TP53 and Decitabine in Acute Myeloid Leukemia and Myelodysplastic Syndromes

Abstract

Background: The molecular determinants of clinical responses to decitabine therapy in patients with acute myeloid leukemia (AML) or myelodysplastic syndromes (MDS) are unclear.

Methods: We enrolled 84 adult patients with AML or MDS in a single-institution trial of decitabine to identify somatic mutations and their relationships to clinical responses. Decitabine was administered at a dose of 20 mg per square meter of body-surface area per day for 10 consecutive days in monthly cycles. We performed enhanced exome or gene-panel sequencing in 67 of these patients and serial sequencing at multiple time points to evaluate patterns of mutation clearance in 54 patients. An extension cohort included 32 additional patients who received decitabine in different protocols.

Results: Of the 116 patients, 53 (46%) had bone marrow blast clearance (<5% blasts). Response rates were higher among patients with an unfavorable-risk cytogenetic profile than among patients with an intermediate-risk or favorable-risk cytogenetic profile (29 of 43 patients [67%] vs. 24 of 71 patients [34%], P<0.001) and among patients with TP53 mutations than among patients with wild-type TP53 (21 of 21 [100%] vs. 32 of 78 [41%], P<0.001). Previous studies have consistently shown that patients with an unfavorable-risk cytogenetic profile and TP53 mutations who receive conventional chemotherapy have poor outcomes. However, in this study of 10-day courses of decitabine, neither of these risk factors was associated with a lower rate of overall survival than the rate of survival among study patients with intermediate-risk cytogenetic profiles.

Conclusions: Patients with AML and MDS who had cytogenetic abnormalities associated with unfavorable risk, TP53 mutations, or both had favorable clinical responses and robust (but incomplete) mutation clearance after receiving serial 10-day courses of decitabine. Although these responses were not durable, they resulted in rates of overall survival that were similar to those among patients with AML who had an intermediate-risk cytogenetic profile and who also received serial 10-day courses of decitabine. (Funded by the National Cancer Institute and others; ClinicalTrials.gov number, NCT01687400 .).

Figure 1. Clinical Responses in Patients with TP53 Mutations

Panel A shows patients with acute myeloid leukemia (AML) or myelodysplastic syndromes (MDS) in the discovery and extension cohorts. Samples obtained from 67 patients were evaluated in the study with sequence analysis at Washington University in St. Louis, and 32 additional patients, who received treatment at the University of Chicago or on alternative protocols at Washington University in St. Louis, were also included in the extension analysis. Panels B and C show mutation clearance observed in 2 patients with TP53 mutations. The variant allele frequency is the percentage of total reads that support a specific somatic mutation. The variant allele frequency of each identified mutation is indicated across time points. Variants in TP53 are indicated in red, variants in DNMT3A are indicated in blue, mutations in genes that are not recurrently mutated in AML are indicated in purple, and blast counts are indicated in orange. Relapse occurred in the patients shown in Panels B and C at day 28 of cycle 8. In Panel C, patient 1023 also had hematologic improvement in platelet and neutrophil counts but incomplete recovery of red cells. Panel D shows the rate of clearance of 21 TP53 mutations identified in 16 patients. Panel E shows the rate of clearance of mutations (specifically, in genes that were mutated in at least 5 of the 54 patients with samples that could be evaluated), with a median of four time points evaluated in each patient. One-way analysis of variance and the Tukey multiple-comparison test were performed to compare the means of mutation clearance for each gene. Horizontal bars with an asterisk denote P<0.05 for the indicated comparisons. Short horizontal bars within the data points denote the mean mutation clearance rate for the indicated gene.

Figure 2. Correlation between Somatic Mutations and Clinical Responses

Panel A shows the results of enhanced exome sequencing in the discovery cohort among patients who had a response (complete remission, complete remission with incomplete count recovery, or morphologic complete remission) or no response (partial response, stable disease, or progressive disease). Panel B shows the results of gene-panel sequencing (264 genes in 15 patients and 8 genes in 45 patients) in patients in the extension cohort. ND denotes not done. Panel C shows the proportions of patients among all those in whom samples were sequenced (99 patients) who had a response or did not have a response, according to the presence of the indicated mutations. Panel D shows the locations and predicted consequences of TP53 mutations identified in this trial and in patients with AML in the Cancer Genome Atlas Research Network cohort.

Figure 3. Correlation between Clinical Responses and Pharmacologic and DNA Methylation Measurements

Panel A shows steady-state plasma decitabine levels that were determined during day 4 of cycle 1 (deviation of up to 1 day for sample collection was permitted for patient convenience) (P=0.09 by analysis of variance for the comparison among all response groups). CR denotes complete remission, CRi complete remission with incomplete count recovery, mCR morphologic complete remission, meCpG methylated CpG, NA not applicable, PD progressive disease, PR partial response, and SD stable disease. The horizontal lines indicate pairwise comparisons with statistical differences. Panel B shows the absolute difference between the proportions of meCpGs on day 0 and day 10 of cycle 1, as determined with the use of Illumina Human Methylation 450 Bead Chip profiling (P=0.19 by analysis of variance for the comparison among all response groups). The I bars indicate standard deviation. Panel C shows the mean fraction of meCpGs per sample on day 0 and day 10, according to data shown in Panel B. Panel D shows the differences in the mean fraction of differentially methylated CpGs (those with a change in methylation of ≥25%) between day 0 and day 10. In Panels C and D, the asterisks indicate P<0.001 for the indicated comparisons. Panels E and F show examples of CpG methylation values across approximately 450,000 unique CpGs in representative patients. Samples from total bone marrow cells on day 0 and on day 10 of cycle 1 are compared. Red squares indicate CpGs with a difference of 25% or more in the proportion of cells with methylation at the indicated CpG, and gray squares indicate CpGs with a difference of less than 25%. Hypomethylation predominantly occurs at highly methylated CpGs, and hypomethylation is incomplete within the total bone marrow population. P values were calculated with the use of one-way analysis of variance and the Tukey multiple-comparison test. Similar results were obtained with the use of analysis of covariance (see the Methods section in the Supplementary Appendix).

Figure 4. Correlation between Clinical Variables and Survival

Panel A shows the rate of overall survival among 116 patients as a function of responses to decitabine. CR/CRi/mCR denotes complete remission, complete remission with incomplete count recovery, or morphologic complete remission; PR/SD partial response or stable disease; and PD/NA progressive disease or not assessed. Panel B shows the correlation between karyotype and the presence of TP53 mutations. Panel C shows the rate of survival among 114 patients with karyotypes associated with unfavorable risk or with karyotypes associated with either intermediate risk or favorable risk. (Karyotype data were not available for 2 patients.) Panel D shows the rate of survival among patients with TP53 mutations and patients with wild-type TP53. Panel E shows the rate of survival among 116 patients according to whether they had undergone allogeneic stem-cell transplantation. Half the patients received a conditioning regimen of fludarabine and busulfan, a third received fludarabine and melphalan, and the others received busulfan and cyclophosphamide, cladribine and melphalan, or fludarabine and total-body irradiation. No differences in survival were noted on the basis of the conditioning regimen. Panel F shows the rate of survival among patients who underwent allogeneic stem-cell transplantation according to the type of TP53 mutation. P values were calculated with the use of pair-wise Wilcoxon analysis (Panels A and F) and log-rank analysis (Panels C through E).