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ИННОВАЦИОННЫЕ ТЕХНОЛОГИИ В ПЕДИАТРИИ И ДЕТСКОЙ ХИРУРГИИ
sis and were identified in our laboratory between the years 2000 and 2015. Of 458 CML cases 24 patients (5.2%) exhibited vPhs with overall 29 different breakpoints; the most frequent repeated breakpoint was 16p11.2, and chromosome 12 was the most frequently involved in our vPhs cases. The majority of the cases were simple translocations (75%) and 25% of these cases were complex translocations. Deletions in 9q34 on the der(9) were observed in four of the 22 vPh cases (18.1%), which was not associated with a lower rate of ABL1 or BCR deletion status compared to the average of ~40% reported in the literature. Mainly vPhs cases revealed a one-step mechanism of translocation formation (22 of 24 vPhs cases, 91.6%), despite one case exhibiting most likely a two-step mechanism, and only one case was due to a complex multi-step mechanism.
INFLUENCE OF GLUTATHIONE S-TRANSFERASE (GSTT1 AND GSTM1) GENE POLYMORPHISM ON IMATINIB FAILURE IN CHRONIC MYELOID LEUKEMIA PATIENTS
Al Achkar W.1, Moassass F.1, Aroutiounian R.2, Harutyunyan T.2, Liehr T.3, Wafa A.1
'Human Genetics Division, Department of Molecular Biology and Biotechnology, Atomic Energy Commission of Syria, Damascus, Syria 2Department of Genetics and Cytology, Yerevan State University, Yerevan, Armenia 3Institute of Human Genetics, Jena University Hospital, Jena, Germany
Chronic myeloid leukemia (CML) is genetically characterized by the occurrence of a reciprocal translocation t(9;22)(q34;q11), resulting in a BCR/ABL1 gene fusion on the derivative chromosome 22, i.e. the Philadelphia (Ph) chromosome. Imatinib (IM) is a chemically designed drug which blocks BCR/ABL1 tyrosine kinase activity and is successfully used in CML patients. However, some CML patients fail to reach adequate hematologic, cytogenetic and molecular responses or develop resistance to IM treatment. It has been reported that mutations or amplification of BCR-ABL1 kinase domain or variation in the bioavailability of IM may be linked to inherited genetic differences of enzymes involved in IM metabolism and thus those could explain interindividual differences in treatment outcome. Glutathione S-transferases (GSTs) are phase II xenobiotic metabolizing enzymes known to be involved in the detoxification of carcinogens and anticancer drugs. Individual genetic variation expressed as inherited polymorphisms of GSTT1 and GSTT1 may lead to complete loss of enzyme activity; this could expose subjects to develop cancer and/or to induce drug resistance. Here we assessed the role of such GSTs in relation to IM treatment outcome in 96 Syrian CML patients. Interestingly, patients carrying GSTT1 and GSTT1 deletions did not suffer from failed IM treatment more frequently than those having the genes not-deleted (P = 0.271). However, patients with GSTT1 and/or GSTT1 deletion developed more likely only minimal cytogenetic response and IM treatment was more likely to fail (P = 0.001). Also, the
same effect was present in patients with deletion of both copies of GSTT1 (P = 0.009). On the other hand However, 5/8patients with minimal cytogenetic response developed clonal cytogenetic evolution; four of those who had ho-mozygous deletion of GSTT1. GSTT1 gene presence or absence had no influence on the cytogenetic responses in the studied CML patients (P = 0.279). Cytogenetic response and GSTT1 and/or GSTT1 gene loss are therefore important determinants of IM failure in CML. Screening for GSTT1 and GSTT1 gene deletions at time of diagnosis may identify patients who may be better treated by an alternative therapy.
BIRC3 ALTERATIONS IN CHRONIC AND B-CELL ACUTE LYMPHOCYTIC LEUKEMIA PATIENTS
Alhourani E.1, Othman Moneeb A.K.2, Melo J.B.2-3, Carreira I.M.2-3, Grygalewicz B.4, Vujic D.5-6, Zecevic Z.6, Joksic G.7, Glaser A.1, Pohle B.1, Schlie C.1, Hauke S.8, Liehr T.1
•Jena University Hospital, Friedrich Schiller University, Institute of Human Genetics, Jena, Germany 2Laboratory of Cytogenetics and Genomics, Faculty of Medicine, University of Coimbra, Coimbra, Portugal 3CIMAGO, Centro de Investiga gaoem, Meio Ambiente, Genéticae Oncobiologia, Coimbra, Portugal "Cytogenetic Laboratory, Maria Sklodowska-Curie Memorial Cancer Centre and Institute, Warsaw, Poland 5University of Belgrade Faculty of Medicine, Belgarde, Serbia
6Mother and Child Health Care Institution of Serbia "Dr. Vukan Cupic", Belgrade, Serbia 7Vinca Institute of Nuclear Sciences, Belgrade, Serbia 8ZytoVision GmbH, Bremerhaven, Germany
Deletions within the long arm of chromosome 11, subbands q22 to q23 are considered to be among the most common chromosomal aberrations in chronic lympho-cytic leukemia (CLL). They are associated with a poor outcome. Besides the ATM also the BIRC3 gene is located in this region. The latter is a negative regulator of the non-canonical NF-kB protein. Moreover, BIRC3 disruption is known to be restricted to CLL fludarabine refractory patients. The aim of this study was to determine the frequency of copy number changes of BIRC3 gene and to align with two known negative predictors of CLL-outcome, i.e. ATM and TP53 deletions. Besides, the yet suggested specificity of BIRC3 alterations to CLL patients (117 patients were included in this study) BIRC3 copy numbers were also studied in 45 B-cell acute lymphocytic leukemia (B-ALL) patients. A commercially available multiplex ligation dependent probe amplification probe set was applied, which includes 4 probes for detection of TP53, and 4 others for ATM gene region. In-terphase-directed fluorescence in situ hybridization was done applying commercially available probes for BIRC3, ATM, and TP53. High resolution array-comparative genomic hybridization was applied for selected cases. Genetic abnormalities of BIRC3 were detected in 23/117 (~20%) CLL and also in 2/45 (~4%) B-ALL cases. Over-
РОССИЙСКИЙ ВЕСТНИКПЕРИНАТОЛОГИИ И ПЕДИАТРИИ, 4, 2015
Раздел 2 Педиатрия
all 20 CLL and one B-ALL case showed BIRC3 deletion, and three CLL and one B-ALL case BIRC3 duplication. All cases with ATM deletion went together with deletion in BIRC3. Only two CLL cases showed deletion in BIRC3, ATM, and TP53simultaneously. Obviously both, deletion and duplication of BIRC3 can be observed rarely in B-ALL. Also BIRC3 duplication can be found in CLL and the prognostic meaning has to be determined in future. The likelihood that TP53 deletions go together with BIRC3 and/or ATM aberrations is quite low. However, as ATM-deletion can, but must not go together with BIRC3-deletions, both regions should be tested both in future CLL-diagnostics to draw correct treatment decisions, namely the treatment with or without fludarabine. Supported in parts by KAAD (fellowship to EA), DAAD (fellowship to MAKO; PROBRAL 57054562 to TL; uni-versitypartnership program of FSU Jena to TL).
CLINICAL AND MOLECULAR CHARACTERIZATION OF A PATIENT WITH A 2Q22.3 TO 2Q24.1 DELETION
Babameto-Laku A.1, Mokini V.1, Roko D.1, Angioni A.2, Mingarelli R.2, Liehr T.3, Dallapiccola B.2
'Service of Medical Genetics, University Hospital "Mother Teresa", Faculty of Medicine, Tirana, Albania 2Direzione Scientifica IRCCS-Ospedale Pediatrico Bambino Gesu, Roma, Italy
Jena University Hospital, Friedrich Schiller University, Institute of Human Genetics, Jena, Germany
Most of the previously reported cases with interstitial deletion in the long arm of a chromosome 2 have been detected by banding cytogenetic techniques, showing poor breakpoint characterization. Applying array-based comparative genomic hybridization (array-CGH), the detection rate of submicroscopic chromosomal abnormalities improved considerably. Here we report an Albanian boy, five years old, with mental retardation, hypotonia, dysmorphic features and hand/foot abnormalities. A complex cryptic rearrangement of chromosome 2 including a small deletion 547 kb in size (arr[GRCh37/hgl9] 2p12(75,145,737-75,692,546)xl), that overlaps with a copy-number variant region and a deletion of 9.4 Mb (arr[GRCh37/hg19] 2q2 2.3q24.1(145,809,240-155,187,403)x1) were identified by array-CGH. Region 2q22.3 to 2q24.1 encompassed the genes MBD5 and EPC2. Interestingly, haploinsufficiency of MBD5 is known to be associated with microcephaly, intellectual disabilities, severe speech impairment, and seizures, as also seen in the present patient. Furthermore, the present case shares some clinical features with previously reported carriers of deletions within the chromosomal region 2q22 to 2q24.
The present case provides further evidence for a contiguous gene syndrome located in 2q23q24. It adds new information on the potentially critical region, involved genes and clinical features. Even though the ~500bp deletion in 2p12 just seems to be a copy number variant, according to the 'two-hit-model', an influence of such "copy number variants" cannot be completely neglected and suggest other studies.
INFLUENCE OF AFLATOXIN B1 ON COPY NUMBER VARIANTS IN HUMAN LEUKOCYTES IN VITRO
Harutyunyan T.1, Hovhannisyan G.1, Babayan N.1,2, Othman Moneeb A.K..3, Liehr T.3, Aroutiounian R.1
'Department of Genetics and Cytology, Yerevan State University, Yerevan, Armenia 2Institute of Molecular Biology, National Academy of Sciences, Yerevan, Armenia
3Institute of Human Genetics, Jena University Hospital, Friedrich Schiller University, Jena, Germany
Aflatoxin B1 (AFB1) is a mycotoxin produced by Aspergillus spec. The latter are worldwide contaminants of food with mutagenic and carcinogenic activities in animals and humans. AFB1 was shown to have deleterious effects on metabolism of eukaryotes in many model systems, including the ability to inhibit DNA replication. An agent that disturbs DNA replication may also have the potential to induce de novo DNA copy number variations (CNVs). Blood samples of three clinically healthy carriers were treated in vitro with AFB1 and chromosome preparations were subjected to parental origin determination fluorescence in situ hybridization (pod-FISH). Probes able to visualize CNVs in 8p21.2 and 15q11.2 were applied. In this setting here for the first time an influence of AFB1 on molecular-cytoge-netically detectable CNVs could be shown. The obtained results indicate that: (i) pod-FISH is a single cell directed, sensitive and suitable method for the analysis of mutagen induced CNVs, (ii) AFB1 has the potential to induce in vitro instability of known CNVs in human leukocytes. This research was supported by MES BMBF (grant number 12GE-004) and partially by State Committee of Science of RA (grant number 14A-1f16).
PARENTAL ORIGIN OF MULTIPLE SMALL SUPERNUMERARY MARKER CHROMOSOMES: EVIDENCE FROM A RARE PRENATAL AND POSTNATAL CASE
Hochstenbach R.1, Nowakowska B.2, Volleth M.3, Ummels A.1, Kutkowska-Kazmierczak A.2, Obersztyn E.2, Ziemkiewicz K.2, Schanze D.3, Zenker M.3, Muschk P.3, Schanze I.3, GerloffC.4, Poot M.1, Liehr T.5 1Department of Medical Genetics, University Medical Centre Utrecht, Utrecht, The Netherlands 2Department of Medical Genetics, Institute of the Mother and Child, ul. Kasprzaka, Warsaw, Poland 3Department of Human Genetics, Otto-von-Guericke University, Magdeburg, Germany
4Universitätsfrauenklinik, Otto-von-Guericke Universität, Magdeburg, Germany
5Universitätsklinikum Jena, Institut für Humangenetik, Jena, Germany
We present two novel cases with different combinations of de novo multiple supernumerary marker chromosomes (sSMCs), each derived from a different chromosome. We determined the gene content, contribution to the clinical phenotype and parental origin of the sSMCs using karyo-typing, fluorescence in situ hybridization, and oligonucle-
РОССИЙСКИЙ ВЕСТНИКПЕРИНАТОЛОГИИ И ПЕДИАТРИИ, 4, 2015
