Georg-Nikolaus Franke

Graft-Versus-Host Disease and Graft-Versus-Tumor Effects After Allogeneic Hematopoietic Cell Transplantation

PURPOSE We designed a minimal-intensity conditioning regimen for allogeneic hematopoietic cell transplantation (HCT) in patients with advanced hematologic malignancies unable to tolerate high-intensity regimens because of age, serious comorbidities, or previous high-dose HCT. The regimen allows the purest assessment of graft-versus-tumor (GVT) effects apart from conditioning and graft-versus-host disease (GVHD) not augmented by regimen-related toxicities. PATIENTS AND METHODS Patients received low-dose total-body irradiation ± fludarabine before HCT from HLA-matched related (n = 611) or unrelated (n = 481) donors, followed by mycophenolate mofetil and a calcineurin inhibitor to aid engraftment and control GVHD. Median patient age was 56 years (range, 7 to 75 years). Forty-five percent of patients had comorbidity scores of ≥ 3. Median follow-up time was 5 years (range, 0.6 to 12.7 years). RESULTS Depending on disease risk, comorbidities, and GVHD, lasting remissions were seen in 45% to 75% of patients, and 5-year survival ranged from 25% to 60%. At 5 years, the nonrelapse mortality (NRM) rate was 24%, and the relapse mortality rate was 34.5%. Most NRM was a result of GVHD. The most significant factors associated with GVHD-associated NRM were serious comorbidities and grafts from unrelated donors. Most relapses occurred early while the immune system was compromised. GVT effects were comparable after unrelated and related grafts. Chronic GVHD, but not acute GVHD, further increased GVT effects. The potential benefit associated with chronic GVHD was outweighed by increased NRM. CONCLUSION Allogeneic HCT relying on GVT effects is feasible and results in cures of an appreciable number of malignancies. Improved results could come from methods that control progression of malignancy early after HCT and effectively prevent GVHD.

Low tumor burden is associated with early B-cell reconstitution and is a predictor of favorable outcome after non-myeloablative stem cell transplant for chronic lymphocytic leukemia

Abstract Reconstitution, engraftment kinetics and tumor cell clearance were analyzed after reduced intensity conditioning hematopoietic cell transplant (RIC-HCT) in patients with chronic lymphocytic leukemia (CLL). Patients were transplanted from unrelated (n = 40) or related (n = 10) donors after fludarabine and 2 Gy total body irradiation followed by cyclosporine and mycophenolate mofetil. The vast majority of patients (96%) engrafted with absolute neutrophil count (ANC) > 0.5 × 109/L at day + 22. CLL cells decreased (median 2%, range 0–69%) within 28 days, but disappeared by day + 180 after HCT. Donor T-cell chimerism increased to > 95% at day 56 and donor B-cell chimerism to 94% at day + 360. Overall survival was 51 ± 8%, incidence of progression 37 ± 7% and non-relapse related mortality (NRM) 30 ± 7% at 4 years. The most common causes of NRM were graft-versus-host disease (GvHD) (14%) and sepsis (6%). Disease status at HCT was significantly associated with early B-cell reconstitution (p = 0.04) and with increased risk of relapse/progression in univariate and multivariate analysis (p = 0.022). Tumor cells were undetectable by day + 180, although B-cell reconstitution did not occur until 1.5 years after RIC-HCT. The best predictors for progression-free survival (PFS) and overall survival (OS) were complete response (CR) or first partial response (PR1) and the absence of bulky disease at transplant, respectively.

Digital droplet PCR-based absolute quantification of pre-transplant NPM1 mutation burden predicts relapse in acute myeloid leukemia patients

Allogeneic hematopoietic stem cell transplantation is an established consolidation therapy for patients with acute myeloid leukemia. However, relapse after transplantation remains a major clinical problem resulting in poor prognosis. Thus, detection of measurable (“minimal”) residual disease to identify patients at high risk of relapse is essential. A feasible method to determine measurable residual disease may be digital droplet PCR (ddPCR) that allows absolute quantification with high sensitivity and specificity without the necessity of standard curves. Using ddPCR, we analyzed pre-transplant peripheral blood and bone marrow of 51 NPM1-mutated acute myeloid leukemia patients transplanted in complete remission or complete remission with incomplete recovery. Mutated NPM1 measurable residual disease-positive patients had higher cumulative incidence of relapse (P < 0.001) and shorter overall survival (P = 0.014). Restricting the analyses to patients receiving non-myeloablative conditioning, mutated NPM1 measurable residual disease positivity is associated with higher cumulative incidence of relapse (P < 0.001) and shorter overall survival (P = 0.006). Positive mutated NPM1 measurable residual disease status determined by ddPCR before allogeneic stem cell transplantation is associated with worse prognosis independent of other known prognostic markers—also for those receiving non-myeloablative conditioning. In the future, mutated NPM1 measurable residual disease status determined by ddPCR might guide treatment and improve patients’ outcomes.

Prognostic relevance of DNMT3A R882 mutations in AML patients undergoing non-myeloablative conditioning hematopoietic stem cell transplantation

Mutations in the epigenetic modifying gene DNA methyltransferase 3A (DNMT3A) are found in 14–35% of acute myeloid leukemia (AML) patients [1–6]. The majority of DNMT3A mutations cluster at codon R882 in exon 23 [1–6]. Most of the studies that have investigated the prognostic impact of these alterations reported inferior outcomes associated with the presence of DNMT3A mutations—particularly in codon R882 [1–4, 6–9]. However, the negative impact on outcomes may be dependent on the clinical and biological context, such as the presence of concurrent gene mutations (i.e. NPM1 mutations and/or presence of FLT3ITD) and the DNMT3A mutation type (i.e. R882 vs. nonR882) [1–6, 9]. In older AML patients, DNMT3A mutations are more prevalent [7] and have been associated with poor outcomes [1, 6, 8]. DNMT3A mutations are often found to persist during remission (possibly due to a preexisting clonal hematopoiesis) and at AML relapse, and might contribute to chemotherapy resistance [10]. This raises the issue of allogeneic hematopoietic stem cell transplantation (HSCT) as a therapeutic approach specifically for DNMT3A mutated AML. Studies that included HSCT treated patients—most of which focused on younger AML patients—observed no influence of an HSCT on the negative prognostic impact of DNMT3A mutations [3–5, 9, 11]. For example, Gaidzik et al. [5]. described a shorter relapse-free survival in younger DNMT3A R882 mutated AML patients independent of the use of HSCT. Similarly, Ahn et al. [9]. and Xu et al. [11]. reported a negative impact of DNMT3A R882 mutations after HSCT in mainly younger patients with normal karyotype (NK) AML. Since DNMT3A R882 mutations occur more frequently with age [7] and data on the influence of DNMT3A R882 mutations on outcome in older AML patients receiving HSCT are limited, we analyzed the prognostic impact of DNMT3A R882 mutations at diagnosis among mainly older (>60 years) AML patients who underwent nonmyeloablative (NMA)-HSCT. NMA-HSCT relies on an immunological donor leukocyte-mediated GvL effect and, because of the lower treatment-related toxicity, represents a feasible consolidating therapy for older AML patients and/or those with comorbidities [12, 13]. The patients studied here were treated with cytarabinebased chemotherapies followed by conditioning with fludarabine and 2 Gy TBI or 2 Gy TBI only before NMAHSCT (for further details, see Supplementary Information) at the University Hospital Leipzig between February 2003 and December 2012. We retrospectively analyzed the presence of DNMT3A R882 mutations in pre-treatment bone marrow (n= 98) or peripheral blood (n= 10) samples of 108 AML patients (for further details, see Supplementary Information). All patients gave written informed consent in accordance with the declaration of Helsinki. Pre-treatment cytogenetics and the mutation status of NPM1, CEBPA, and the presence of FLT3-ITD were determined as previously Laura K. Schmalbrock and Lynn Bonifacio contributed equally to this work.

High BAALC copy numbers in peripheral blood prior to allogeneic transplantation predict early relapse in acute myeloid leukemia patients

High BAALC expression levels at acute myeloid leukemia diagnosis have been linked to adverse outcomes. Recent data indicate that high BAALC expression levels may also be used as marker for residual disease following acute myeloid leukemia treatment. Allogeneic hematopoietic stem cell transplantation (HSCT) offers a curative treatment for acute myeloid leukemia patients. However, disease recurrence remains a major clinical challenge and identification of high-risk patients prior to HSCT is crucial to improve outcomes. We performed absolute quantification of BAALC copy numbers in peripheral blood prior (median 7 days) to HSCT in complete remission (CR) or CR with incomplete peripheral recovery in 82 acute myeloid leukemia patients using digital droplet PCR (ddPCR) technology. An optimal cut-off of 0.14 BAALC/ABL1 copy numbers was determined and applied to define patients with high or low BAALC/ABL1 copy numbers. High pre-HSCT BAALC/ABL1 copy numbers significantly associated with higher cumulative incidence of relapse and shorter overall survival in univariable and multivariable models. Patients with high pre-HSCT BAALC/ABL1 copy numbers were more likely to experience relapse within 100 days after HSCT. Evaluation of pre-HSCT BAALC/ABL1 copy numbers in peripheral blood by ddPCR represents a feasible and rapid way to identify acute myeloid leukemia patients at high risk of early relapse after HSCT. The prognostic impact was also observed independently of other known clinical, genetic, and molecular prognosticators. In the future, prospective studies should evaluate whether acute myeloid leukemia patients with high pre-HSCT BAALC/ABL1 copy numbers benefit from additional treatment before or early intervention after HSCT.

Prognostic Impact of the CD34+/CD38‐ Cell Burden in Patients with Acute Myeloid Leukemia receiving Allogeneic Stem Cell Transplantation

In acute myeloid leukemia (AML), leukemia‐initiating cells exist within the CD34+/CD38− cell compartment. They are assumed to be more resistant to chemotherapy, enriched in minimal residual disease cell populations, and responsible for relapse. Here we evaluated clinical and biological associations and the prognostic impact of a high diagnostic CD34+/CD38− cell burden in 169 AML patients receiving an allogeneic stem cell transplantation in complete remission. Here, the therapeutic approach is mainly based on immunological graft‐versus‐leukemia effects. Percentage of bone marrow CD34+/CD38− cell burden at diagnosis was measured using flow cytometry and was highly variable (median 0.5%, range 0%–89% of all mononuclear cells). A high CD34+/CD38− cell burden at diagnosis associated with worse genetic risk and secondary AML. Patients with a high CD34+/CD38− cell burden had shorter relapse‐free and overall survival which may be mediated by residual leukemia‐initiating cells in the CD34+/CD38− cell population, escaping the graft‐versus‐leukemia effect after allogeneic transplantation. Evaluating the CD34+/CD38− cell burden at diagnosis may help to identify patients at high risk of relapse after allogeneic transplantation. Further studies to understand leukemia‐initiating cell biology and develop targeting therapies to improve outcomes of AML patients are needed.