Kristiina Heinonen

BAALC, the human member of a novel mammalian neuroectoderm gene lineage, is implicated in hematopoiesis and acute leukemia

The molecular basis of human leukemia is heterogeneous. Cytogenetic findings are increasingly associated with molecular abnormalities, some of which are being understood at the functional level. Specific therapies can be developed based on such knowledge. To search for new genes in the acute leukemias, we performed a representational difference analysis. We describe a human gene in chromosome 8q22.3, BAALC (brain and acute leukemia, cytoplasmic), that is highly conserved among mammals but evidently absent from lower organisms. We characterized BAALC on the genomic level and investigated its expression pattern in human and mouse, as well as its complex splicing behavior. In vitro studies of the protein showing its subcellular localization suggest a function in the cytoskeleton network. Two isoforms are specifically expressed in neuroectoderm-derived tissues, but not in tumors or cancer cell lines of nonneural tissue origin. We show that blasts from a subset of patients with acute leukemia greatly overexpress eight different BAALC transcripts, resulting in five protein isoforms. Among patients with acute myeloid leukemia, those overexpressing BAALC show distinctly poor prognosis, pointing to a key role of the BAALC products in leukemia. Our data suggest that BAALC is a gene implicated in both neuroectodermal and hematopoietic cell functions.

Cytogenetic abnormalities in childhood acute myeloid leukaemia: a Nordic series comprising all children enrolled in the NOPHO-93-AML trial between 1993 and 2001

Summary. Between 1993 and 2001, 318 children were diagnosed with acute myeloid leukaemia (AML) in the Nordic countries. The patient group comprised 237 children < 15 years of age with de novo AML, 42 children < 15 years with Down syndrome (DS) and de novo AML, 18 adolescents 15–18 years of age with de novo AML, and 21 children < 15 years with treatment‐related AML (t‐AML). The first group was all‐inclusive, yielding an annual childhood de novo AML incidence of 0·7/100 000. Cytogenetic analyses were successful in 288 cases (91%), and clonal chromosomal abnormalities were detected in 211 (73%). The distribution of ploidy levels were pseudodiploidy (55%), hyperdiploidy (34%) and hypodiploidy (11%). The most common aberrations (> 2%) were + 8 (23%) (as a sole change in 6·2%), 11q23‐translocations, including cryptic MLL rearrangements (22%) [t(9;11)(p21–22;q23) in 11%], t(8;21)(q22;q22) (9·0%), inv(16)(p13q22) (6·2%), −7/7q– (5·2%), and t(15;17)(q22;q12) (3·8%). Except for +8, these abnormalities were rare in group 2; only one DS patient had a t(8;21) and none had 11q23‐translocations, t(15;17) or inv(16). In the t‐AML group, three cases displayed 11q23‐rearrangements, all t(9;11); and there were no t(8;21), t(15;17) or inv(16). Overall, the observed frequencies of t(8;21) and t(15;17) were lower, and frequencies of trisomy 8 and 11q23‐translocations higher, than in previous studies. Furthermore, seven abnormalities that were previously reported as only single AML cases were also seen, meaning that der(4)t(4;11)(q26–27;q23), der(6)t(1;6)(q24–25;q27), der(7)t(7;11)(p22;q13), inv(8)(p23q11–12), t(11;17)(p15;q21), der(16)t(10;16)(q22;p13) and der(22)t(1;22)(q21;q13) are now classified as recurrent abnormalities in AML. In addition, 37 novel aberrations were observed, 11 of which were sole anomalies.

Cell lineage involvement in four patients with myelodysplastic syndrome and t(1;7) or trisomy 8 studied by simultaneous immunophenotyping and fluorescence in situ hybridization☆

Four patients with myelodysplastic syndrome (MDS), one with t(1;7) and three with trisomy 8, were studied by immunophenotyping and fluorescence in situ hybridization (FISH) to assess cell lineage involvement. The t(1;7) was detected using a biotin-labeled chromosome 1 centromere-specific DNA probe. This aberration was present in CD34-positive stem cells, the erythroid cell lineage (GPA+), and the granulocytic/monocytic (CD13+ and CD64+) cell lineages. We were not able to demonstrate the abnormality in the lymphoid cell lineages. In the patients with trisomy 8, the aberration was detected with chromosome 8 centromere-specific DNA probe or by chromosome in situ suppression hybridization (CISS) with a chromosome 8-specific library probe. The trisomy was detected in stem cells, erythroid precursor cells, megakaryocytes, and granulocytes/monocytes. In these MDS patients, the chromosome aberrations appear to occur only in cells of myeloid lineage.

Chromosome abnormalities in 16 finnish patients with Burkitt's lymphoma or l3 acute lymphocytic leukemia☆

Eleven patients with Burkitts lymphoma (BL), i.e., small noncleaved non-Hodgkins lymphoma, and 5 patients with Burkitt-type acute lymphocytic leukemia (ALL-L3) were selected for chromosome study. Two of the 16 patients had no B-cell markers, but the erythrocyte marker--glycophorin A--was present on the surface of the leukemic blasts. The critical breakpoint at 8q24 was detected in 14 of the 16 patients, whereas this aberration was not detected in any of the 134 patients belonging to other subgroups of non-Hodgkins lymphoma or ALL that we studied during the same period. In addition to the t(8;14)(q24;q32), the following translocations with the breakpoint at 8q24 were seen: t(2;8)(p11;q24), t(8;11)(q24;q13) in BL, and t(2;8;14)(p11 or p12;q24;q32) in ALL. Additional aberrations seen more than once were trisomy #7 and abnormalities in chromosomes #1, #11, and #13.

High modal number and triple trisomies are highly correlated favorable factors in childhood B-cell precursor high hyperdiploid acute lymphoblastic leukemia treated according to the NOPHO ALL 1992/2000 protocols

Between 1992 and 2008, 713 high hyperdiploid acute lymphoblastic leukemias in children aged 1–15 years were diagnosed and treated according to the Nordic Society for Pediatric Hematology and Oncology acute lymphoblastic leukemia 1992/2000 protocols. Twenty (2.8%) harbored t(1;19), t(9;22), der(11q23), or t(12;21). The median age of patients with “classic” high hyperdiploidy was lower than that of patients with translocation-positive high hyperdiploidy (P<0.001). Cases with triple trisomies (+4, +10, +17), comprising 50%, had higher modal numbers than the triple trisomy-negative cases (P<0.0001). The probabilities of event-free survival and overall survival were lower for those with white blood cell counts ≥50×109/L (P=0.017/P=0.009), ≥5% bone marrow blasts at day 29 (P=0.001/0.002), and for high-risk patients (P<0.001/P=0.003), whereas event-free, but not overall, survival, was higher for cases with gains of chromosomes 4 (P<0.0001), 6 (P<0.003), 17 (P=0.010), 18 (P=0.049), and 22 (P=0.040), triple trisomies (P=0.002), and modal numbers >53/55 (P=0.020/0.024). In multivariate analyses, modal number and triple trisomies were significantly associated with superior event-free survival in separate analyses with age and white blood cell counts. When including both modal numbers and triple trisomies, only low white blood cell counts were significantly associated with superior event-free survival (P=0.009). We conclude that high modal chromosome numbers and triple trisomies are highly correlated prognostic factors and that these two parameters identify the same subgroup of patients characterized by a particularly favorable outcome.

Clinical and cytogenetic features of a population-based consecutive series of 285 pediatric T-cell acute lymphoblastic leukemias: Rare T-cell receptor gene rearrangements are associated with poor outcome.

Clinical characteristics and cytogenetic aberrations were ascertained and reviewed in a population‐based consecutive series of 285 pediatric T‐cell acute lymphoblastic leukemias (T‐ALLs) diagnosed between 1992 and 2006 in the Nordic countries. Informative karyotypic results were obtained in 249 (87%) cases, of which 119 (48%) were cytogenetically abnormal. Most (62%) of the aberrant T‐ALLs were pseudodiploid. Structural changes were more common than numerical ones; 86% displayed at least one structural abnormality and 41% at least one numerical anomaly. The most frequent abnormalities were T‐cell receptor (TCR) gene rearrangements (20%) [TCR;11p13 (10%), TCR;10q24 (3%), TCR;other (8%)], del(9p) (17%), +8 (14%), del(6q) (12%), and 11q23 rearrangements (6%). The TCR;other group comprised the rare rearrangements t(X;14)(p11;q11), t(X;7)(q22;q34), t(1;14)(p32;q11), ins(14;5)(q11;q?q?), inv(7)(p15q34), t(8;14)(q24;q11), t(7;11)(q34;p15), and t(12;14)(p13;q11). The clinical characteristics of this Nordic patient cohort agreed well with previous larger series, with a median age of 9.0 years, male predominance (male/female ratio 3.1), median white blood cell (WBC) count of 66.5 × 109/l, and a high incidence of mediastinal mass and central nervous system involvement (59% and 9.5%, respectively). These features did not differ significantly among the various genetic subgroups. 5‐year event‐free survival (EFS) and overall survival for all patients were 0.61 (±0.03) and 0.67 (±0.03), respectively. In a multivariate analysis, two factors affected negatively the EFS, namely a WBC count of ≥200 × 109/l (P < 0.001) and the presence of rare TCR rearrangements (P = 0.001). In conclusion, in this large series of childhood T‐ALLs from the Nordic countries, the cytogenetic findings were not associated with risk of therapy failure with the exception of the TCR;other group. However, further prospective and collaborative investigations of this genetically heterogeneous entity are needed to confirm these results.

Multiple karyotypic abnormalities in three cases of small cell variant of T-cell prolymphocytic leukemia

Cytogenetic, clinical, and laboratory findings of three patients with a small cell variant of T-cell prolymphocytic leukemia (T-PLL) are presented. Immunophenotypic studies of the morphologically typical small cell variant prolymphocytes showed a mature helper T-cell phenotype (CD4+CD8-) in one patient and a common thymocyte phenotype (CD4+ CD8+) in two other patients. The cytogenetic analysis revealed complex karyotypes with several structural aberrations in the peripheral blood lymphocytes of all three patients. In all cases chromosome 14 was affected with the breakpoint at 14q11. Inversion (14) and isochromosome 8q, often reported as an additional aberration in T-PLL, were detected in two of the patients. In two patients a translocation of the short arm of chromosome 12 was also seen. The T-cell receptor beta-chain gene showed a clonal rearrangement in all three patients, whereas no rearrangements were detected in the immunoglobulin genes. The survival of the patients ranged from 10 weeks to 48 months. The association between cytogenetic, clinical, and laboratory data is discussed.

Acquired X-chromosome aneuploidy in children with acute lymphoblastic leukemia

BACKGROUND A cytogenetic study of 75 consecutive children with ALL revealed a normal karyotype, a low hyperdiploid karyotype (including 47-50 chromosomes), and a high hyperdiploid karyotype (including > 50 chromosomes) in 10, 12, and 33 patients, respectively. An acquired extra X-chromosome was detected at diagnosis by conventional cytogenetics in 29 (88%) of 33 children with a high hyperdiploid karyotype and in 4 (33%) of 12 children with a low hyperdiploid karyotype. X-chromosome aneuploidy was retrospectively studied by fluorescence in situ hybridization (FISH) in eight and 20 patients with a normal and a hyperdiploid karyotype, respectively. PROCEDURE A classical cytogenetic study was performed according to standard methods. FISH with the centromeric probe specific to X-chromosome was used to study interphase cells of bone marrow or blood samples. RESULTS An extra X-chromosome was found by FISH in all 13 patients with a high hyperdiploid or tetraploid, in 6 of 7 patients with a low hyperdiploid, and in none with a normal karyotype. Two children with a normal karyotype displayed monosomy X. Altogether, 57.3% of newly diagnosed children displayed X-chromosome aneuploidy. CONCLUSIONS Out study indicates that X-chromosome aneuploidy may be the most common chromosome abnormality in childhood ALL. It can be detected in nearly all children with a high hyperdiploid karyotype and up to one-half of the patients with a low hyperdiploid karyotype. FISH with an X-chromosome centromeric probe is a rapid and simple tool to detect an abnormal clone at diagnosis in the majority of children with ALL and is useful in confirming remission in these patients.

Analysis of phenotype and genotype of individual cells in neoplasms

The authors describe a combination technique enabling detection of in situ hybridization (ISH) signals from chromosome-specific probes in interphase or mitotic cells that still retain the alkaline phosphatase antialkaline phosphatase (APAAP) or Sudan black B (SBB) staining reactions (simultaneous detection) or have been first classified morphologically and then by APAAP or SBB. The technique can be used on cell suspensions, in situ cultures and tissue sections. Examples from leukemias (chronic lymphocytic, myeloid, and acute myeloid leukemia) and solid tumors (chondromyxoid fibroma and glioblastoma) illustrate the potential of the technique in investigation of cancer tissue heterogeneity. In leukemias, it can be used to study cell lineage involvement, stem cells, and minimal residual disease, as well as to monitor therapy. In solid tumors, it can be used to identify neoplastic areas of tissue and to track the site of origin of neoplastic cells. Finally, it can be used to study the significance of chromosome abnormalities in carcinogenesis.

Detection of numerical chromosome abnormalities by FISH in childhood acute lymphoblastic leukemia

Fluorescence in situ hybridization (FISH) was performed on interphase bone marrow cells to study numerical chromosome abnormalities in childhood acute lymphoblastic leukemia (ALL). Ten patients were selected for this study on the basis of having an extra chromosome 6 in the abnormal clone of the bone marrow at diagnosis. The numerical changes that were detected by FISH with a chromosome 6 specific alpha-satellite DNA probe correlated well with the cytogenetic and clinical data in all patients. Three hybridization signals were seen in 43.8--83.0% of interphase cells in the specimens with a hyperdiploid karyotype. The diagnostic bone marrow sample of the patient with a tetraploid karyotype revealed four signals in 67.0% of cells. Two signals were detected in the majority of the cells in the three nonleukemic control bone marrow samples (97.0--97.7%), assessing the cut-off value of about 1% for trisomy 6. This study demonstrates that FISH analysis is a useful and sensitive tool to screen for the presence of extra chromosomes in interphase cells and is important clinically for evaluating the achievement and maintenance of remission in hyperdiploid childhood ALL. However, to detect structural chromosome aberrations which carry important diagnostic and prognostic information, as seen in our patient with the translocation t(1;19) at diagnosis but not at relapse, conventional cytogenetic analysis should be performed both at diagnosis and at relapse.