A Study Of The Priming Regimen’s Efficacy And Mechanism In Eradicating Leukemic Cells When Applied On Refractory And Relapsed ALL

Ⅰ. Clinical Observation of CAG Priming Regimen’s Effect on Refractory and Relapsed AMLObjective To evaluate CAG priming regimen’s effect on refractory and relapsed acute myelogenous leukemia (AML), and to clarify the role played by CAG priming regimen in the salvage therapy of refractory and relapsed AML.Methods From October 2003 to September 2009, a total of 36 consecutive patients with refractory or relapsed AML received a course of re-remission-induction therapy of CAG regimen, the response to therapy was assessed after the course. All the patients were followed-up after therapy till February 2010, or the time when disease progressed or the death occurred.Results Morphologic examination of 36 enrolled patients showed M0 3, M1 4, M2 21, M4 3, M5 3, M6 2, and all exhibited myelogenous phenotype. Cytogenetic analysis revealed complex karyotype in 3, normal in 7, t (8;21) in 9, t (8;21) combined with other additional abnormal karyotypes in 11, and other abnormal karyotypes involved with chromosome 7, 8, 9, t (5;11) in 1, 1, 3, 1 patient, respectively. Before the patients received CAG regimen, they had got 78 times chemotherapy all together, failed 62 times. All the patients tolerated the CAG priming regimen well during the course, no serious adverse effect occurred. 27 patients got CR after one course treatment, of which 4 received allo-genetic hematopoietic stem cell transplantation (allo-HSCT) afterwards and acquired continuous complete remission (CR), 5 received autologous hematopoietic stem cell transplantation (auto-HSCT) and only 2 were in CR, 3 relapsed. 18 patients out of 27 patients who achieved CR after CAG regimen got continuous consolidation chemotherapy, 8 were in CR during the follow-up days. The event-free-survival time after achieving CR by CAG priming regimen in HSCT group was longer than chemotherapy group, and the difference was significantly.Conclusions CAG priming regimen was an effective treatment protocol for refractory and relapse AML, and it would be a good choice in salvage therapy of these patients. Moreover, the application of this regimen would create chance for refractory and relapsed AML patients to receive curative therapy--allo-HSCT.Ⅱ. A Pilot Study of CAG Priming Regimen’s Efficacy on Refractory and Relapsed ALLObjective To evaluate CAG priming regimen’s efficacy on refractory and relapsed acute lymphocytic leukemia (ALL), and to clarify the role acted by CAG priming regimen in the salvage therapy of refractory and relapsed ALL.Methods From April 2004 to nowadays, a total of 25 consecutive patients with refractory or relapsed ALL were enrolled in this study. All the enrolled patients received a course of CAG regimen to re-induce remission, and the response to re-induction therapy was assessed after the treatment. All the patients were followed-up after finishing therapy till February 2011, or the time when disease progressed or the death occurred.Results Of the enrolled 25 patients, 11 patients showed T cell immunophenotype, which included 1 pro-T cell ALL, 2 mature-T cell ALL, 8 immature-T cell ALL; 14 patients exhibited B cell immunophenotype, which included 11 common B-ALL. Cytogenetically, 13 patients had normal karyotype, 1 patient had pseudodiploid karyotype combined i(7q), 3 patients had complex karyotype, of whom 2 had Philadelphia chromosome. Furthermore, each of t(1,19), 46, XY, ins(1), 13q-, XX, t(4,11), 46, XX, t(10,14) was found in 1 patient, respectively. Before the patients received CAG regimen, they had got 34 times chemotherapy all together, failed 13 times. All the patients had a good toleration to the CAG priming regimen during the re-induction course, no serious adverse effect happened. 16 patients got good response (CR+PR) after one course treatment, and the response rate was 64%. In 11 T-ALL patients, 10 achieved CR, 1 got PR, the response rate was 100%. In 14 B-ALL patients, 4 achieved CR, 1 got PR, the response rate was 35.7%. The response rate in T-ALL was higher than that in B-ALL, the difference was significantly (χ2/P, 11.0491/0.0009). Follow-up study showed of the patients who got CR after CAG priming only 1 received allo-HSCT and was in continous CR, others all relapsed even those received auto-HSCT.Conclusions CAG priming regimen was an effective treatment protocol for refractory and relapse ALL, and it could be a good choice in salvage therapy of these patients. More importantly, the application of CAG priming could create chance for refractory and relapsed ALL patients to receive curative therapy--allo-HSCT.Ⅲ. A Study of G-CSFR Expression Characteristics on Lymphocytic Lineage CellsObjective To investigate G-CSFR expression characteristics on normal lymphocytes and ALL cells.Methods G-CSFR expression level and G-CSFR mRNA transcripts were examined by flow cytometry with the use of fluorescence-labeled antibodies and RT-PCR in 22 normal people and 25 ALL samples, respectively.Results Lower expression level of G-CSFR was detected in lymphocytes from normal people compared with that detected in normal neutrophil (χ2/P, 32.0006/<.0001), and the G-CSFR expression level of different lymphocyte subgroup was different from each other. Examination results of 25 ALL samples detected by flow cytometry revealed 21 samples expressed G-CSFR and a G-CSFR positive rate of 84%. Among them, T-ALL’s G-CSFR positive rate was 91% (10/11), B-ALL’s G-CSFR positive rate was 77% (10/13), and there was no significant difference between them (χ2/P, 0.8392/0.3596). Only 1 sample of bi-phenotype of T and B-ALL was detected, and it showed G-CSFR positive.Conclusions Lymphocytic lineage cells including ALL cells and normal lymphocytes could express G-CSFR.Ⅳ. A Study of the Proliferative Effect on G-CSFR Positive ALL Cells by G-CSF and Synergetically anti-leukemic Effect on these Cells by G-CSF and Cytotoxic agentObjective To observe the proliferative effect on ALL cells by G-CSF, and to evaluate the synergetically anti-leukemic effect on these cells played by G-CSF and Ara-C.Methods G-CSFR expression density of 14 ALL cell samples (including 5 ALL cell lines) were detected, and expression density changes due to G-CSF treatment were evaluated. After treatment by G-CSF of various concentrations, changes of cell cycle and cell proliferation in ALL cell lines were examined by DNA-PREP-TM kit and Cell Counting Kit (CCK-8), respectively. After treatment by different concentration’s Ara-C combined with different concentration’s G-CSF, ALL samples’existing cell viability were detected by CCK-8.Results All 14 ALL samples expressed G-CSFR; Cell cycle analysis showed that S phase cell proportion in MOLT-4, Jurkat,Clone E6-1, A3 cell lines had a growing tendency after G-CSF’treatment, while no similar tendency was found in CCRF-CEM, 697 cell lines. A relatively proliferative advantage was found in MOLT-4, Jurkat,Clone E6-1, A3 cell lines after G-CSF’s treatment, but only in MOLT-4 there was a statistically significance. A synergetically anti-leukemic effect was observed in MOLT-4, Jurkat,Clone E6-1, A3 cell lines after Ara-C’s treatment combined with G-CSF. Among 14 ALL samples, 7 ALL samples including 3 T-ALL cell lines, that were MOLT-4, Jurkat,Clone E6-1, A3, showed G-CSFR expression down-regulation tendency after G-CSF’s treatment, which was similar in neutrophil. While 7 ALL samples including CCRF-CEM and 697 exhibited G-CSFR expression up-regulation tendency after G-CSF’s treatment. Among G-CSFR expression down-regulation group when treated by G-CSF, synergetically anti-leukemic effect occurred more frequently than in the G-CSFR expression up-regulation group when treated by G-CSF and the difference was significantly (χ2/P, 10.5/0.0012).Conclusions The biological characteristics of ALL cells expressing G-CSFR was different from each other. Synergetically anti-leukemic effect produced by cytotoxic agent and G-CSF could occur in ALL cells which showed a growing S phase cell proportation after G-CSF’s treatment. ALL cell’s response to G-CSF, which means down-regulation of G-CSFR’s expression density, seems a promising index that a synergetically effect could be produced by Ara-C plus G-CSF. Before this index could be applied for guiding ALL patients’management, prospective clinical researches are needed to confirm it.