We report on a child with mild mental retardation, hypotelorism, blepharophimosis, face slight asymmetry and partial hypoplasia of corpus callosum, with an interstitial deletion of a chromosome 15. The deletion was molecularly characterized by array-CGH and FISH techniques. This rearrangement has a 7.18Mb extension and maps to 15q21.2q22.1. To date, there have been only six individuals reported with a deletion of 15q21; in three cases, the rearrangement was characterized by molecular cytogenetic techniques. After a comparison with these three cases, it appeared that the deletion we found is one of the smallest and it overlaps the distal portion of the ones taken into account. Finally, we tried to delineate the genotype–phenotype correlation in patients with a deletion of 15q21.

BACKGROUND In virtually all Ph1 chromosome-positive CML patients, the breakpoint on chromosome 22 maps in a very restricted area of 5.8 Kb, which has been named "breakpoint cluster region" or "bcr". Several molecular probes of this region are presently available, and this makes the molecular diagnosis of CML a useful approach which can be particularly important in those cases in which cytogenetic analysis does not reveal the presence of a Ph1 chromosome. Here we report the problems and our experience during the molecular analysis of the 478 patients examined so far. METHODS Molecular analyses were performed after digestion of the DNA with 2 to 4 restriction enzymes and hybridization with different probes. Individual samples were subjected to PCR since no rearrangements had been obtained with Southern blotting. RESULTS Rearrangement bands were detected in all the samples examined. In 473 cases the breakpoint was located within the bcr. In one of these cases, it was detected only after PCR analysis, and in two cases only after the use of the PHL/BCR probe. In 5 cases the breakpoint was localized either 5' or 3' with respect to the bcr. CONCLUSIONS In this paper, the criteria for a correct molecular diagnosis of CML are presented. The "PHL/BCR" probe appeared to be very specific and time-saving, since it required only one digestion to evidence the rearrangement. Our results confirm the high specificity of the breakpoint on chromosome 22 in CML and the relatively rare incidence of molecular variants.

We report a patient with M2 acute non-lymphocytic leukemia and a complex karyotype: 47,XY,t(5;7)(q34; q21), +8. After chemotherapy with Daunomycin and Arabinosyl Cytosine, a complete remission was reached, but two months later he relapsed and died because of sepsis. Only 5 other cases with translocations involving chromosomes 5 and 7 have been described, but with different breakpoints. Several genes related to cell proliferation and maturation have been identified on the long arms of chromosomes 5 and 7. The possible involvement of specific genes located at or very close to the breakpoints is hypothesized.

Allogeneic bone marrow transplantation is the only way to cure patients with Ph1+ chronic myeloid leukemia. It is commonly assumed that, in order to obtain a cure for the patients, the leukemic clone must be completely destroyed by the conditioning treatment and the donor's bone marrow must repopulate the hemopoietic niches leading to a “complete chimera”. However, cytogenetic analyses, supported by molecular ones, indicate that Ph1+ cells, far from being completely destroyed by chemo-radiotherapy may persist for a long time, probably in the majority of the patients. As demonstrated by the outcome of patients receiving T-cell depleted marrow, immune mechanisms must be involved in controlling and progressively reducing the size of the residual leukemic clone. Furthermore, immunomodulating therapeutic strategies, represented by cyclosporin-A discontinuation or alpha interferon treatment, may successfully reduce the Ph1+ cell population even after a full relapse.

Comparative analyses of the leukocyte differential counting were performed using a Coulter VCS Hematology Flow Cytometer and direct microscopic observation on 547 unselected individuals analyzed at the outpatient clinic of the Institute of Hematology "L. e A. Seràgnoli" of Bologna. The Coulter VCS is able to provide leukocyte differential counting by measuring cell volume, conductivity and laser light scatter. Negative (true negative: 50.8%) results were observed in 278 subjects by both automated and direct observation methods. Abnormal cells in the blood samples (true positive results) were detected both by VCS and by direct observation in 199 cases (36.4%). Regional distribution of the cells in specific "flags" on the scatterplot was often associated with specific cell types. In 66 cases (12.1%) no abnormal cell was detected by direct observation, while Coulter VCS gave an abnormal pattern, even if only to a slight extent (false positive cases). In 4 cases only (0.7%), false negative results were given by the VCS system. The correlation of the results given by the VCS system with those given by direct microscopic analysis was very high; however careful control by the operator was essential in evaluating the data given by the automated system and in identifying the type of abnormal cells detected.

A patient with Philadelphia (Ph 1 )‐negative, breakpoint cluster region (bcr)‐positive chronic myeloid leukemia (CML) is reported. Pulsed‐field gel electrophoretic analysis demostrated the comigration of both ABL and BCR sequences on the same Bss 1–111 and Sac 11 fragment. Moreover, in situ hybridization studies demonstrated that ABL sequences had been moved from band 9q34 to 22q11 and that the additional t(12;12)(q13;p12) was not involved in the ABL/BCR related translocation. Neverthless, a possible role of oncogenes or regulatory sequences activated or inhibited by the additional translocation cannot be excluded.

Out of 105 Philadelphia (Ph) positive chronic myeloid leukemia patients analyzed, six (5.7%) carried a variant Ph translocation, namely t(6;9;9;10;22)(q24;p13;q34;p15;q11); t(9;13;22)(q34;q21;q11);der(2)(2pter----2q31::9q21---- 9q34::22q11----22qter) and der(9)t(2;9) (9pter----9q21::2q31----2qter);t(7;9;22)(q11;q34 ;q11), 14q + ;t(7;9;22)(q35;q34;q11), and t(9;11;22) (q34;q13;q11), respectively. Five of these patients were analyzed with Southern blotting. Three of them showed an atypical molecular pattern; namely, the patient with t(9;13;22) showed no rearrangement in the breakpoint cluster region (bcr), the patient with t(7;9;22)(q35;q34;q11) showed a 3' deletion, and the patient with t(7;9;22), 14q + showed a bcr rearrangement 3' to the exon 4 of the M-BCR. Chromosome in situ hybridization studies demonstrated that in patient one, a two-step translocation occurred: the first step moved the 3' bcr from chromosome 22 to chromosome 9, and the second moved the terminal part of 22q, carrying the c-sis protooncogene, to 10p. Variant Ph translocations appear to be associated with atypical molecular breakpoints.

A patient with M2-ANLL and a 46,XX,del(5)(q22q33), t(2;11)(p21;q24) karyotype is described. The diagnosis was made after a short period of myelodysplastic syndrome. After chemotherapy consisting of Daunorubicin and Arabinosylcytosine in continuous infusion, the patient reached a complete remission. The chromosome pattern described here has been observed in two other patients with refractory anemia and refractory anemia with excess of blasts, respectively. The breakpoints on the chromosomes 2, 5 and 11 allow us to hypothesize the involvement of N-myc, c-fms, GM-CSF and IL-3 genes.

British Journal of HaematologyVolume 69, Issue 3 p. 424-426 CYTOGENETIC AND MOLECULAR ANALYSES IN PHILADELPHIA CHROMOSOME POSITIVE ACUTE LYMPHOBLASTIC LEUKAEMIA Alfonso Zaccaria, Alfonso Zaccaria Istituto de Ematologia ‘Lorenzo e Ariosto Seàgnoli’, Universita’di BolognaSearch for more papers by this authorAngela Tassinari, Angela Tassinari Istituto de Ematologia ‘Lorenzo e Ariosto Seàgnoli’, Universita’di BolognaSearch for more papers by this authorGiuseppe Saglio, Giuseppe Saglio Dipartimento di Scienze Biomediche e Oncologia Umana, Istituto di Clinica Medica A, Universita’de Torino, ItalySearch for more papers by this authorAngelo Guerrasio, Angelo Guerrasio Dipartimento di Scienze Biomediche e Oncologia Umana, Istituto di Clinica Medica A, Universita’de Torino, ItalySearch for more papers by this authorNicoletta Testoni, Nicoletta Testoni Istituto de Ematologia ‘Lorenzo e Ariosto Seàgnoli’, Universita’di BolognaSearch for more papers by this authorBommina Celso, Bommina Celso Istituto de Ematologia ‘Lorenzo e Ariosto Seàgnoli’, Universita’di BolognaSearch for more papers by this authorGianantonio Rosti, Gianantonio Rosti Istituto de Ematologia ‘Lorenzo e Ariosto Seàgnoli’, Universita’di BolognaSearch for more papers by this authorSante Tura, Sante Tura Istituto de Ematologia ‘Lorenzo e Ariosto Seàgnoli’, Universita’di BolognaSearch for more papers by this author Alfonso Zaccaria, Alfonso Zaccaria Istituto de Ematologia ‘Lorenzo e Ariosto Seàgnoli’, Universita’di BolognaSearch for more papers by this authorAngela Tassinari, Angela Tassinari Istituto de Ematologia ‘Lorenzo e Ariosto Seàgnoli’, Universita’di BolognaSearch for more papers by this authorGiuseppe Saglio, Giuseppe Saglio Dipartimento di Scienze Biomediche e Oncologia Umana, Istituto di Clinica Medica A, Universita’de Torino, ItalySearch for more papers by this authorAngelo Guerrasio, Angelo Guerrasio Dipartimento di Scienze Biomediche e Oncologia Umana, Istituto di Clinica Medica A, Universita’de Torino, ItalySearch for more papers by this authorNicoletta Testoni, Nicoletta Testoni Istituto de Ematologia ‘Lorenzo e Ariosto Seàgnoli’, Universita’di BolognaSearch for more papers by this authorBommina Celso, Bommina Celso Istituto de Ematologia ‘Lorenzo e Ariosto Seàgnoli’, Universita’di BolognaSearch for more papers by this authorGianantonio Rosti, Gianantonio Rosti Istituto de Ematologia ‘Lorenzo e Ariosto Seàgnoli’, Universita’di BolognaSearch for more papers by this authorSante Tura, Sante Tura Istituto de Ematologia ‘Lorenzo e Ariosto Seàgnoli’, Universita’di BolognaSearch for more papers by this author First published: July 1988 https://doi.org/10.1111/j.1365-2141.1988.tb02389.xCitations: 2AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Citing Literature Volume69, Issue3July 1988Pages 424-426 RelatedInformation

By analyzing a total of 107 patients affected by chronic myelogenous leukemia (CML; chronic and blast crisis) or lymphoid and myeloid Philadelphia chromosome (Ph′) positive acute leukemias, we have investigated the relationship between the molecular defect on the Ph′ chromosome and the associated hematologic phenotype. As expected, approximately half of the Ph′ positive acute leukemias showed a breakpoint on chromosome 22 falling outside the “breakpoint cluster region” (bcr) known to be involved in CML. Surprisingly, seven of 80 CML cases in chronic phase also showed rearrangements falling outside the bcr region. In two of these cases the breakpoint on chromosome 22 was mapped between 9 and 12 kb upstream to the bcr region. In another case, the breakpoint was located approximately 16 kb downstream to bcr. In the remaining four cases, the precise position of the rearrangement could not be localized with the available bcr probes. DNAs from patients with CML blast crises showed classical bcr rearrangements. No molecular changes were observed during the progression of the disease in six patients whose DNA from both a chronic and acute phase was available. Our results seem to indicate a greater degree of variability of chromosome 22 breakpoints in CML than previously observed, and the lack of additional rearrangements on the Ph′ chromosome in CML blast crises with respect to chronic phase.