Claudia Lengerke

Ruxolitinib in corticosteroid-refractory graft-versus-host disease after allogeneic stem cell transplantation: A multicenter survey

Despite major improvements in allogeneic hematopoietic cell transplantation over the past decades, corticosteroid-refractory (SR) acute (a) and chronic (c) graft-versus-host disease (GVHD) cause high mortality. Preclinical evidence indicates the potent anti-inflammatory properties of the JAK1/2 inhibitor ruxolitinib. In this retrospective survey, 19 stem cell transplant centers in Europe and the United States reported outcome data from 95 patients who had received ruxolitinib as salvage therapy for SR-GVHD. Patients were classified as having SR-aGVHD (n=54, all grades III or IV) or SR-cGVHD (n=41, all moderate or severe). The median number of previous GVHD-therapies was 3 for both SR-aGVHD (1–7) and SR-cGVHD (1–10). The overall response rate was 81.5% (44/54) in SR-aGVHD including 25 complete responses (46.3%), while for SR-cGVHD the ORR was 85.4% (35/41). Of those patients responding to ruxolitinib, the rate of GVHD-relapse was 6.8% (3/44) and 5.7% (2/35) for SR-aGVHD and SR-cGVHD, respectively. The 6-month-survival was 79% (67.3–90.7%, 95% confidence interval (CI)) and 97.4% (92.3–100%, 95% CI) for SR-aGVHD and SR-cGVHD, respectively. Cytopenia and cytomegalovirus-reactivation were observed during ruxolitinib treatment in both SR-aGVHD (30/54, 55.6% and 18/54, 33.3%) and SR-cGVHD (7/41, 17.1% and 6/41, 14.6%) patients. Ruxolitinib may constitute a promising new treatment option for SR-aGVHD and SR-cGVHD that should be validated in a prospective trial.

Novel therapeutic options in Acute Myeloid Leukemia

Acute myeloid leukemia (AML) is a biologically complex and molecularly and clinically heterogeneous disease, and its incidence is increasing as the population ages. Cytogenetic anomalies and mutation testing remain important prognostic tools for tailoring treatment after induction therapy. Despite major advances in understanding the genetic landscape of AML and its impact on the pathophysiology and biology of the disease, as well as the rapid development of new drugs, standard treatment options have not experienced major changes during the past three decades. Especially for patients with intermediate or high-risk AML, which often show relapse. Allogeneic hematopoietic stem cell transplantation (HSCT) remains the best chance for cure. Here we review the state of the art therapy of AML, with special focus on new developments in immunotherapies and cellular therapies including HSCT and particularly discuss the impact of new conditioning and haplo-identical donor regimens for HSCT, post-transplant strategies for preventing and treating relapse, and emerging novel therapeutic options.

Long-term observation reveals high-frequency engraftment of human acute myeloid leukemia in immunodeficient mice

Repopulation of immunodeficient mice remains the primary method for functional assessment of human acute myeloid leukemia. Published data report engraftment in ~40–66% of cases, mostly of intermediate- or poor-risk subtypes. Here we report that extending follow-up beyond the standard analysis endpoints of 10 to 16 weeks after transplantation permitted leukemic engraftment from nearly every case of xenotransplanted acute myeloid leukemia (18/19, ~95%). Xenogeneic leukemic cells showed conserved immune pheno-types and genetic signatures when compared to corresponding pre-transplant cells and, furthermore, were able to induce leukemia in re-transplantation assays. Importantly, bone marrow biopsies taken at standardized time points failed to detect leukemic cells in 11/18 of cases that later showed robust engraftment (61%, termed “long-latency engrafters”), indicating that leukemic cells can persist over months at undetectable levels without losing disease-initiating properties. Cells from favorable-risk leukemia subtypes required longer to become detectable in NOD/SCID/IL2Rγnull mice (27.5±9.4 weeks) than did cells from intermediate-risk (21.9±9.4 weeks, P<0.01) or adverse-risk (17±7.6 weeks; P<0.0001) subtypes, explaining why the engraftment of the first was missed with previous protocols. Mechanistically, leukemic cells engrafting after a prolonged latency showed inferior homing to the bone marrow. Finally, we applied our model to favorable-risk acute myeloid leukemia with inv(16); here, we showed that CD34+ (but not CD34−) blasts induced robust, long-latency engraftment and expressed enhanced levels of stem cell genes. In conclusion, we provide a model that allows in vivo mouse studies with a wide range of molecular subtypes of acute myeloid leukemia subtypes which were previously considered not able to engraft, thus enabling novel insights into leukemogenesis.

In vitro biomimetic engineering of a human hematopoietic niche with functional properties

Significance The development of an in vitro human bone marrow (BM) tissue appears essential to compile information on human hematopoiesis. Conventional systems fail at both capturing the complexity of the bone marrow niche while allowing the maintenance of functional hematopoietic stem cells (HSCs). Here, we report the development of a human 3D (BM) analogue in a perfusion-based bioreactor system, partially recapitulating structural, compositional, and organizational features of the native human osteoblastic niche environment. The engineered tissue supports the maintenance of some hematopoietic stem and progenitor cell (HSPC) properties. This provides an advanced technological platform of broad fundamental and translational relevance, including the study of human HSPC biology and interactions with their niche, the manipulation of functional human HSPCs, or the identification of factors influencing human hematopoiesis. In adults, human hematopoietic stem and progenitor cells (HSPCs) reside in the bone marrow (BM) microenvironment. Our understanding of human hematopoiesis and the associated niche biology remains limited, due to human material accessibility and limits of existing in vitro culture models. The establishment of an in vitro BM system would offer an experimentally accessible and tunable platform to study human hematopoiesis. Here, we develop a 3D engineered human BM analog by recapitulating some of the hematopoietic niche elements. This includes a bone-like scaffold, functionalized by human stromal and osteoblastic cells and by the extracellular matrix they deposited during perfusion culture in bioreactors. The resulting tissue exhibited compositional and structural features of human BM while supporting the maintenance of HSPCs. This was associated with a compartmentalization of phenotypes in the bioreactor system, where committed blood cells are released into the liquid phase and HSPCs preferentially reside within the engineered BM tissue, establishing physical interactions with the stromal compartment. Finally, we demonstrate the possibility to perturb HSPCs’ behavior within our 3D niches by molecular customization or injury simulation. The developed system enables the design of advanced, tunable in vitro BM proxies for the study of human hematopoiesis.

In Vitro Tumorigenic Assay: The Tumor Spheres Assay

Cancer stem cells (CSCs) are a subpopulation of cells within cancer tissues that are thought to mediate tumor initiation. CSCs are furthermore considered the cause of tumor progression and recurrence after conventional therapies, based on their enhanced therapy resistance properties. A method commonly used to assess CSC potential in vitro is the so-called tumor spheres assay in which cells are plated under non-adherent culture conditions in serum-free medium supplemented with growth factors. Tumor spheres assays have been used in cancer research as an intermediate in vitro cell culture model to be explored before performing more laborious in vivo tumor xenograft assays.

In silico analysis of anti-leukemia immune response and immune evasion in acute myeloid leukemia

The induction of antitumor immune response is one of the most relevant factors contributing to clinical outcome in cancer patients. Composition of tumor infiltrating lymphocytes (TILs) subtypes and...

High mortality in hematopoietic stem cell transplant-associated thrombotic microangiopathy with and without concomitant acute graft-versus-host disease

Transplant-associated thrombotic microangiopathy (TA-TMA) remains a major complication of allogeneic hematopoietic stem cell transplantation (allo-HSCT). We hypothesized that TA-TMA correlates with steroid-refractory acute graft-vs.-host disease (aGvHD) and assessed 660 patients suffering from either AML n = 248, ALL n = 79, CML n = 23, CLL n = 36, lymphoma/myeloma n = 127, MDS/MPN n = 124 or bone marrow failure n = 22, who met the study inclusion criteria and had undergone myeloablative (78%) and non-myeloablative (22%) allo-HSCT between 2006 and 2016. Sixty-five (9.8%) of these patients matched the established diagnostic criteria for TA-TMA, and TA-TMA was shown to be a relevant independent risk factor for mortality (RR 3.27; 95% CI 2.07–5.16). Patients with TA-TMA and concomitant aGvHD had a markedly reduced OS compared to patients with TA-TMA or aGvHD alone (median 5.6 months vs. 7.6 months vs. 55.4 months, respectively; p < 0.0001). Risk factors for development of TA-TMA were aGvHD ≥ grade 2, higher aGvHD grade, steroid-refractory aGvHD, CMV reactivation/end-organ disease, but not the conditioning regimen (RIC or MAC), usage of TBI or TBI dose, underlying disease, donor type, age or sex. TA-TMA, with or without concomitant aGvHD, is a significant complication after allo-HSCT and a high-risk factor for a poor survival outcome. Thus, allo-HSCT recipients with grade 2–4 aGvHD or CMV viremia should be closely monitored for the presence of TA-TMA.