Minimal residual disease diagnostics in patients with acute myeloid leukemia in the post-transplant period: comparison of peripheral blood and bone marrow analysis.

Considering the high relapse rates of AML after allogeneic hematopoietic stem cell transplant, research aims to improve post-transplant surveillance. To determine the value of peripheral blood (PB) for post-transplant minimal residual disease monitoring, we compared 38 PB and bone marrow (BM) sample pairs in 25 stem cell recipients with NPM1-mutated AML (12 males, 13 females, ages 21-73 years). NPM1A mutation levels and chimerism ratios were determined in non-separated BM/PB. We observed congruent results in 28/38 (74%). In 14/38 sample pairs (37%), BM and PB were negative for the NPM1A mutation. Fourteen sample pairs were positive in BM and PB, albeit at higher mutation levels in the BM in 11 cases (4- to 278-fold). Results were discordant in 10 cases (26%), with weakly positive mutation levels in the BM but negative levels in the PB. Cases with ≥0.2% NPM1A mutation level in BM were always positive in PB. Chimerism was concordant in BM and PB in 21/34 (62%) of sample pairs. In conclusion, MRD monitoring with qPCR for the NPM1 mutation and chimerism from non-separated PB contributes to surveillance in patients with AML in the post-transplant period, but even with highly sensitive qPCR there is a risk of failure to detect the mutation in PB.

Post-transplant immune reconstitution after unrelated allogeneic stem cell transplant in patients with acute myeloid leukemia.

We evaluated immune recovery in 67 patients with acute myeloid leukemia (AML) with a median age of 40 years (4-69) following allo-SCT after reduced (n = 35) or myeloablative (n = 32) conditioning. The following lymphocyte populations were determined on days +30, +90, +180, +270, and +365 by flow associated cell sorting: CD3+, CD3+CD4+, CD3+CD8+, CD3+CD4+/CD3+CD8+ ratio, CD3-CD56+, and CD19+ cells. Peripheral blast count >5% was related to lower number of CD3+CD4+ (day +30) and NK cells (day +180; p = 0.02). Intensity of conditioning did not have any significant impact on the kinetics of immune recovery. Patients with normal CD3+CD4+/CD3+CD8+ ratio (day +30) and NK cell count (day +90; p <0.05) experienced better survival than those with decreased parameters. Post-transplant sepsis/severe infections impaired CD3+CD8+ (day +90; p = 0.015) and CD19+ (day +90; p = 0.02) recovery. Relapse in patients following allo-SCT showed an association with decreased numbers of CD19+ (day +270) and NK cells (day +365). Acute GvHD (II-IV) was accompanied by reduced CD19+ and CD3+CD4+ cells. Thus, the evaluation of post-transplant immune reconstitution in patients with AML might improve risk stratification concerning either relapse or TRM and remains to be further explored.

Bone marrow mesenchymal stromal cells remain of recipient origin after allogeneic SCT and do not harbor the JAK2V617F mutation in patients with myelofibrosis.

The close association of the myeloproliferative neoplasms with the activating non-receptor tyrosine kinase JAK2V617F mutation is well established. To further clarify the pathomechanisms of this mutation in patients with myelofibrosis, we performed screening with quantitative real-time PCR for the respective mutation in in vitro expanded bone marrow (BM) mesenchymal stromal cells (MSCs) and compared the results with BM/peripheral blood (PB). Eight patients with primary/secondary myelofibrosis were investigated before (n = 4) or after allogeneic stem cell transplantation (n = 4). All patients had systemic evidence of the JAK2V617F mutation in BM/PB (mutation ratios 0.2-23.5) at the time of investigation in contrast to negative results in the MSCs (n = 7) or a very low (0.004) mutation ratio (n = 1) which was probably due to hematopoietic contamination. The four patients post-transplant had systemic donor chimerism between 96.5 and 100% in BM/PB, while MSCs showed no evidence of donor-specific alleles. In conclusion, in myelofibrosis, the JAK2V617F mutation is restricted to hematopoietic cells, and cannot explain the stromal alterations being observed in this disorder. Further, the MSCs remain of recipient origin after allogeneic SCT, which might contribute to the increased risk of graft dysfunction or failure in myelofibrosis patients after allogeneic transplantation.

Second allogeneic stem cell transplantation in myeloid malignancies.

For patients with myeloid malignancies who relapse after allogeneic stem cell transplantation (allo-SCT), one salvage option is a second SCT. We retrospectively analyzed outcomes of the second allo-SCT in 25 patients who received at least 2 allografts from related/unrelated donors due to relapse of acute myeloid leukemia, myelodysplastic syndrome or myelofibrosis after the first SCT. A minority of the acute myeloid leukemia/myelodysplastic syndrome patients had reached complete hematological remission before the second SCT (6/25, 24%). Reduced conditioning strategies were performed in the majority (n = 23). Complete remission was achieved in all 21 cases with available data after the second SCT, but relapse was seen in 11/25 patients (44%). After a median follow-up of 18 months (range 6-47), 8/25 patients (32%) were still alive, and of those, 6 (24%) were in stable remission. In 9 cases mortality was associated to relapse and in 8 cases to transplant-related causes (treatment-related mortality; 8/25, 32%). In conclusion, a second SCT offers the chance of stable remission for some patients relapsing with a myeloid malignancy after a first allo-SCT, although high treatment-related mortality and relapse rates remain a problem. Efforts should concentrate on an optimization of conditioning strategies, immunosuppression and post-transplant surveillance for this specific situation.