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Genomic imbalances are confined to non-proliferating cells in paediatric patients with acute myeloid leukaemia and a normal or incomplete karyotype.

Ballabio E, Regan R, Garimberti E, Harbott J, Bradtke J, Teigler-Schlegel A, Biondi A, Cazzaniga G, Giudici G, Wainscoat JS, Boultwood J, Bridger JM, Knight SJ, Tosi S - PLoS ONE (2011)

Bottom Line: Leukaemia is often associated with genetic alterations such as translocations, amplifications and deletions, and recurrent chromosome abnormalities are used as markers of diagnostic and prognostic relevance.The majority of these copy number alterations (CNAs) were confirmed by FISH and found to localize to the interphase rather than metaphase nuclei.Interestingly, these experiments showed that, in the vast majority of cases, the changes appeared to be confined to interphase nuclei in a non-proliferative status.

View Article: PubMed Central - PubMed

Affiliation: Centre for Cell and Chromosome Biology and Brunel Institute of Cancer Genetics and Pharmacogenomics, Division of Biosciences, Brunel University, West London, United Kingdom.

ABSTRACT
Leukaemia is often associated with genetic alterations such as translocations, amplifications and deletions, and recurrent chromosome abnormalities are used as markers of diagnostic and prognostic relevance. However, a proportion of acute myeloid leukaemia (AML) cases have an apparently normal karyotype despite comprehensive cytogenetic analysis. Based on conventional cytogenetic analysis of banded chromosomes, we selected a series of 23 paediatric patients with acute myeloid leukaemia and performed whole genome array comparative genome hybridization (aCGH) using DNA samples derived from the same patients. Imbalances involving large chromosomal regions or entire chromosomes were detected by aCGH in seven of the patients studied. Results were validated by fluorescence in situ hybridization (FISH) to both interphase nuclei and metaphase chromosomes using appropriate bacterial artificial chromosome (BAC) probes. The majority of these copy number alterations (CNAs) were confirmed by FISH and found to localize to the interphase rather than metaphase nuclei. Furthermore, the proliferative states of the cells analyzed by FISH were tested by immunofluorescence using an antibody against the proliferation marker pKi67. Interestingly, these experiments showed that, in the vast majority of cases, the changes appeared to be confined to interphase nuclei in a non-proliferative status.

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Representative FISH images confirming CNAs in the nuclei of patient samples.(A) Dual-colour FISH signals on nucleus of patient no. 1 confirm trisomy 7 and trisomy 8 in the same clone. Green signals correspond to chromosome 7 centromere, and red signals correspond to chromosome 8 centromere. (B) Trisomy 19 is shown on a nucleus of patient no, 2 using probe RP11-197O4 (hybridization signals in red). (C) Three red signals corresponding to probe RP11-399H11confirm the presence of three copies of 9q in patient no. 5. (D, E and F) FISH performed on patient no. 11 confirmed: (D) a monosomy of 7q22 as shown by the presence of one red hybridization signal corresponding to probe RP11-193I7; (E) a trisomy of 8q24.13 as shown by the presence of three red signals corresponding to probe RP11-16G11 and (F) a monosomy of 16p12 as shown by one red signals corresponding to probe CTD-2515A14. (G) In patient no. 12, monosomy of 15q21.1 is shown by one red signal corresponding to probe RP11-485O10 (H) Dual-colour FISH on a nucleus from patient no. 19 confirms the presence of a trisomy for 13q32.2 (three red signals corresponding to probe RP11-383H17) and monosomy of 4q35.2 (one green signal corresponding to RP11-45F23) in the same leukaemic clone.
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pone-0020607-g002: Representative FISH images confirming CNAs in the nuclei of patient samples.(A) Dual-colour FISH signals on nucleus of patient no. 1 confirm trisomy 7 and trisomy 8 in the same clone. Green signals correspond to chromosome 7 centromere, and red signals correspond to chromosome 8 centromere. (B) Trisomy 19 is shown on a nucleus of patient no, 2 using probe RP11-197O4 (hybridization signals in red). (C) Three red signals corresponding to probe RP11-399H11confirm the presence of three copies of 9q in patient no. 5. (D, E and F) FISH performed on patient no. 11 confirmed: (D) a monosomy of 7q22 as shown by the presence of one red hybridization signal corresponding to probe RP11-193I7; (E) a trisomy of 8q24.13 as shown by the presence of three red signals corresponding to probe RP11-16G11 and (F) a monosomy of 16p12 as shown by one red signals corresponding to probe CTD-2515A14. (G) In patient no. 12, monosomy of 15q21.1 is shown by one red signal corresponding to probe RP11-485O10 (H) Dual-colour FISH on a nucleus from patient no. 19 confirms the presence of a trisomy for 13q32.2 (three red signals corresponding to probe RP11-383H17) and monosomy of 4q35.2 (one green signal corresponding to RP11-45F23) in the same leukaemic clone.

Mentions: Ten of the putative CNAs identified by aCGH were investigated further by FISH, using probes specifically targeted to the chromosomes involved in the abnormalities. Importantly, the cell preparations used for FISH were derived from samples obtained at the same time as those used for DNA extraction for the aCGH experiments. The BAC clones used for FISH are listed in Table 2 together with a summary of the results showing that ten putative CNAs were verified by FISH. Examples of FISH images obtained using samples from patients no. 1, 2, 5, 11, 12 and 19 are shown in Figure 2.


Genomic imbalances are confined to non-proliferating cells in paediatric patients with acute myeloid leukaemia and a normal or incomplete karyotype.

Ballabio E, Regan R, Garimberti E, Harbott J, Bradtke J, Teigler-Schlegel A, Biondi A, Cazzaniga G, Giudici G, Wainscoat JS, Boultwood J, Bridger JM, Knight SJ, Tosi S - PLoS ONE (2011)

Representative FISH images confirming CNAs in the nuclei of patient samples.(A) Dual-colour FISH signals on nucleus of patient no. 1 confirm trisomy 7 and trisomy 8 in the same clone. Green signals correspond to chromosome 7 centromere, and red signals correspond to chromosome 8 centromere. (B) Trisomy 19 is shown on a nucleus of patient no, 2 using probe RP11-197O4 (hybridization signals in red). (C) Three red signals corresponding to probe RP11-399H11confirm the presence of three copies of 9q in patient no. 5. (D, E and F) FISH performed on patient no. 11 confirmed: (D) a monosomy of 7q22 as shown by the presence of one red hybridization signal corresponding to probe RP11-193I7; (E) a trisomy of 8q24.13 as shown by the presence of three red signals corresponding to probe RP11-16G11 and (F) a monosomy of 16p12 as shown by one red signals corresponding to probe CTD-2515A14. (G) In patient no. 12, monosomy of 15q21.1 is shown by one red signal corresponding to probe RP11-485O10 (H) Dual-colour FISH on a nucleus from patient no. 19 confirms the presence of a trisomy for 13q32.2 (three red signals corresponding to probe RP11-383H17) and monosomy of 4q35.2 (one green signal corresponding to RP11-45F23) in the same leukaemic clone.
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Related In: Results  -  Collection

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pone-0020607-g002: Representative FISH images confirming CNAs in the nuclei of patient samples.(A) Dual-colour FISH signals on nucleus of patient no. 1 confirm trisomy 7 and trisomy 8 in the same clone. Green signals correspond to chromosome 7 centromere, and red signals correspond to chromosome 8 centromere. (B) Trisomy 19 is shown on a nucleus of patient no, 2 using probe RP11-197O4 (hybridization signals in red). (C) Three red signals corresponding to probe RP11-399H11confirm the presence of three copies of 9q in patient no. 5. (D, E and F) FISH performed on patient no. 11 confirmed: (D) a monosomy of 7q22 as shown by the presence of one red hybridization signal corresponding to probe RP11-193I7; (E) a trisomy of 8q24.13 as shown by the presence of three red signals corresponding to probe RP11-16G11 and (F) a monosomy of 16p12 as shown by one red signals corresponding to probe CTD-2515A14. (G) In patient no. 12, monosomy of 15q21.1 is shown by one red signal corresponding to probe RP11-485O10 (H) Dual-colour FISH on a nucleus from patient no. 19 confirms the presence of a trisomy for 13q32.2 (three red signals corresponding to probe RP11-383H17) and monosomy of 4q35.2 (one green signal corresponding to RP11-45F23) in the same leukaemic clone.
Mentions: Ten of the putative CNAs identified by aCGH were investigated further by FISH, using probes specifically targeted to the chromosomes involved in the abnormalities. Importantly, the cell preparations used for FISH were derived from samples obtained at the same time as those used for DNA extraction for the aCGH experiments. The BAC clones used for FISH are listed in Table 2 together with a summary of the results showing that ten putative CNAs were verified by FISH. Examples of FISH images obtained using samples from patients no. 1, 2, 5, 11, 12 and 19 are shown in Figure 2.

Bottom Line: Leukaemia is often associated with genetic alterations such as translocations, amplifications and deletions, and recurrent chromosome abnormalities are used as markers of diagnostic and prognostic relevance.The majority of these copy number alterations (CNAs) were confirmed by FISH and found to localize to the interphase rather than metaphase nuclei.Interestingly, these experiments showed that, in the vast majority of cases, the changes appeared to be confined to interphase nuclei in a non-proliferative status.

View Article: PubMed Central - PubMed

Affiliation: Centre for Cell and Chromosome Biology and Brunel Institute of Cancer Genetics and Pharmacogenomics, Division of Biosciences, Brunel University, West London, United Kingdom.

ABSTRACT
Leukaemia is often associated with genetic alterations such as translocations, amplifications and deletions, and recurrent chromosome abnormalities are used as markers of diagnostic and prognostic relevance. However, a proportion of acute myeloid leukaemia (AML) cases have an apparently normal karyotype despite comprehensive cytogenetic analysis. Based on conventional cytogenetic analysis of banded chromosomes, we selected a series of 23 paediatric patients with acute myeloid leukaemia and performed whole genome array comparative genome hybridization (aCGH) using DNA samples derived from the same patients. Imbalances involving large chromosomal regions or entire chromosomes were detected by aCGH in seven of the patients studied. Results were validated by fluorescence in situ hybridization (FISH) to both interphase nuclei and metaphase chromosomes using appropriate bacterial artificial chromosome (BAC) probes. The majority of these copy number alterations (CNAs) were confirmed by FISH and found to localize to the interphase rather than metaphase nuclei. Furthermore, the proliferative states of the cells analyzed by FISH were tested by immunofluorescence using an antibody against the proliferation marker pKi67. Interestingly, these experiments showed that, in the vast majority of cases, the changes appeared to be confined to interphase nuclei in a non-proliferative status.

Show MeSH
Related in: MedlinePlus