Fariba Chinian

Bone Marrow Mesenchymal Stromal Cells to Treat Tissue Damage in Allogeneic Stem Cell Transplant Recipients: Correlation of Biological Markers with Clinical Responses

Bone marrow mesenchymal stromal cells (BMSCs) have been used to treat acute graft‐versus‐host disease (GVHD) and other complications following allogeneic hematopoietic stem cell transplantation (SCT). We conducted a phase I trial using third party, early passage BMSCs for patients with steroid‐refractory GVHD, tissue injury, or marrow failure following SCT to investigate safety and efficacy. To identify mechanisms of BMSC immunomodulation and tissue repair, patients were serially monitored for plasma GVHD biomarkers, cytokines, and lymphocyte phenotype. Ten subjects were infused a fixed dose of 2 × 106 BMSCs/kg intravenously weekly for three doses. There was no treatment‐related toxicity (primary endpoint). Eight subjects were evaluable for response at 4 weeks after the last infusion. Five of the seven patients with steroid‐refractory acute GVHD achieved a complete response, two of two patients with tissue injury (pneumomediastinum/pneumothorax) achieved resolution but there was no response in two subjects with delayed marrow failure. Rapid reductions in inflammatory cytokines were observed. Clinical responses correlated with a fall in biomarkers (Reg 3α, CK18, and Elafin) relevant for the site of GVHD or tissue injury. The GVHD complete responders survived significantly longer and had higher baseline absolute lymphocyte and central memory CD4 and CD8 counts. Cytokine changes also segregated with survival. These results confirm that BMSCs are associated with rapid clinical and biomarker responses in GVHD and tissue injury. However, BMSCs were ineffective in patients with prolonged GVHD with lower lymphocyte counts, which suggest that effective GVHD control by BMSCs requires a relatively intact immune system. Stem Cells 2014;32:1278–1288

Alemtuzumab in T-cell large granular lymphocytic leukaemia: interim results from a single-arm, open-label, phase 2 study.

Background T-cell large granular lymphocytic leukemia (T-LGL) is a lymphoproliferative disease presenting with immune-mediated cytopenias and characterized by clonal expansion of cytotoxic CD3+CD8+ lymphocytes. Methotrexate, cyclosporine, or cyclophosphamide improve cytopenias in 50% of patients as first therapy, but the activity of an anti-CD52 monoclonal antibody, alemtuzumab, is not defined in T-LGL. Methods Twenty-five consecutive subjects with T-LGL were enrolled from October 2006 to March 2015 at the National Institutes of Health (www.clinicaltrials.gov-NCT00345345). Alemtuzumab was administered at 10 mg/day intravenously for 10 days. The primary endpoint was haematologic response at 3 months. Analysis was intention to treat. Here we report the protocol specified interim benchmark of a phase II clinical trial using alemtuzumab in T-LGL. Findings In this heterogeneous, previously treated cohort, 14/25 (56%; 95% CI, 37–73%) subjects had a haematological response at 3 months. In T-LGL cases not associated with myelodysplasia or marrow transplantation, the response rate was 14/19 (74%; 95% CI, 51–86%). First dose infusion reactions were common which improved with symptomatic therapy. EBV and CMV reactivations were common and subclinical. In only 2 patients pre-emptive anti-CMV therapy was instituted. There were no cases of EBV or CMV disease. Alemtuzumab induced sustained reduction of absolute clonal population of T-cytotoxic lymphocytes, as identified by TCRBV-receptor phenotype, but the abnormal clone serendipitously persisted in responders. STAT3 mutations in the SH2 domain, identified in ten subjects, did not correlate with response. When compared with healthy volunteers, T-LGL subjects showed a distinct plasma cytokine and JAK-STAT signature prior to treatment, but neither correlated to response. Interpretation This is the largest and only prospective cohort of T-LGL subjects treated with alemtuzumab yet reported. The high activity with a single course of a lymphocytotoxic agent in a mainly relapsed and refractory suggests that haematologic response outcomes can be accomplished without the need for continued use of oral immunosuppression. Funding This research was supported by the Intramural Research Program of the NIH, National Heart, Lung, and Blood Institute.BACKGROUND T-cell large granular lymphocytic leukaemia (T-LGL) is a lymphoproliferative disease that presents with immune-mediated cytopenias and is characterised by clonal expansion of cytotoxic CD3+ CD8+ lymphocytes. Use of methotrexate, ciclosporin, or cyclophosphamide as first therapy improves cytopenias in 50% of patients, but long-term use of these can lead to toxicity. We aimed to explore the activity and safety of alemtuzumab, an anti-CD52 monoclonal antibody, in patients with T-LGL. METHODS We did this single-arm, phase 2 trial in consecutively enrolled adults with T-LGL referred to the National Institutes of Health in Bethesda, MD, USA. Alemtuzumab was given intravenously at 10 mg per day for 10 days. The primary endpoint was haematological response at 3 months after infusion. A complete response was defined as normalisation of all affected lineages, and a partial response was defined in neutropenic patients as 100% increase in the absolute neutrophil count to more than 5 × 10(8) cells per L, and in those with anaemia, as any increase in haemoglobin of 20 g/L or higher observed in at least two serial measurements 1 week apart and sustained for 1 month or longer without exogenous growth factors support or transfusions. Analysis was by intention to treat. We report results from the first stage of this Simon two-stage design trial; enrolment into the second stage is continuing. This study is registered with ClinicalTrials.gov, number NCT00345345. FINDINGS From Oct 1, 2006, to March 1, 2015, we enrolled 25 patients with T-LGL. 14 patients (56%; 95% CI 35-76) had a haematological response at 3 months. Four patients with associated myelodysplastic syndrome and two who had received haemopoietic stem cell transplantation had either no response or were not evaluable, meaning 14 (74% [49-91]) of the 19 patients with classic T-LGL responded. All patients had an infusion reaction (24 [96%] patients grade 1-2, one [4%] patient grade 3), which improved with symptomatic therapy. All patients developed lymphopenia, with 22 (88%) patients having grade 3 or 4 lymphopenia. The other most common grade 3 and 4 adverse events were leukopenia (eight [32%]) and neutropenic infections (five [20%]). Seven patients died; all were non-responders. INTERPRETATION This is the largest and only prospective study of alemtuzumab in patients with T-LGL. The activity reported with a single course of a lymphocytotoxic drug in patients with mainly relapsed and refractory disease suggests that haematological response can be achieved without continued use of oral immunosuppression. FUNDING National Heart, Lung, and Blood Institute.

Cellular immune profiling after sequential clofarabine and lenalidomide for high risk myelodysplastic syndromes and acute myeloid leukemia

Patients with high risk myelodysplastic syndromes (MDS) and acute myelogenous leukemia (AML) are commonly older with multiple co-morbidities, rendering them unsuitable for intensive induction chemotherapy or transplantation. We report preliminary cellular immune profiling of four cases receiving sequential clofarabine and lenalidomide for high risk MDS and AML in a phase I study. Our results highlight the potential of immune profiling for monitoring immune-modifying agents in high risk MDS and AML.

Over-expression of PD-1 does not predict leukemic relapse after allogeneic stem cell transplantation

Blockade of the T-cell exhaustion marker PD-1 to re-energize the immune response is emerging as a promising cancer treatment. Relapse of hematologic malignancy after allogeneic stem cell transplantation limits the success of this approach, and PD-1 blockade may hold therapeutic promise. However, PD-1 expression and its relationship with post-transplant relapse is poorly described. Because the donor immunity is activated by alloresponses, PD-1 expression may differ from nontransplanted individuals, and PD-1 blockade could risk graft-versus-host disease. Here we analyzed T-cell exhaustion marker kinetics and their relationship with leukemia relapse in 85 patients undergoing myeloablative T-cell-depleted HLA-matched stem cell transplantation. At a median follow-up of 3.5 years, 35 (44%) patients relapsed. PD-1 expression in CD4 and CD8 T cells was comparably elevated in relapsed and nonrelapsed cohorts. Helios+ regulatory T cells and CD8 effector memory cells at day 30 emerged as independent predictors of relapse. Although leukemia antigen-specific T cells did not overexpress PD-1, single-cell analysis revealed LAG3 and TIM3 overexpression at relapse. These findings indicate that PD-1 is an unreliable marker for leukemia-specific T-cell exhaustion in relapsing patients but implies other exhaustion markers and suppressor cells as relapse biomarkers.

Distinct Biomarker Profiles in Ex Vivo T Cell Depletion Graft Manipulation Strategies: CD34+ Selection versus CD3+/19+ Depletion in Matched Sibling Allogeneic Peripheral Blood Stem Cell Transplantation

Various approaches have been developed for ex vivo T cell depletion in allogeneic stem cell transplantation to prevent graft-versus-host disease (GVHD). Direct comparisons of T cell depletion strategies have not been well studied, however. We evaluated cellular and plasma biomarkers in 2 different graft manipulation strategies, CD3+CD19+ cell depletion (CD3/19D) versus CD34+ selection (CD34S), and their associations with clinical outcomes. Identical conditions, including the myeloablative preparative regimen, HLA-identical sibling donor, GVHD prophylaxis, and graft source, were used in the 2 cohorts. Major clinical outcomes were similar in the 2 groups in terms of overall survival, nonrelapse mortality, and cumulative incidence of relapse; however, the cumulative incidence of acute GVHD trended to be higher in the CD3/19D cohort compared with the CD34S cohort. A distinct biomarker profile was noted in the CD3/19D cohort: higher levels of ST2, impaired Helios- FoxP3+Treg reconstitution, and rapid reconstitution of naïve, Th2, and Th17 CD4 cells in the early post-transplantation period. In vitro graft replication studies confirmed that CD3/19D disproportionately depleted Tregs and other CD4 subset repertoires in the graft. This study confirms the utility of biomarker monitoring, which can be directly correlated with biological consequences and possible future therapeutic indications.