difference in the CC genotype and C allelic frequencies in the RPL women was observed ( P = .014 and P = .064, respectively). The patients having compound heterozygous (677 CT/1298AC) were associated with an estimated 4.86-fold increase in risk of pregnancy loss compared to individuals with a wild type ( P = .012). Our findings indicate that RPL women with homozygous genotype for (C677T and A1298C) either alone or compound heterozygous genotypes have a high risk of pregnancy loss in Syrian women.

First report of prevalence c.IVS1+1G>A and del (GJB6-13S1854) mutations in Syrian families with non-syndromic sensorineural hearing loss.

OBJECTIVE: Mutations in GJB2 and GJB6 genes are a frequent cause of congenital non-syndromic hearing loss (NSHL). Mutational screening has usually focused on coding region of GJB2 gene. A few studies have been conducted on the non-coding region and exon 1. c.IVS1+1G>A (a splice site mutation in GJB2 gene have been detected as disruptive mutation. Del (GJB6 D13S1830) is found in many populations, but del (GJB6 D13S1854) is reported from a few restricted countries. This study was carried out to investigate the prevalence of splice site mutation c.IVS1+1G>A and two common deletions in GJB6 gene as the genetic etiology of hearing impairment in 70 Syrian families. METHODS: The frequency of the c.IVS1+1G>A mutation and two deletions were determined by PCR-RFLP and A multiplex PCR assay. RESULT: Our results showed a high prevalence of IVS1+1G>A mutation (20%) and del(GJB6-D13S1854) (15.7%) in deaf families. The homozygous genotype (c.IVS1+1G>A/c.IVS1+1G>A) was observed in one family and the compound heterozygous genotypes (c.35delG/c.IVS1+1G>A) and (c.IVS1+1G>A/V153I) were observed in 7 families and one family respectively. Also, the heterozygous state (c.IVS1+1G>A/unknown) was detected in 5 families. The study of del((GJB6-D13S1854) was showed a compound heterozygous genotype del((GJB6-D13S1854)/c.IVS1+1G>A) in the same families (5 families) having heterozygous genotype of c.IVS1+1G>A mutation. Also, del(GJB6-D13S1854) is combined with c.35delG mutation in 2 families and it was observed in the heterozygous state del(GJB6-D13S1854)/unknown) in 4 families. In contrast, the del(GJB6-D13S1830) described in many population was absent in our patients. CONCLUSION: Our findings indicate to significant contribution of the splice site mutation and del(GJB6-D13S1854) in our deaf families and these mutations were important causes of hearing impairment.

A high complex karyotype involving eleven chromosomes including three novel chromosomal aberrations and monoallelic loss of TP53 in case of follicular lymphoma transformed into B-cell lymphoblastic leukemia.

BACKGROUND: Follicular lymphoma (FL) is one of the most common B-cell non-Hodgkin's lymphoma (NHL). A subset of FL cases transform into more aggressive malignancies, most often to diffuse large B-cell lymphoma (DLBCL); in addition, lymphoblastic lymphoma and acute lymphoblastic leukemia (ALL) have also been rarely reported. The most common cytogenetic abnormalities associated with FL are translocation t(14;18)(q32;q21) with BCL2 rearrangements, present in 80-90% of all FL. However, that translocation alone is insufficient to cause FL and additional genomic events specifically leading to this kind of disease are still to be determined. The most frequently reported secondary changes are gains of chromosomes 7p or 7q, Xp, 12q and 18q, as well as losses on 6q and mutations within BCL2 and/or BCL6 genes. The presence of additional genomic aberrations, in particular 17p and 6q deletions is more frequent in grade 2 and 3 FL patients and correlated with shorter survival and a higher rate of transformation into DLBCL. CASE PRESENTATION: We describe here, an adult FL grade 2 patient that had transformed to B-ALL at diagnosis. Banding cytogenetics, refined by multi-color fluorescence in situ hybridization including array-proven multicolor banding revealed a unique complex karyotype involving eleven chromosomes, translocation t(X;20)(p21.3;q11.2), translocation t(3;20)(q26.2;q12), and a dicentric dic(17;18). Interestingly, the dicentric chromosome led to monosomy of the tumor suppressor gene TP53. The case had an immunophenotype consistent with follicular center cell lymphoma according to the World Health Organization (WHO) recommendations. CONCLUSIONS: To the best of our knowledge, a comparable adult FL grade 2 case that transformed to B-ALL associated with such a complex karyotype and loss of TP53 was not previously reported. Most of complex aberrations were found simultaneously in approximately 85% of studied malignant cells and the remained cells studied were non-clonal; mechanisms explaining this may be either multiple-step mechanisms or single step in sense of chromothripsis. TRIAL REGISTRATION: Identifying number: 3842. Registered 09 July 2012.

Acute promyelocytic leukemia with the translocation t(15;17)(q22;q21) associated with t(1;2)(q42~43;q11.2~12): a case report.

BACKGROUND: Acute promyelocytic leukemia is characterized by a typical reciprocal translocation t(15;17)(q22;q21). Additional chromosomal abnormalities are reported in only 23-43 % of cases of acute promyelocytic leukemia. CASE PRESENTATION: Here we report the case of a 46-year-old Syrian Alawis woman with acute promyelocytic leukemia with the typical t(15;17) translocation, but with a second clone presenting a t(1;2)(q42~43;q11.2~12) translocation as an additional abnormality. To the best of our knowledge, an association between these chromosomal abnormalities has not previously been described in the literature. Our patient started treatment with all-trans retinoic acid 10 days after diagnosis but died the same day of treatment initiation due to hemolysis, intracranial hemorrhage, thrombocytopenia, and disseminated intravascular coagulation. CONCLUSION: The here reported combination of aberrations in a case of acute promyelocytic leukemia seems to indicate an adverse prognosis, and possibly shows that all-trans retinoic acid treatment may be contraindicated in such cases.

Correlation of p210 BCR-ABL transcript variants with clinical, parameters and disease outcome in 45 chronic myeloid leukemia patients.

PURPOSE: The aim of this study was to search the BCR/ABL 1 fusion gene in 45 chronic myeloid leukemia (CML) Syrian patients using nested reverse transcription polymerase chain reaction (RT-PCR) and compare our results with those of conventional cytogenetics and molecular cytogenetics methods. METHODS: 45 bone marrow or peripheral blood samples from untreated CML patients in chronic phase (CP) were obtained at diagnosis, and analyzed by nested RT-PCR, conventional cytogenetics and molecular cyto-genetics methods. RESULTS: 45 patients examined were positive for some type of BCR/ABL1 fusion gene rearrangement. Out of 45 studied CML patients, 23 (51.1%) expressed b3a2 fusion transcript, 21 (46.7%) b2a2 transcript, and 1 (2.2%) a rare b2a3 transcript. No patient co-expressed both b3a2/b2a2 types. CONCLUSIONS: The distribution BCR-ABL1 transcript types found in Syria were similar to that of Indian Far-Eastern, African or European populations and the M-BCR rearrangement types were not dependent on white blood count (WBC), platelet count, hemoglobin level or gender of the patients. Overall, we could show that patients with b3a2 rearrangements were younger than patients with b2a2 transcripts, thus our young patients may have a worse prognosis.

Chromosomal aberration leads to recurrent pregnancy loss and partial trisomy of 5p12-15.3 in the offspring: report of a Syrian couple and review of the literature .

Here we describe a Syrian couple having recurrent pregnancy loss in the first trimester, fetal malformations, and/or neonatal death. The father had a balanced chromosomal translocation t(5;15), an sY125 microdeletion of locus b in the azoospermia factor (AZF) gene, and an MTHFR C677T homozygous polymorphism with normal phenotype. Interestingly, his healthy wife had another MTHFR A1298C homozygous polymorphism. The couple experienced two pregnancy losses and had two stillborn children with severe malformations due to partial trisomy of the short arm of chromosome 5. The couple does not have any living offspring after 10 years of marriage.

An adult B-cell precursor acute lymphoblastic leukemia with multiple secondary cytogenetic aberrations.

BACKGROUND: We report a clinically diagnosed acute lymphoblastic leukemia (ALL) with yet unreported secondary chromosomal aberrations. RESULTS: A complete cytogenetic and molecular cytogenetic analysis, using GTG banding, fluorescence in situ hybridization (FISH) and array-proven multicolor banding (aMCB), for a female patient with clinically diagnosed ALL and immunophenotypically confirmed pre-B ALL (FAB classifications), revealed the presence of a complex structural rearrangement, der (2) (20qter- > 20q13.33::2q21- > 2p14::2q21 > 2qter) along with t (9;22) (q34;q11), t (12;14) (q12;p12) and a monosomy of chromosome 7. CONCLUSIONS: Molecular cytogenetic studies are suited best for identification and characterization of chromosomal rearrangements in acute leukemia. Single case reports as well as large scale studies are necessary to provide further insights in karyotypic changes taking place in human malignancies.

Influence of CYP1A1, GST polymorphisms and susceptibility risk of chronic myeloid leukemia in Syrian population.

In the present study, we investigated the associations of polymorphisms in cytochrome P450 gene (CYP1A1), glutathione S-transferase genes (GSTM1 and GSTT1) with chronic myelogenous leukemia (CML). A total of 126 patients with CML and 172 healthy volunteers were genotyped, and the DNA was isolated from their blood samples. The polymorphisms were assessed by polymerase chain reaction (PCR) restriction fragment length polymorphism-based methods and multiplex PCR. Logistic regression analyses showed significant risk of CML associated with CYP1A1 Val allele [odds ratio (OR) 3.3, 95% confidence intervals (CI) 1.96-5.53], (p < 0.0001) while CYP1A1 Val/Val homozygotes were observed only in the CML patients. There was statistically significant difference in the frequency of GSTM1 and GSTT1 null genotypes. The GSTT1-null genotype was slightly higher in 27% of CML cases and 16.7% of controls (OR 1.98, 95% CI 1.12-3.5) (p < 0.020). The GSTM1 null was higher in 42.8% of CML cases and 22.7% of controls (OR 2.55, 95% CI 1.54-4.22) (p < 0.00024). The individuals carrying CYP1A1 Ile/Val (AG) and GSTM1 null genotype have 9.9 times higher risk to be CML than those carrying CYP1A1 Ile/Ile (AA) and GSTM1 present genotype (OR 9.9, 95% CI 2.7-36.3) (p < 0.0001). This suggests that the association of the GSTM1 null genotype, either alone or in combination with GSTT1 null, with CYP1AI heterozygous leads to the CML risk.

Hyperdiploidy associated with T315I mutation in BCR-ABL kinase domain in an accelerated phase-chronic myeloid leukemia case.

BACKGROUND: Chronic myeloid leukemia (CML) is genetically characterized by the occurrence of a reciprocal translocation t(9;22)(q34;q11), resulting in a BCR/ABL gene fusion on the derivative chromosome 22, i.e. the Philadelphia (Ph) chromosome. During CML progression 60-80% of the cases acquire additional genetic changes. Even though hyperdiploidy is not a rare finding in advanced phase-CML, hyperdiploidy together with a T315I kinase domain (KD) mutation in the BCR-ABL gene has not yet been reported. RESULTS: A complete cytogenetic and molecular cytogenetic analysis; molecular biology methods such as quantitative reverse transcription polymerase chain reaction (RQ-PCR) and allele-specific oligonucleotide (ASO)-PCR; and immunophenotypically confirmed CML in acceleration phase (AP). Our case revealed the presence of hyperdiploidy including multiple copies of the Ph chromosome, presence of b3a2 fusion transcript,T315I mutation in BCR-ABL KD in pre imatinib mesylate (IM) treatment. The ratio of BCR-ABL/ABL expression in post nilotinib treatment was 0.07% on international scale. CONCLUSIONS: The patient demonstrated a good response to nilotinib after imatinib failure; while the hyperdiploid clone disappeared the T315I mutation remained during follow-up. The underlying mechanisms and prognostic implications of these cytogenetic abnormalities are discussed.

Multiple copies of BCR-ABL fusion gene on two isodicentric Philadelphia chromosomes in an imatinib mesylate-resistant chronic myeloid leukemia patient.

The so-called Philadelphia (Ph) chromosome is present in more than 90% of chronic myeloid leukemia (CML) cases. Amplification or duplication of the BCR-ABL gene has been found to be one of the key factors leading to drug resistance to imatinib mesylate (IM). In the present study, we identified the presence of isodicentric Ph chromosomes [idic(Ph)] in an IM-resistant patient. Fluorescence in situ hybridization (FISH) analysis on metaphase chromosomes confirmed the heterogeneity and amplification of the fused BCR-ABL gene. FISH analysis superimposed on G-banding confirmed the presence of idic(Ph) chromosomes. Reverse transcription-polymerase chain reaction (RT-PCR) products revealed the presence of the BCR-ABL fusion transcript b3a2. The idic(Ph) chromosomes in CML were shown to be fused at the satellite regions of the short arms. The patient did not respond to IM chemotherapy and did not achieve remission. In this study, the impact of the idic(Ph) chromosomes on genomic instability, heterogeneity and amplification of the BCR-ABL gene in IM-resistant patients is discussed.

Cytogenetic abnormalities and Y-chromosome microdeletions in infertile Syrian males.

Infertility is an important health issue affecting numerous couples. Approximately 30-50% of the cases of male infertility is due to unknown reasons. The main genetic factors involved in male infertility are chromosomal abnormalities and Y chromosome microdeletions within the Yq11 region. The genes controlling spermatogenesis located in the Yq11 region are termed azoospermia factor genes (AZF). Klinefelter syndrome (KS) is the most common of the chromosomal anomalies in the infertile male. AZF microdeletions on the Y chromosome are the most frequent genetic cause of male infertility. Screening for microdeletions in the AZFa, b and c regions of the Y chromosome showed a marked variation among different studies. The present study aimed to investigate the prevalence of such deletions in Syrian men. A total of 162 infertile males (97 azoospermic, 49 oligospermic and 16 severely oligospermic) were screened for chromosomal abnormalities and Y chromosome microdeletions using 28 markers in the AZF region. Twenty (12.34%) patients had chromosomal rearrangements, 17 of them showed sex chromosome abnormalities (11 of 17 patients within the azoospermic group had a KS of 64.7%), 2 patients had apparently balanced autosomal rearrangements, while 1 patient had an inversion. Of the 162 infertile men, 46 patients (28.4%) had Y chromosome microdeletions within the AZF-regions. Most frequently hit were the AZFc (34.8%), followed by the AZFbc, AZFa, AZFac, AZFbc, AZFb, AZFd, AZFab, AZFad, AZFbd, AZFabc and the AZFbcd. Combined AZF deletions involving three regions with chromosomal abnormalities were observed in one case. The higher frequency of AZF deletions in our study was comparable with frequencies in other countries and regions of the world, possibly due to the elevated number of the sequence-tagged site (STS) markers used for this screening.

A novel dic (17;18) (p13.1;q11.2) with loss of TP53 and BCR/ABL rearrangement in an Imatinib resistant chronic myeloid leukemia.

BACKGROUND: The so-called Philadelphia (Ph) chromosome is present in more than 90% of chronic myeloid leukemia (CML) cases. It results in juxtaposition of the 5' part of the BCR gene on chromosome 22 to the 3' part of the ABL gene on chromosome 9. Since the majority of CML cases are currently treated with Imatinib, variant rearrangements in general have no specific prognostic significance, although the mechanisms involved in resistance to therapy have yet to be investigated. The T315I mutation within the abl-gene is the most frequent one associated with resistance to tyrosine kinase inhibitors. RESULTS: This study evaluated a Ph chromosome positive CML case resistant to imatinib mesylate. A dic(17;18), loss of TP53 gene, co-expression of b2a2 and b3a2 fusions transcript and a T315I mutation were found. CONCLUSIONS: We reported here a novel case of a Ph chromosome positive CML with a secondary abnormality [dic(17;18)], resulting to Glivec resistance but good response to nilotinib. The dic(17;18) might be a marker for poor prognosis in CML. Our finding indicated for an aggressive progression of the disease. The patient died under the treatment due to unknown reasons.