INTRODUCTION

Chronic Myeloid Leukemia (CML) is a clonal myeloproliferative disorder of the pleuripotent hemopoietic progenitor cell, characterized by excessive proliferation of marrow granulocytes, erythroid precursors, megakaryocytes and connective tissue forming cells. CML is a biphasic or triphasic disease.1 The disease progresses through three distinct phases, chronic phase, accelerated phase and blast crisis.

Allogenic SCT is currently the only curative therapy for CML and when feasible is the treatment of choice. Today the goal of therapy is complete molecular remission and cure. Imatinib (formerly STI-571), a tyrosine kinase inhibitor is a revolution in the treatment of CML.

The hematological remission rate documented is more than 90% and the cytogenetic remission rate is around 60% with major cytogenetic remission rate of upto 30% in patients who are in chronic phase of CML. However, the molecular remission, which is the goal of present day therapy is not well documented. This study besides evaluating hematological response aims to evaluate molecular response to Imatinib in CML patients by the measurement of BCR-ABL transcript level in blood using real time reverse transcriptase quantitative polymerase chain reaction (RT Q-PCR).

METHODS AND MATERIALS

Sixteen patients, thirteen males and three females in the age group of 26- 51 years who were diagnosed as having Chronic Myeloid Leukemia, chronic phase, at Kasturba Medical College Hospital, Attavar, Mangalore, during the period of two years from January 2004 to January 2006 were started on Imatinib mesylate. All the patients were diagnosed on the basis of clinical features, peripheral blood smear (PBS) findings, Bone marrow aspiration (BMA) findings and real time reverse transcriptase quantitative polymerase chain reaction (RT Q-PCR).

The dose of imatinib administered was 400mg/day orally. They were followed closely over a period of 1 year using the following parameters: (1) monthly clinical examination, (2) monthly peripheral blood smear examination, (3) RT Q-PCR for BCR-ABL at the end of every 6 months.

A patient was considered as having responded at the hematological level2 if he/she achieved normal hemoglobin, leucocyte and platelet indices within 3 months of starting treatment with imatinib and the patient was free of all signs and symptoms.

Real time RT Q-PCR provides an accurate, sensitive and non-invasive method of measurement of residual leukemia. BCR-ABL transcript quantification is expressed as percentage of control gene. The sample showing amplification curve for both BCR-ABL target gene and control gene are considered as positive for the test and reported as % expression of BCR-ABL. The sample showing no amplification curve for BCR-ABL target gene but shows amplification curve for control gene is considered as negative for the test and reported as 0.0% expression. In the absence of amplification curve for control gene test is reported as invalid.

The findings were analyzed after the end of one-year therapy with imatinib to know the hematological and molecular response as well as the side effects.

RESULTS

Thirteen (81.25%) out of sixteen patients were males and three (18.75%) females (Table 1). The most common symptoms with which the patients presented were weakness and abdominal discomfort. Other symptoms were abdominal pain, fever and weight loss (Table 1). All the patients, 16 (100%) had moderate to massive splenomegaly at the time of presentation. Ten (62.5%) patients had mild to moderate hepatomegaly at the time of presentation. Eleven (68.75%) patients had pallor at the time of diagnosis (Table 1). All the patients were diagnosed as having chronic phase CML on the basis of PBS findings, BMA findings and real time RT Q-PCR.

The most common side effects were edema and arthralgia. Seven(43.75%) patients complained of arthralgia which occurred mostly during the first two months of therapy and then gradually subsided.

DISCUSSION

CML is a clonal disorder in which cells of myeloid lineage undergo massive clonal expansion. The disease progresses through three distinct phases, chronic phase, accelerated phase and blast crises during which the leukemic clone progressively loses its ability to differentiate. The underlying genetic abnormality is a translocation described by the notation t(9;22)(q34;q11).3 This results in the formation of Philadelphia chromosome (Ph) which is the chromosomal abnormality seen in 95-100% of marrow cell metaphases of >90% of CML patients. The molecular abnormality seen is the BCR-ABL gene. The BCR-ABL fusion gene product is a 210 kDa protein which acts as a constitutively activated tyrosine kinase. It subsequently activates down stream kinases that prevent apoptosis and is responsible for abnormal proliferation of cancer cells. Conventional treatment of CML has been single agent therapy with either busulphan or hydroxyurea, which was developed in 1960. But the disadvantages of these two historical drugs were median survival of less than 3 years with a hematological remission of 80% and no cytogenetic remission. The side effects of prolonged treatment with these agents are skin pigmentation, infertility, and interstitial fibrosis of lung. Allogenic SCT is currently the only curative therapy for CML. When allogenic SCT is not feasible, IFN-a therapy was the treatment of choice before imatinib become available. The BCR-ABL tyrosine kinase is a well-validated therapeutic target in CML. All of the transforming activities of BCR-ABL are dependent on its tyrosine kinase activity. Thus, an inhibitor of the BCR-ABL kinase would be predicted to be an effective and selective therapeutic agent for CML. The treatment guidelines for patients with CML are undergoing substantial change due to emerging evidence regarding the effectiveness of imatinib mesylate. Imatinib induced hematological response in 82% of CML patients.4 The rate of major cytogenetic response was 24% (complete in 17%).4 Orally administered imatinib is an effective and well-tolerated treatment for patients with CML in accelerated phase.4 Imatinib has substantial activity and a favorable safety profile when used as a single agent in patients with CML in blast crisis.5 Complete hematologic responses typically occurred within four weeks after the initiation of therapy.6

All the patients were diagnosed as having chronic phase CML and the dose of imatinib administered was 400mg/day. CML is more common in males. This is evidenced by the fact that there were thirteen males and three females diagnosed and treated in this study. The median age at diagnosis was 34 years. Splenomegaly is a constant finding in CML as all the 16 patients had it at the time of diagnosis. Hepatomegaly and pallor was also present in the majority of patients at the time of diagnosis. Lymphadenopathy, however is not a feature of chronic phase CML as evidenced by the fact that none of the patients had it at the time of diagnosis.

Fifteen (93.75%) patients achieved CHR i.e. normal blood indices within a period of three months. Nine (56.25%) patients achieved CHR within the first month of therapy with imatinib. One (6.25%) patient who did not achieve complete hematological response after three months of therapy did not achieve it even after one year of therapy. The median time taken for CHR is one month. These values suggest strongly that imatinib is very effective at the hematological level of response in chronic phase CML.

But the goal of present day therapy of CML is complete molecular remission. Evaluation of molecular response is a more sensitive method than cytogenetic response to find out residual leukemia and also the response to the imatinib therapy. It also saves the patient from pain due to multiple bone marrow aspirations and can also predict the cytogenetic response, long term disease control, indicate imatinib resistance or relapse if present, facilitate rationale patient management, allow comparison of different imatinib based treatment strategies.

Six (37.5%) patients achieved CMR in the first six months of imatinib therapy i.e. BCR-ABL/ABL ratio equal to 0.0%. None of the patients who did not achieve complete molecular response within first six months of therapy achieved it after one year of therapy, however they showed slight improvement in their BCR-ABL/ABL ratio. None of the patients who achieved complete molecular response within first six months lost the response. The results are very encouraging and show that Imatinib is highly effective at molecular level as compared to earlier used drugs.

The median BCR ABL/ABL value at the end of the six months was 11%. The median BCR ABL/ABL value at the end of the one year was 3.38%. Thus, we see that with imatinib therapy the rate of molecular response is maximum during the first six months and the probability and extent of molecular response decreases with time if the patient does not respond in first six months. Thus molecular response evaluation after six months can predict the subsequent molecular response after one year. Several mechanisms of resistance to imatinib have been identified from in intro studies of CML cell lines. For clinical purposes two types of resistance to imatinib have been described7, (1) upfront resistance to Imatinib which implies that patient fails to respond to therapy, and (2) point mutation in BCR-ABL that prevented imatinib from inhibiting it kinase activity or amplification of BCR-ABL gene.

Molecular response evaluation can also predict the cytogenetic response as demonstrated by the study done by Wang L et al,8 they found that in patients who had complete cytogenetic remission (CCR) within six months, their BCR ABL / ABL ratio was less than 0.08% at the end of the six months. In our study there were six (37.5%) patients with BCR ABL / ABL value equal to 0.0% after six months. So these six (37.5%) people can be considered to be in CCR. In the study done by Wang L et al,8 patients who achieved a partial cytogenetic response had their BCR ABL/ABL values between 0.08 and 10% after six months of treatment with imatinib. In our study we have two (12.5%) patients who have BCR-ABL / ABL values between 0.08% and 10% after six months of therapy. These two (12.5%) patients can be considered to have achieved a partial cytogenetic response in 6 months of therapy. However, at the end of one year there were six (37.5%) patients with BCR-ABL/ABL values between 0.08% and 10%, so these six (37.5%) patients can be considered to have achieved partial cytogenetic response after one year of therapy. In the study done by Wang L et al,8 cytogenetic non-responders had BCR ABL/ABL values more than 11%. In our study we have eight (50%) patients with BCR-ABL / ABL values more than 11% after six months of therapy and can be considered to have had a minor or no cytogenetic response to imatinib after six months of therapy. However at the end of one year we have only four (25%) such patients, which can be considered as cytogenetic nonresponders. Thus, real time RT Q-PCR can be used as a surrogate monitor of the marrow cytogenetic response to imatinib therapy in CML.

All the patients generally tolerated imatinib therapy very well with minor side effects. The most common side effects were edema and arthralgia. Seven (43.75%) patients complained of arthralgia, which occurred mostly during the first two months of therapy and then gradually subsided. Other side effects at the start of therapy were vomiting, diarrhea, skin rash which were mild and subsided in few days.

CONCLUSION

By this study we have concluded that imatinib mesylate is highly effective in treating chronic phase CML at the hematological and molecular level and so should be considered as the drug of first choice in CML. Complete hematological response was observed in fifteen (93.75%) patients within first three months of treatment and was not lost during one year of follow up. Six (37.5%) patients achieved complete molecular response in the first six months of imatinib therapy as demonstrated by measurement of BCR ABL/ABL values using real time RT Q-PCR, which is a highly sensitive method, used to measure residual leukemia. The molecular response was also not lost during the one year of follow up. Molecular response evaluation after six months can predict the subsequent molecular response and can also be used as a surrogate monitor of the marrow cytogenetic response to imatinib therapy in CML. Molecular response evaluation can be used as a parameter for long term evaluation of patients on imatinib.

However, we still lack long term follow up data on the efficacy of Imatinib in CML. Therefore, clinical trials like this must continue for longer durations to find out the durability of response and long term responses to imatinib in terms of disease free survival and long term survival and also to find out if it is of any advantage to use imatinib in combination with other drugs or with other treatment modalities like allogenic stem cell transplant. Imatinib mesylate has demonstrated the potential for development of anti-cancer drugs based on the specific molecular abnormality present in a human cancer. And so the future of cancer chemotherapy seems to revolve around targeted molecular therapy and promises cure of various other human cancers, which are at present untreatable.

The study was granted in part by Indian Council of Medical Research under Short Term Research Studentship-2005. We are thankful to the Indian Council of Medical Research.

REFERENCES

1. Kantargian HM et al. Chronic leukemias - in Principles and Practice of Oncology. Devita, et al (2433-2445), 6th edition 2000.

2. Goldman JM et al. Clinical decisions for Chronic Myeloid Leukemia in the imatinib Era. Seminars in Hematology. 2003;40:98-103.

3. Oxford textbook of Oncology (1648-1658) 1995.

4. Talpaz Mosche et al. Imatinib induces durable hematologic and cytogenetic responses in patients with accelerated phase myeloid leukemia : Results of a phase 2 study. Blood 2002;99:192837.

5. Sawyers Charles L et al. Imatinib induces hematologic and cytogenetic responses in patients with chronic myelogenous leukemia in myeloid blast crises : results of a phase II study. Blood 2002;99:3530-9.

6. Drunker BJ, Talpaz M, Resta DJ, et al. Efficacy and safety of a specific inhibitor of the BCR-ABL tyrosine kinase in CML. N Engl J Med 2001;344:1031-7.

7. Sauyers et al. Chronic Myeloid Leukemia - Hematology 2001:87-98.