Marker Of Chronic Myeloid Leukemia Research Articles

Dasatinib in chronic myeloid leukemia: a review

Deregulated BCR-ABL tyrosine kinase (TK) activity is the molecular marker for chronic myeloid leukemia (CML), which provides an identifiable target for developing therapeutic agents. Imatinib mesylate, a BCR-ABL TK inhibitor, is the frontline therapy for CML. Despite the stunning efficacy of this agent, a small number of patients develop a suboptimal response or resistance to imatinib. In newly diagnosed patients with chronic phase CML, the rate of resistance to imatinib at 4 years was up to 20%, increasing to 70% to 90% for patients in the accelerated/blastic phase. Resistance to imatinib led to the development of novel TK inhibitors such as dasatinib. Several clinical trials have reported more durable complete hematologic and cytogenetic responses with this agent in patients who are resistant or intolerant to imatinib. Dasatinib is well tolerated and has broad efficacy, resulting in durable responses in patients with any BCR-ABL mutation except for T3151 and mutations in codon 317 - most commonly F317L - including mutations that were highly resistant to imatinib, such as L248, Y253, E255, F359, and H396. Dasatinib is recommended for CML in chronic, blastic or accelerated phase that is resistant or intolerant to imatinib. Dasatinib was approved by the FDA at 100 mg once daily as the starting dose in patients with chronic phase CML and at 70 mg twice daily in patients with accelerated or blastic phase CML. Various clinical trial results provided evidence that resistance to one TK inhibitor can be reversed with the use of a different TK inhibitor (TKI). Other second-generation TKIs with activity in CML include nilotinib, bosutinib and INNO 406. New molecules, such as the inhibitor of Aurora family serine-threonine kinases, MK0457, which has antileukemic activity in CML associated with a T315I mutation, are being investigated. Allogeneic hematopoietic stem cell transplantation remains an option for selected patients.

Detection of bcr-abl gene expression at a low level in blood cells of some patients with essential thrombocythemia

The major bcr-abl fusion gene is seen as a major marker of chronic myeloid leukemia (CML). However, whether the bcr-abl transcript can be detected in patients with essential thrombocythemia (ET) is still a matter of controversy. We detected the messenger RNA expression of the bcr-abl gene using reverse transcription–polymerase chain reaction in peripheral-blood leukocytes (PBLs) from 63 patients with myeloproliferative disorders (including CML, ET, and polycythemia vera [PV]) and 51 normal, healthy volunteers. The bcr-abl transcript was detected in 4 of the 30 ET patients (13.3%), 17 of the 17 CML patients (100%), none of the 16 PV patients (0%), and 1 of the 51 normal subjects (1.9%). Compared with the normal controls, ET patients have a greater tendency to express the bcr-abl transcript in PBLs ( P = .06, Fisher's exact test). Further semiquantitative analysis showed that the intensity of bcr-abl transcript expression in 4 ET patients and a normal individual was 10 3 to 10 4 times less than that in the CML patients. We conclude that the bcr-abl transcript can be detected in the PBLs of Philadelphia chromosome (Ph)-negative ET patients but that the level of expression is markedly less than that in CML patients. The clinical significance of this finding merits further investigation.

Simple competitive two-step RT-PCR assay to monitor minimal residual disease in CML patients after bone marrow transplantation.

Quantitative reverse transcription-polymerase chain reaction (Q-RT-PCR) assessing the amount of transcripts of the BCR/ABL gene, the molecular marker of chronic myeloid leukemia (CML), is the only method sensitive enough for monitoring of minimal residual disease (MRD) in CML patients after bone marrow transplantation (BMT). In this study we present a simple modification of competitive Q-RT-PCR using natural competitors from cell lines K562 and BV173. The competitors were used in the form of unpurified RNA in cell lysates which ensured their high stability. Mixing competitors and samples before RNA extraction eliminated problems with quantification of cDNA or RNA entering the competitive reaction and with checking for the RNA quality and reverse transcription (RT) efficiency. The bulk of the malignant clone was expressed as the number of leukemic cells in 10(6) leukocytes when the overproduction of the BCR/ABL mRNA in the cell lines we used as competitors was taken into account. It was found to be 82-fold and 14-fold in K562 and BV173, respectively, in comparison with 100% Ph-positive CML standard. The assay reliability was verified by comparison of results with the mathematical model of competitive PCR. The assay is highly reproducible and sensitive (10(-5)). Its accuracy was proved to be excellent in a wide range of malignant cell concentrations. The method is demonstrated on three CML patients suffering from MRD after BMT. In conclusion, this method fulfills all criteria of competitive Q-RT-PCR. Because of its simplicity it is suitable for clinical laboratories and due to the high stability of the lysates used it may serve in the standardization of results between different laboratories.

Properties of the K562 cell line, derived from a patient with chronic myeloid leukemia.

The K562 cell line derived from a CML patient in blast crisis was examined for properties of B and T lymphocytes and cell lines. K562 lacks the B markers of immunoglobulins, Epstein-Barr virus (EBV) genome and associated nuclear antigen, and receptors for EBV. A low proportion of cells from rosettes with sheep erythrocytes, the frequency of which is considerably increased after neuraminidase treatment. Unlike B lines but like T lines, K562 cells are lysed rapidly by C'/Fc receptor-positive human blood leukocytes and do not stimulate MLC reactions. On the other hand, K562 lacks T antigen, high radiosensitivity and sensitivity to growth inhibition by thymidine. The cells do not contain N-APase, an enzyme found in all lines derived from lymphoid cells and in lymphoproliferative diseases. By scanning electron microscopy, K562 cells were seen to be rounded and relatively smooth, with small numbers of short microvilli resembling undifferentiated leukemic cells. A few cells had narrow ridge-like profiles and small ruffles similar to granulocytic leukemic cells. K562 is strongly positive for immunoglobuln Fc receptors and pinocytosis, but does not phagocytose or mediate antibody-dependent phagocytosis or cytolysis. Among histochemical stains, K562 is positive for esterase, lipid, and acid phosphatase. There seems to be no doubt that K562 is not a B cell line. While it has some T cell properties, these are not exclusive. Some of its characteristics indicate that it is probably not lymphoid. Due to its low level of differentiation, its nature cannot be stated with certainty. On the basis of the possible presence of the cellular marker of chronic myeloid leukemia, the Ph chromosome, it may be regarded as belonging to the granulocytic series of cells.