Telomere aggregates in non-Hodgkin lymphoma patients at different disease stages.

Telomeres of tumor nuclei tend to form aggregates (TA). The same phenomenon was also observed in premalignant states. The aim of this study was to estimate TA formation in leukocytes of patients with non-Hodgkin lymphoma (NHL) at different disease stages (diagnosis, treatment, relapse, and remission). The peptide nucleic acid Telomere Kit was used to evaluate TA formation, using two-dimensional fluorescence microscopy. A higher rate of TA was found in all the NHL stages (including remission) than in the control group. Significantly higher TA formation was also observed in the relapse group, compared to the diagnosis group. It may be possible that patients with higher TA numbers are prone to relapse. From our previous results involving replication pattern, random aneuploidy rate, and (recently) TA formation, it can be concluded that the patients in remission are at higher risk of developing relapse than the normal population throughout their life span. The genetic instability parameters remain in the cells of these patients, who must continue to be monitored throughout their life.

Random aneuploidy in CML patients at diagnosis and under imatinib treatment.

Chronic myeloid leukemia (CML) is characterized by the presence of a BCR-ABL fusion gene, which is the result of a reciprocal translocation between chromosomes 9 and 22, and is cytogenetically visible as a shortened chromosome 22 (Philadelphia). Research during the past two decades has established that BCR-ABL is probably the pathogenetic pathway leading to CML, and that constitutive tyrosine kinase activity is central to BCR-ABL capacity to transform hematopoietic cells in vitro and in vivo. The tyrosine kinase inhibitor imatinib mesylate was introduced into the treatment regimen for CML in 1998. During the last few years, reports on chromosomal changes during imatinib treatment have been described. In this study, we evaluated the random aneuploidy rate with chromosomes 9 and 18 in bone marrow from treated and untreated patients. We found higher aneuploidy rates in both treated and untreated patients compared to the control group. In three patients who were treated with imatinib mesylate for more than 1.5 years, triploidy also appeared in some nuclei. To our knowledge, this is the first report on new chromosomal changes such as random aneuploidy and triploidy under imatinib treatment, but more studies are needed to investigate the long-term effect of the imatinib treatment on genetic instability.

The influence of different chromosomal aberrations on molecular cytogenetic parameters in chronic lymphocytic leukemia.

B-cell chronic lymphocytic leukemia (B-CLL) is the most common leukemia of adults in Western countries. The most frequent recurring chromosomal aberrations identified in B-CLL patients are trisomy 12 and deletions of 13q, 17p, and 11q. Cases with deletions of 11q and 17p have a poor prognosis, whereas cases with deletions in 13q have a favorable prognosis. It was previously shown that CLL patients with trisomy 12 and del(13)(q14) have a higher rate of asynchronous replication of normal structural genes when compared to those with normal karyotypes. We studied the replication pattern of the structural locus 21q22 and the imprinted gene SNRPN and its telomere (15qter) and the random aneuploidy of chromosomes 9 and 18 in CLL patients with trisomy 12 and deletions of 11q and 17p, and compared the results to those of CLL patients without these aberrations and to healthy controls. Random aneuploidy rate was higher in the group of patients with trisomy 12 as compared to all other groups. The replication pattern with higher asynchronous pattern was found in both aberration groups compared to the CLL patients without the aberrations and to the control group with involvement of 21q22 and 15qter, whereas the highest synchronous group was found in the 2 aberrations CLL patient groups compared to the other groups with the imprinted locus SNRPN. The existence and significance of chromosomal aberrations in CLL have a deleterious effect on the processes of cell cycle and gene replication and may have biological and prognostic implications.

Random aneuploidy and telomere capture in chronic lymphocytic leukemia and chronic myeloid leukemia patients.

Telomeric regions of the human genome are of particular interest, because rearrangements of these regions are difficult to identify by conventional chromosome banding technology. With the advent of molecular cytogenetic techniques such as fluorescence in situ hybridization (FISH), it has been possible to investigate the terminus in cytogenetically visible terminal deletions and telomere rearrangements. We investigated telomere capture and aneuploidy rates in chronic lymphocytic leukemia (CLL) and chronic myeloid leukemia (CML) patients, as well as in healthy control subsets. Using a FISH technique, we estimated the random aneuploidy and telomere capture of the 21q22, SNRPN, and 15qter loci. Higher aneuploidy rates were found in the leukocytes of CLL and CML patients, compared with the control group, for the 21q22 and SNRPN loci. There was no difference in the aneuploidy rate between the CML and CLL groups. Telomere capture was found in the two groups (CLL and CML), but not in the control group. We propose that the telomere capture phenomenon is much more common than has been reported in the literature; however, its prognostic significance is yet to be established.

Random aneuploidy in neoplastic and pre-neoplastic diseases, multiple myeloma, and monoclonal gammopathy.

In this study we evaluated the aneuploidy rate of cells from patients considered to have a premalignant condition (monoclonal gammopathy or MGUS) and patients with multiple myeloma, as well as healthy controls. By applying a fluorescence situ hybridization technique, we estimated the random aneuploidy rate of alpha-satellite (centromeres) probes from chromosomes 9 and 18. The monosomy and total aneuploidy rates were higher in the two study groups compared to the control group. The monosomy rate was significantly higher in the MGUS group compared to the group with chromosome 18 alpha-satellite probes, a finding that was reported before in preneoplastic conditions. Our results support the cancer aneuploidy theory that carcinogenesis is initiated by a random aneuploidy, which is induced either spontaneously or by a carcinogen. The resulting karyotype instability sets a chain reaction of aneuploidization, which generates even more abnormal and eventually cancer-specific combinations and rearrangements of chromosomes.

Deletion of 5q31 and 7q31 in patients with stable melphalan treated multiple myeloma.

Multiple myeloma represents a malignant proliferation of plasma cells derived from a single clone. It is well known that alkylating agents are capable of inducing myelodysplastic syndromes (MDS) and acute myelocytic leukemias (AML). This risk of both diseases in patients with multiple myeloma has been estimated to be 10-20% after 10 years. We aimed to evaluate the time course and the type of genetic abnormalities in melphalan-treated patients in the chronic stage of the disease. We applied fluorescence in situ hybridization methods with probes to 5q31 and 7q31 to mononuclear peripheral blood leukocytes of 18 melphalan-treated patients and compared the results to those of 8 untreated myeloma patients. We found three patients (17%) with a 5q31 deletion in their peripheral white blood cells, but no 7q31 deletion. These findings suggest that 5q- occurs before the overt development of MDS/AML and raise important concerns regarding long-term treatment of myeloma patients with alkylating agents. Also, the performance of cytogenetic evaluation should be considered before autologous transplantation. The clinical and biological implications of these findings should be evaluated in larger clinical and laboratory studies.

The effect of chlorambucil treatment on cytogenetic parameters in chronic lymphocytic leukemia patients.

The most common treatment of chronic lymphocytic leukemia (CLL) is the alkylating agent chlorambucil (CLB), with or without prednisone. In the present study, our aim was to evaluate whether treatment with CLB for more than one year induced genetic changes manifested by comparative genomic hybridization (CGH) as new chromosomal aberrations. We also studied whether CLB affected the pattern of replication by using fluorescence in situ hybridization (FISH). We found a similar rate of asynchronous pattern of replication in both treated and untreated patients with CLL. Most of the aberrations found with CGH were previously reported in CLL. More prognostically unfavorable aberrations and more cases with genetic changes were found in the treated group. The changes found were not typical of the secondary genetic aberrations associated with alkylating agents. Thus, we conclude that treatment of CLL with CLB for at least a year does not affect the parameters analyzed in this study. Longer studies are needed to further explore the effects of alkylating agents on normal and malignant cells.