I. Overview: Chronic Myelogenous Leukemia (CML) was the first malignant disease identified with a specific chromosomal abnormality. abnormal chromosome -t (q34; q11), named chromosome Ph1, was found in the bone marrow cells of CML patients in 1960; chromosome Ph1 can detected in 95% of CML patients, and translocations in the long arms of chromosomes 9 and 22 form a persistently abnormal activation of bcr/abl tyrosine kinase, leading to malignant transformation.
The prognosis of CML before the 1990s was relatively poor, with less than 20% 5-year survival in most patients, except for allogeneic hematopoietic stem cell transplantation, which can successfully cure CML. Patients treated with conventional chemotherapy and alpha interferon eventually developed mostly resistance to therapy, manifested by chromosomal clonal evolution, eosinophilia, peripheral blood progranulocytosis and promyelocytosis, and after 3-18 months 75% of patients would undergo an acute transformation with a presentation similar to high-risk or secondary acute leukemia, and 90% would die within 6 months.
In recent years with the study of the bcr/abl fusion gene, an in-depth understanding of its tyrosine kinase function has been gained, leading to the introduction of the concept of targeted therapy for the treatment of CML. The development of the first targeted therapy drug imatinib mesylate (Gleevec, a tyrosine kinase inhibitor) not only changed the prognosis of the disease, but also contributed to the advancement of efficacy monitoring methods, introducing molecular biology techniques based on cytogenetics to better detect microscopic residual disease.
Second, epidemiology: CML accounts for 15-20% of all adult leukemias worldwide, with an annual incidence of 1-2 cases/100,000 population. The incidence rate of our survey results in the 1980s was 0.36 cases/100,000 population.
The median age of onset of CML patients is 55-60 years, and the incidence increases with age, accounting for 30% of patients over 60 years of age. The American Cancer Society estimates that there will be 850 deaths due to CML in the United States in 2005, accounting for 3.7% of all leukemia deaths.
III. Molecular pathophysiological studies.
The discovery of the Ph1 chromosome was a milestone in the cytogenetic study of CML. In 1982, Heisterkamp et al. reported the discovery of the c-abl oncogene homologous sequence from Abelson mouse leukemia virus on chromosome 9, suggesting that the Ph1 chromosome is not only a marker of CML, but also a possible key factor in the pathogenesis of CML. Recent findings have demonstrated that the Ph chromosome contains the bcr/abl fusion gene, an abnormal gene that results from a broken fragment of the abl proto-oncogene on 9q34 shifted to the broken end of the bcr gene on 22q11. Under normal conditions, in most cells the Abl gene is located in the nucleus and is involved in cell cycle regulation, inhibiting G1 phase cell proliferation, G1/S phase transition, and possibly increasing S phase gene transcription through RNA polymorphase II phosphorylation. The bcr/abl mRNA has no exon 1, so bcr is directly fused to abl exon a2. bcr gene breakpoints on chromosome 22 occur in three regions, with 95% of CML and 1/3 of ALL occurring in the 5.8-kb region called the major The breakpoint cluster region, which has 5 exons, used to be called b1-b5, but is now named e12-e16 according to its true position in the gene. most breakpoints occur in e13 (b2) or e14 (b3), forming e13a2 (b2a2) or e14a2 (b3a2) fusion genes. bcr breakpoints in 2/3 Ph+ALL and a few CML and AML occur upstream of the major breakpoint cluster region, called the minor breakpoint cluster region, with the break occurring between exons e2′ and e2. The third breakpoint region is between exons e19 and e20 at the 3′ end of the bcr gene. bcr/abl-transfected mice show abnormal proliferation of red lineage, macrophages and lymphocytes, and all three different types of bcr/abl-transfected animals equally cause CML-like myeloproliferative syndrome.
BCR/ABL tyrosine kinase phosphorylation and growth signaling mainly down through the Ras and PI3K pathways lead to cancer-like hyperproliferation; Raf and JNK/SAPK are also tyrosine signaling pathways. In addition tumor cell transformation requires transcription of NF-κB, c-Myc, and c-Jun; and transformed tumor cell growth and proliferation requires STAT5 phosphorylation.
The molecular weight of BCR/ABL chimeric proteins varies depending on the bcr gene breakpoint. 210 kDa protein is expressed by most CML patients, whereas Ph+ acute lymphoblastic leukemia expresses both 210 and 190 kDa proteins. p190BCR-ABL tyrosine kinase activity is stronger than p210BCR-ABL. In addition, a larger molecular weight BCR/ABL chimeric protein, P230BCR-ABL, was also found in some patients with specific subclasses of CML, with clinical manifestations of no increase or even decrease in leukocytes.
IV. Clinical manifestations: Half of the patients with CML are asymptomatic at the time of presentation. Common symptoms include malaise, anorexia, left upper abdominal discomfort, abdominal distension, wasting and night sweats; although extreme leukocyte elevation is rare, once it occurs patients may develop hyperviscosity, headache, confusion, tinnitus and abnormal penile erection. More than 50% of patients have splenomegaly on physical examination, and some patients may have mild hepatomegaly.
As the disease progresses patients may develop unexplained fever, bone pain, progressive splenomegaly, and some patients may have leukemic skin infiltrates, hemorrhage, and extramedullary infiltrates at other sites.
Laboratory tests: leukocytes are often >10×109/L, classified as intermediate to mature granulocytes, anemia is rare, and some patients may have thrombocytosis. The bone marrow shows extremely active myeloid proliferation, with predominantly intermediate and late juvenile proliferation, accompanied by an increase in the proportion of eosinophils and basophils. The histochemical alkaline phosphatase score is reduced or even absent. Ph chromosomes can be detected in 95% of patients, and bcr/abl fusion gene can be detected in 100% of patients.
Clinical staging: CML is clinically divided into chronic, accelerated and acute phases, and some patients may enter the acute phase without obvious accelerated phase. As the disease progresses, clinical and laboratory manifestations vary. Recently, WHO has proposed diagnostic criteria for the accelerated phase, which has not yet been universally adopted clinically.
Diagnosis: The diagnosis of CML can be established when the clinical manifestations include splenomegaly, increased peripheral blood leukocytes, middle- to late-stage granulocytes, marked → extremely active bone marrow myeloid hyperplasia, predominantly middle- to late-stage granulocytes, Ph chromosome in cytogenetic examination or bcr/abl fusion gene in molecular biological examination.
In the past, the Ph chromosome was the gold standard for the diagnosis of CML, but the test was time-consuming and could not be reported in a timely manner, and the sensitivity could not reach 100%, and the Ph chromosome could not be found in 5% of CML patients by conventional methods. In recent years, improvements in molecular biology methods have made clinical tests faster and more accurate. Multiplex enzyme chain reaction (PCR) can not only detect the bcr/abl fusion gene, but also determine the exact breakpoint of the fusion gene, and fluorescence in situ hybridization (FISH) also has high sensitivity, is reproducible, and is simple to operate, and results can be reported on the same day. Molecular biology tools are now increasingly used for clinical diagnosis and disease monitoring.
VII. Assessment of prognostic factors: The clinical course of patients with CML varies greatly, with some patients surviving for more than 5-10 years with chemotherapy alone, while others undergo acute changes very quickly. There are many clinical features associated with prognosis, and a number of prognostic evaluation models have been established using these prognostic indicators, currently in common use are the Sokal scoring system and the European scoring system. the Sokal system was established based on the clinical characteristics of patients taking oral hydroxyurea and Maryland alone, and is divided into low, intermediate and high risk, with a 4-year survival rate of 62% for low risk, 43% for intermediate risk and 33% for high risk. The application of interferon greatly improved the prognosis of CML, and the Sokal system could not meet the clinical needs, thus Hasford et al. proposed a new staging system based on the information from the European Collaborative Group on Interferon Therapy for CML.
Imatinib mesylate has changed the status quo in the treatment of CML. Even in patients who failed interferon therapy and received imatinib, more than half of them still achieved complete cytogenetic remission, and the impact of the above indicators on prognosis is no longer significant. In patients treated with imatinib, the response to treatment, the presence or absence of clonal evolution, and the presence or absence of severe granular deficiency at 45-90 days of treatment may have an impact on long-term survival. A prognostic evaluation model for imatinib has been established, and the significance of its impact on prognosis cannot be determined because this drug has been available for a short time.
VIII. Treatment.
(A) Treatment goals and monitoring: CML prognosis has been greatly improved in recent years, the survival of conventional chemotherapy patients is only 35-65 months, the advent of α-interferon has extended the median survival to 65-90 months, and the 10-year survival of patients who obtained cytogenetic complete remission (CCR) is as high as 78%, of which 100% are PCR persistent negative, PCR one-off negative is 76%, and only 46% with a positive PCR (P < 0.001). Therefore, the goal of CML treatment should be cytogenetic complete remission or even molecular biological complete remission. In the IRIS study, 97% of patients who achieved major cytogenetic remission (MCR) at 6 months remained in chronic phase at 2.5 years, while only 89% of patients who did not achieve MCR at 6 months were in chronic phase (P < 0.001), with 30-month survival rates of 97% and 92%, respectively (P = 0.0162). Based on the above findings the basic goal for CML treatment should be to achieve MCR at 6 months, CCR at 12 months and preferably BCR/ABL >3log decline.
The general initial disease monitoring interval is 3 months, mainly monitoring cytogenetics, BCR/ABL fusion gene changes. Although chromosome examination has shortcomings, such as the need for mid-phase division, time-consuming, not 100% sensitive, and failure due to myelosuppression after chemotherapy, it is still the gold standard for evaluating efficacy due to the ability to detect cytogenetic clonal evolution and the standardization of methods. /105 cells) and can be detected in peripheral blood specimens, but they cannot detect cytogenetic clonal evolution. The combination of chromosomal and molecular biology methods is the current trend. The advantages and disadvantages of commonly used monitoring methods are compared in.
(ii) Traditional treatment of CML.
1, Maryland and hydroxyurea: Maryland is an alkylating agent, while hydroxyurea selectively inhibits cellular DNA synthesis, two
Oral chemotherapeutic agents used to be the main treatment for CML. 50-80% of patients can achieve hematological remission after treatment, but few patients can achieve cytogenetic remission and cannot delay disease progression. It is now mostly used in combination regimens and is no longer used as a first-line treatment option alone. In addition, hydroxyurea has replaced maricylan as the first choice due to its fewer side effects and survival advantage over maricylan. Hydroxyurea is generally given at 0.5-2.0g per dose, 2-3 times daily, with the dose adjusted according to the white blood cell count.
2. α-interferon: α-interferon was the first drug reported to achieve cytogenetic remission in CML patients. In a large randomized trial by the German CML Study Group comparing the efficacy of interferon, mariculan and hydroxyurea, it was found that survival was significantly better in the interferon group than in the mariculan group (63.2mo. vs 45.4mo., P=0.008), but similar to that of hydroxyurea (63.2mo. vs 56mo., P=0.44); to determine whether interferon was beneficial to patients A subsequent meta-analysis of 1554 cases from seven large randomized clinical trials worldwide (17) showed that survival was significantly better in the interferon group than in the hydroxyurea (P=0.001) and mariculan groups (P=0.00007).
The dose of interferon has been reported differently; 274 patients with CML-CP treated at the MD Anderson Oncology Center with an interferon dose of 5 million units/m2/d had 80% hematologic remission and 58% cytogenetic remission (26% complete remission) (18); CALGB also used 5 million U/m2/d, 18% cytogenetic complete remission and 21% cytogenetic partial remission (19); Schofield (20) tried interferon 2 million U/m2 three times a week and although hematologic remission was 70%, cytogenetic complete remission was only 7% and partial remission 22%. Therefore interferon dose of 5 million U/m2 is recommended.
To investigate whether combination chemotherapy increases the efficacy of interferon, the German CML-II study compared interferon combined with hydroxyurea with hydroxyurea alone, and survival was significantly better in the combination group than in the single-agent chemotherapy. In contrast, the French and Italian collaborative groups (23,24) compared the efficacy of interferon combined with low-dose cytarabine with that of interferon alone. Among 721 patients with primary CML in France, patients receiving interferon + cytarabine had 35% major cytogenetic remission (MCR) at 12 months compared with 21% in the interferon alone group (P=0.001), with median survival of 89 months and 77 months, respectively ( In the Italian group of 538 patients receiving interferon + cytarabine, the MCR at 24 months was 28% compared with 18% in the interferon group (P=0.003), but there was no significant difference in survival. These results confirm the superiority of interferon combination chemotherapy over interferon monotherapy, but the survival results of the Italian Collaborative Group leave open the question of whether the difference is in the number of cases or the duration of observation, which needs to be further explored.
The European Collaborative Group for the Treatment of CML with Interferon (25) reported 317 patients with CML with a treatment goal of CCR, a mean interferon dosage of 3 to 74 million U per week (median 37 million U), a median time from the start of interferon therapy to the achievement of CCR of 19 months (17-21 months), and an overall survival of 86% at 5 years and 72% at 10 years after obtaining CCR; among them Sokal low-risk group had 93% survival at 5 years and 89% survival at 10 years since obtaining CCR, while Sokal high-risk patients had 54% survival at 5 years and very few at 10 years, and almost no one could reach 10 years. This indicates that interferon therapy mainly benefits intermediate and low-risk CML, and although high-risk patients can prolong the chronic phase, they rarely reach 10 years and should be considered for more intense treatment. To achieve CCR goals, interferon should be dosed adequately.
Long-acting interferon (PEG IFN) can be administered weekly, reducing the inconvenience of frequent injections of regular interferon, and there was no significant difference in safety and efficacy between PEG IFN α-26 μg/kg/w) and IFN α-2b (5 million U/m2/d) in a randomized clinical trial (26). In contrast, in the randomized clinical trial of PEG IFN α-2a (450 μg/w) versus IFN α-2a (9 million U/d), the hematologic complete remission rate was significantly higher with PEG IFN α-2a than with IFN α-2a (69% versus 41%, P=0.008), the MCR rate was also better with PEG IFN α-2a than with IFN α-2a (35% versus 18%, P= 0.0162), with 10% death in the PEG IFN α-2a group and 14% death in the IFN α-2a group at two years of follow-up.
Interferon therapy for CML is mainly indicated in the chronic phase, with the best efficacy in the early chronic phase, and in patients in the accelerated phase, although efficacious, it is mostly transient, with only 20% hematologic remission and no cytogenetic remission in patients in the acute phase.
The most common side effect of α-interferon is influenza-like syndrome, which usually disappears after two weeks after symptomatic treatment with antipyretic and analgesic drugs; long-term use of the drug may also cause fatigue, loss of appetite, hair loss, weight loss, depression, decreased libido, impotence; the elderly and patients with previous history of psychiatric disorders may also develop inattention and mental abnormalities; a few patients may also develop immunological abnormalities, such as positive antinuclear antibodies, thyroid A few patients may also have immunological abnormalities, such as positive antinuclear antibodies, hyper- or hypothyroidism, autoimmune hemolytic anemia, etc.
3.Hematopoietic stem cell transplantation: Allogeneic hematopoietic stem cell transplantation is currently considered to be the only cure for CML, but few patients are eligible. Patients need to have an HLA compatible sibling donor, be generally younger than 55 years old, and preferably be transplanted within one year of the diagnosis of CML. The 5-year survival rate is 50-90% for patients in the chronic phase and decreases with disease progression in patients in the accelerated, acute phase. In a group of 131 patients who received transplantation for CML, 78% survived disease-free at 3 years after transplantation, while mortality increased in those transplanted more than one year after disease; despite the occurrence of extensive cGVHD in 60% of patients, the median Kanofski score was 95% and only 10% scored <80%.
Unrelated hematopoietic stem cells increased the chance of CML transplantation, and survival of matched unrelated stem cell transplants was lower than matched sibling transplants. Prognosis also correlates with patient age, stage of disease, and time to initiation of transplantation.
Non-cleared (including reduced intensity pre-treatment regimens) HSCT allows donor stem cell implantation by suppressing the patient’s immune system without complete clearance of the patient’s bone marrow, and clears leukemic cells by graft-versus-leukemia effect (GVL). The long-term survival rate is 40-85%, II-IV° GVHD 20-54%, and non-relapse death 0-35%.
(iii) Molecularly targeted therapy for CML.
It has long been expected that tumor treatment drugs would target only tumor cells and have little or no effect on normal cells. With the study of CML, the BCR/ABL tyrosine kinase activity underlying leukemogenesis became the most attractive target. Imatinib mesylate is the first molecularly targeted agent that is an ATP mimetic with a stronger affinity than ATP and competitively binds to the BCR/ABL tyrosine kinase ATP binding site, thereby preventing ATP binding and hydrolysis, blocking BCR/ABL phosphorylation to inactivate it, and blocking the signaling pathway downstream of BCR/ABL activation (30).
In phase II clinical trials, imatinib was administered at doses of 400-600 mg/d. Hematologic complete remission (HCR) rates were 95%, 34%, and 8%, major cytogenetic remission (MCR) 60%, 24%, and 16%, and complete cytogenetic remission (CCR) 41%, 17%, and 7%, respectively, in patients in the chronic, accelerated, and acute phases of interferon failure ( 31). a large clinical trial was conducted from July 2000 to January 2001 (14), with a total of 1106 newly diagnosed CML-CP patients entering the Interferon and STI571 International Randomized Controlled Study (IRIS), where patients were randomized to receive either imatinib 400 mg or interferon plus low-dose cytarabine. at 30 months, the imatinib group had a CHR of 95%, an At 30 months, the Imatinib group had a CHR of 95%, MCR of 83%, and CCR of 68%, while the interferon plus cytarabine group had a CHR of 56%, MCR of 16%, and CCR of 5%; 30-month progression-free survival (PFS) was 88% and 68%, respectively (P < 0.001), and overall survival was 95% and 92%, respectively. remained in chronic phase, with only 4.5% cytogenetic relapse and 2.4% hematologic relapse. Cytogenetic remission (especially early in the course) is a prognostic factor for prolonged PFS, and similarly molecular biological remission at 12 months is a predictor of improved PFS (32).
To investigate whether increasing the dose of imatinib improves CML prognosis, the TIDEL study (33) compared the difference between imatinib 600 mg and the IRIS study 400 mg, with significant improvements in MCR (94% versus 83%, P=0.0004) and CCR (89% versus 60%, P<0.0001); although there was little difference in the decrease in BCR/ABL > 3S ( TIDEL 47%, IRIS 40%), more patients in the TIDEL study achieved >4S decreases. the MD Anderson Oncology Center used imatinib 800 mg/d to treat the chronic phase of CML, allowing patients to achieve molecular biological remission earlier without a significant increase in toxicities other than bone marrow suppression (34).
The indications for imatinib include 1) patients with newly diagnosed CML in the chronic phase, 2) patients who have failed interferon therapy in the chronic phase, and 3) patients in the accelerated, acute phase. The recommended dose of imatinib is 400-800 mg per day, and efficacy is positively correlated with dose.
The side effects of imatinib include edema, nausea, weakness, diarrhea, rash, muscle tremor, dyspnea, granulocyte deficiency, thrombocytopenia, and anemia.
Some patients may develop resistance to imatinib, manifested as ineffective treatment (primary or endogenous resistance) or relapse after remission (secondary resistance), mainly in patients with progressive disease, with 24% ineffective treatment and 51% relapse after remission in the accelerated phase; 66% ineffective and 88% relapse in patients with acute change. Mechanisms of drug resistance include mutations in the BCR/ABL tyrosine kinase functional region, BCR/ABL overexpression/amplification, clonal evolution, and expression of multidrug resistance protein (MRP-1). Gene mutations are seen in 50-90% of patients with secondary imatinib resistance, resulting in the inability of imatinib to bind tyrosine kinase and thus loss of efficacy; more than 20 different amino acid alterations have been identified, mainly in the ATP-binding region (P-loop), followed by the activation loop and the carboxyl terminus (35). Gene mutations can occur before imatinib treatment, and in a group of 66 CML patients who had never received imatinib treatment before, gene mutations were found in 15 cases (22.7%), all in patients in the accelerated and acute phase, and none in the chronic phase; by multivariate regression analysis, three indicators of cytogenetic clonal evolution, thrombocytopenia, and previous treatment with 6-thioguanine were significantly correlated with the occurrence of gene mutations significant correlation, but a large scale observation is needed because the number of cases is too small (36). In a German study not only BCR/ABL was found to be amplified at the genomic level by FISH, but in a subsequent report BCR/ABL mRNA was also found to be overexpressed in drug-resistant cases, both mechanisms leading to increased BCR/ABL protein expression. In addition they found additional karyotypic abnormalities in more than half of the drug-resistant patients (37). Additional pharmacological mechanisms have also been associated with resistance; MRP-1 levels are an independent prognostic indicator of imatinib efficacy, and liquid phase chromatography revealed reduced intracellular imatinib concentrations in Pgp-positive patients; AGP levels have also been associated with imatinib efficiency, but there is no evidence to confirm that AGP-imatinib binding directly leads to resistance (35).
In addition to imatinib, several new kinase inhibitors are in clinical trials.
AMN107 is a new aminopyrimidine ATP-competitive inhibitor that is 10-50 times more potent than imatinib against BCR/ABL-expressing cell lines. In phase I clinical trials 119 patients with imatinib-resistant CML, Ph+ ALL were treated with AMN107 for 1-385 days (median 120); 60% of patients with BCR/ABL-mutated CML achieved hematologic remission and 41% cytogenetic remission, while 72% of those without BCR/ABL mutations prior to treatment achieved hematologic remission and 59% cytogenetic remission. The recommended dose for phase II clinical trials is 400 mg bid (38).
Dasatinib is a multitargeted kinase inhibitor that acts on both BCR/ABL and SRC family kinase activity and is 325-fold more effective than imatinib against wild-type BCR/ABL-transfected cells. 88% CHR, 40% MCR, 33% CCR in patients with imatinib-resistant CML in chronic phase after treatment; 50% CHR in accelerated phase, 18% in acute granulocytes, and 50% in acute lytic/Ph+ ALL. Dose 70 mg (range 50-100 mg) bid (39,40).