No abstract

Apoptotic protease activating factor 1 (APAF-1) has a critical role in the regulation of apoptosis. In the present study, the mRNA expression analysis of different APAF-1 transcripts (APAF-1S, APAF-1LC, APAF-1LN, and APAF-1XL) was analyzed in bone marrow samples from 37 patients with acute myeloid leukemia (newly diagnosed, with no previous treatment). APAF-1XL and APAF-1LN transcripts (with and without an extra WD-40 repeat region, respectively) were detected in all samples, although the major form expressed was APAF-1XL in 65% of the samples (group 1), while 35% of the samples expressed primarily APAF-1LN (group 2). Only 46% of the patients presented complete remission in response to remission induction therapy (represented by less than 5% marrow blasts and hematological recovery), all but 2 cases being from group 1, 21.6% did not attain complete remission (only 1 case from group 1), and 32.4% of the patients died early. Lower expression of APAF-1XL (APAF-1XL/APAF-1LN ratio <1.2) was associated with a poor response to therapy (P = 0.0005, Fisher exact test). Both groups showed similar characteristics regarding white blood cell counts, cytogenetic data or presence of gene rearrangements associated with good prognosis as AML1-ETO, CBFB-MYH11 and PML/RARA. Since it has been shown that only the isoforms with the extra WD-40 repeat region activate procaspase-9, we suggest that low procaspase-9 activation may also be involved in the deregulation of apoptosis and chemotherapy resistance in acute myeloid leukemia.

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Tumor progress is a multistep process in which clones of cells become abnormal by accumulating growth alterations until they are transformed and/or immortalized. In order to verify a potential role for p53, mdm2, and Ras genes in acute leukemogenesis, we investigated the expression and mutational status of these genes in a cohort of 35 adult patients with acute myeloid leukaemia (AML) at diagnosis with a minimum follow up of 4,5 years. The expression of p21ras, p53, and Mdm2 proteins was studied in bone marrow cytospins stained by APAAP (alkaline phosphatase anti-alkaline phosphatase procedure) using monoclonal antibodies Y13-219, PAb 1801 and 2A10, respectively. N-, K-, H-ras and p53 gene mutations were assessed by PCR-SSCP (polymerase chain reaction/single strand conformation polymorphism) and DNA sequencing to exclude the presence of mutations. We found 2 N-Ras gene and 2 p53 gene mutations in 4 patients. Stratification of AML patients by different cytoplasmic expression patterns (p21ras/p53, p53/mdm2, or p21ras/mdm2, and p21ras/mdm2/p53) revealed that patients with phenotypes p53+/mdm2+ (n=2) had significant best prognosis, patients p53—/mdm2— (n=16) an intermediate prognosis and patients with discrepant phenotypes p53—/mdm2+ or p53+/mdm2— (n=6) a worse prognosis (p=0.003 by Log Rank and p=0.008 by Wilcoxon). Patients with phenotypes p21ras+/mdm2+ (n=2) showed significant best prognosis, patients p53—/mdm2— (n=16) an intermediate prognosis and patients with discrepant phenotypes p53—/mdm2+ or p53+/mdm2— (n=8) a worse prognosis (p=0.07 by Log Rank and p=0.02 by Wilcoxon). Patients with phenotypes p21ras+/p53+/mdm2+(n=2) had a tendency to exhibit best prognosis, patients p21ras—/p53—/mdm2— (n=13) a tendency to intermediate prognosis and patients with discrepant phenotypes p21ras/p53/mdm2 (n=8) had a tendency to worse prognosis (p=0.06 by Log Rank and p=0.06 by Wilcoxon). However, stratification of AML patients by different expression values 0(—), 1(+) or 2(++) for each isolated variable p21ras, p53 and mdm2 using the same survival analysis demonstrated that there were no statistically significant difference among these groups. We found that discrepancies in cytoplasmic expression of p53/mdm2, p21ras/mdm2 and p21ras/p53/mdm2 confer worse long term prognosis to these categories of AML patients. These findings suggest that the signalization pathways p53/mdm2, ras/mdm2 and ras/p53/mdm2 may be active in AML and that an intact p53/Mdm2 axis is important in the prognosis of these patients.