NPM1 Mutations Are Valid Minimal Residual Disease Parameters Following Allogeneic Stem Cell Transplantation: A Study on 116 AML Patients.

Mutations of the Nucleophosmin gene (NPM1) account for the most frequent molecular markers in acute myeloid leukemia (AML). A very limited number of studies analyzed the prognostic impact of NPM1 mutations in minimal residual disease (MRD) diagnostics, but so far no study analyzed the predictive value following allogeneic stem cell transplantation (SCT). We here performed a retrospective study by quantitative PCR in 116 patients with AML who received allogeneic stem cell transplantation (SCT) between January 2002 and May 2007 in the University Medical Center Hamburg-Eppendorf. We retrospectively performed mutational screening by PCR for NPM1 mutations in 116 pts with AML from bone and peripheral blood marrow samples which had been taken before and after SCT. Cases that were positive for the NPM1 mutation type A before SCT were analyzed by quantitative Taqman based real-time PCR as assessed by a standard curve based on the dilution of the OCI/AML3 cell line in the buffy coat of healthy donors. Cases with the mutation type B were analyzed by semiquantitative methods. In total 139 quantitative PCR analyses were performed during a median follow-up of 216 days after SCT (mean 370; range 35–1825 days) at a median of 7 time points (mean 9; r. 2–25) per patient. Results were correlated with cytomorphology, chimerism, and the clinical course. NPM1 mutations of the subtype A were detected in 13/116 pts (11.2%) before SCT, and a mutation of the subtype B was detected in 1/15 pts (0.7%). There were 6 male pts and 8 females (median age: 47 years; range 21–66 years), who received standard conditioning in 8 and reduced intensity conditioning in 7 transplantations (one patient had two allogeneic transplantations due to relapse of AML after the first SCT). Cytogenetics showed a normal karyotype in 12/13 available cases and a del(20q) in one case. 11/14 had de novo AML, 2/14 pts had secondary AML following myelodysplastic syndrome (MDS), and one had CMML-2. At the time of SCT, 3 pts were at first manifestation, 9 at the first and 3 at the second relapse of the disease. The 4 pts with <5% of bone marrow blasts before SCT all had residual levels of the NPM1 mutation (NPM1mut) ≤0.1%. All 3 pts with ≥11% NPM1mut positive cells at the time of SCT relapsed. After SCT, 10/14 pts became NPM1mut negative, whereas 4 remained NPM1mut positive. All 4/14 pts (29%) who achieved stable remissions became PCR-negative after SCT, whereas all 4 pts who remained PCR-positive after SCT developed relapse. The relapse rate after SCT was 10/15 cases (67%) (the patient with two transplantations had another relapse after the 2. SCT). In 9/10 cases (90%) relapse was associated with an increase in NPMmut positivity. In 6/7 cases (86%) for which the exact time point of relapse was established, an increase in NPM1mut levels preceded the morphological manifestation of relapse with a mean interval of 24 days (range 12–38 days). The increase of NPM1mut was detected by PCR earlier than the decrease of chimerism in 6/9 cases (67%) with a mean interval of 15 days (range 1–36 days).

Conclusions: The quantitative assessment of NPM1 mutations provides a reliable MRD marker in pts with AML for the allogeneic transplantation setting and predicts relapse earlier than morphology or chimerism. This might be helpful for earlier therapeutical interventions such as withdrawal of immunsuppression or adoptive immunotherapy by donor lymphocyte infusions (DLI).