Tania Nicole Masmas

Association of HMGB1 polymorphisms with outcome after allogeneic hematopoietic cell transplantation.

Several studies have demonstrated that genetic variation in cytokine genes can modulate the immune reactions after allogeneic hematopoietic cell transplantation (HCT). High mobility group box 1 protein (HMBG1) is a pleiotropic cytokine that functions as a pro-inflammatory signal, important for the activation of antigen presenting cells (APCs) and propagation of inflammation. HMGB1 is implicated in the pathophysiology of a variety of inflammatory diseases, and we have recently found the variation in the HMGB1 gene to be associated with mortality in patients with systemic inflammatory response syndrome. To assess the impact of the genetic variation in HMGB1 on outcome after allogeneic HCT, we genotyped 276 and 146 patient/donor pairs treated with allogeneic HCT for hematologic malignancies following myeloablative (MA) or nonmyeloablative (NMA) conditioning. Associations between genotypes and outcome were only observed in the cohort treated with MA conditioning. Patient homozygosity or heterozygosity for the-1377delA minor allele was associated with increased risk of relapse (hazard ratio [HR] 2.11, P = .02) and increased relapse related mortality (RRM) (P = .03). Furthermore, patient homozygosity for the 3814C > G minor allele was associated with increased overall survival (OS; HR 0.13, P = .04), progression free survival (PFS; HR 0.30, P = .05) and decreased probability of RRM (P = .03). Patient carriage of the 2351insT minor allele reduced the risk of grade II to IV acute graft-versus-host disease (aGVHD) (HR 0.60, P = .01), whereas donor homozygosity was associated with chronic GVHD (cGVHD) (HR 1.54, P = .01). Our findings suggest that the inherited variation in HMGB1 is associated with outcome after allogeneic HCT following MA conditioning. None of the polymorphisms were associated with treatment-related mortality (TRM).

Evidence for involvement of clonally expanded CD8 + T cells in anticancer immune responses in CLL patients following nonmyeloablative conditioning and hematopoietic cell transplantation

We have analyzed the clonotype composition of CD8+ T cells following nonmyeloablative (NMA) conditioning and hematopoietic cell transplantation (HCT), of patients with chronic lymphocytic leukemia (CLL). Consecutive analyses of blood samples taken up to 2 years following HCT, demonstrated that CD8+ T-cell clonality was highly dynamic in the early phases after HCT, but became more stable after 4–5 months. Moreover, donor lymphocyte infusion (DLI) given for disease progression in one of the patients led to establishment of recurrent as well as new T-cell clonotypes. This coincided with disease remission, strongly suggesting that these T cells were engaged with anti-CLL cytotoxicity. To examine the functional capacity of stable clonally expanded T cells after HCT, CD8+ T cells isolated post-transplant from the recipients were stimulated ex vivo with CLL cells and subsequently analyzed by FACS for surface expression of the marker for cytotoxic activity, CD107a. Stimulation with CLL cells indeed led to surface expression of CD107a, and clonotype analyses of sorted cells demonstrated that CD107a positive T cells were stably expanded following HCT. Our data suggest that clonally expanded CD8+ T-cell clones participate in the ongoing T-cell response against CLL cells following HCT with NMA conditioning.

Degree of Predicted Minor Histocompatibility Antigen Mismatch Correlates with Poorer Clinical Outcomes in Nonmyeloablative Allogeneic Hematopoietic Cell Transplantation

In fully HLA-matched allogeneic hematopoietic cell transplantation (HCT), the main mechanism of the beneficial graft-versus-tumor (GVT) effect and of detrimental graft-versus-host disease (GVHD) is believed to be caused by donor cytotoxic T cells directed against disparate recipient minor histocompatibility antigens (miHAs). The most common origin of disparate miHAs is nonsynonymous single nucleotide polymorphism (nsSNP) differences between donors and patients. To date, only some 30 miHAs have been identified and registered, but considering the many different HLA types in the human population, as well as all the possible nsSNP differences between any 2 individuals, it is likely that many miHAs have yet to be discovered. The objective of the current study was to predict novel HLA-A- and HLA-B-restricted miHAs in a cohort of patients treated with nonmyeloablative conditioning allogeneic HCT (matched related donor, n = 70; matched unrelated donor, n = 56) for a hematologic malignancy. Initially, the cohort was genotyped for 53 nsSNPs in 11 known miHA source proteins. Twenty-three nsSNPs within 6 miHA source proteins showed variation in the graft-versus-host (GVH) direction. No correlation between the number of disparate nsSNPs and clinical outcome was seen. Next, miHAs in the GVH direction were predicted for each patient-donor pair. Using the NetMHCpan predictor, we identified peptides encompassing an nsSNP variant uniquely expressed by the patient and with predicted binding to any of the HLA-A or -B molecules expressed by the patient and donor. Patients with more than the median of 3 predicted miHAs had a significantly lower 5-year overall survival (42% vs 70%, P = .0060; adjusted hazard ratio [HR], 2.6, P = .0047) and significantly higher treatment-related mortality (39% vs 10%, P = .0094; adjusted HR, 4.6, P = .0038). No association between the number of predicted miHAs and any other clinical outcome parameters was observed. Collectively, our data suggest that the clinical outcome of HCT is affected not by disparate nsSNPs per se, but rather by the HLA-restricted presentation and recognition of peptides encompassing these. Our data also suggest that 6 of the 11 proteins included in the current study could contain more miHAs yet to be identified, and that the presence of multiple miHAs confers a higher risk of mortality after nonmyeloablative conditioning HCT. Furthermore, our data suggest a possible role for in silico based miHA predictions in donor selection as well as in selecting candidate miHAs for further evaluation in in vitro and in vivo experiments.

The Prognostic Value of YKL-40 Concentrations in Nonmyeloablative Conditioning Allogeneic Hematopoietic Cell Transplantation

Increased plasma concentrations of YKL-40, also called chitinase-3-like-1 protein (CHI3L1), have been correlated with disease severity in a variety of malignant and inflammatory diseases. The objective of the current study was to assess pretransplant recipient and donor CHI3L1 polymorphisms and plasma YKL-40 concentrations as prognostic biomarkers in a cohort of 149 patients treated with hematopoietic cell transplantation (HCT) after nonmyeloablative conditioning for hematologic malignancies. Recipients with pretransplant YKL-40 concentrations above the age-adjusted 95th percentile (high) had higher relapse-related mortality (33% versus 18%, P = .04; hazard ratio (HR) = 4.41, P = .01), lower progression-free survival (38% versus 64%, P < .01; HR = 2.84, P = .01), and overall survival (42% versus 69%, P = .01; HR = 3.09, P = .01). Recipients transplanted with donors with high YKL-40 concentrations had an increased probability and risk of grade 2-4 acute graft-versus-host disease (aGVHD) (93% versus 62%, P < .01; HR = 2.25, P = .02). CHI3L1 polymorphisms were associated with plasma YKL-40 concentrations, but not with clinical outcomes. In conclusion, our study suggests that plasma YKL-40 could function as a biomarker for relapse risk and treatment-related toxicity, and possibly as a tool complementing clinical risk scores such as the HCT comorbidity index.

Natural T-cell responses against minor histocompatibility antigen (mHag) HY following HLA-matched hematopoietic cell transplantation: what are the requirements for a 'good' mHag?

Allogeneic hematopoietic cell transplantation (HCT) represents a potentially curative treatment modality for several hematologic malignancies. Conventional myeloablative conditioning regimens are associated with high treatment-related mortality. To this end, the introduction of nonmyeloablative (NMA) conditioning implies that older and more frail patients can be offered HCT, and recent data suggest comparable outcome of conventional and NMA–HCT, both with regards to efficacy and side effects, at least for selected indications.1 The curative principle in allogeneic HCT with NMA conditioning is solely related to the graft-versus-leukemia (GVL) effect, and several lines of evidence strongly suggest that donor T cells are the main effectors. In human leukocyte antigen (HLA), identical sibling HCT, T-cell responses to minor histocompatibility antigens (mHags) are—at least in part—responsible for the GVL effect but also cause graft-versus-host disease (GVHD). GVHD represents a major side effect of HCT, and it is well established that there exists correspondence between the GVL effect and GVHD. As a consequence, increased knowledge concerning mechanisms at play and the targets recognized may set the stage for development of treatment strategies that focus on induction of GVL in the absence of GVHD.2 Obviously, increased insight into the cells, molecules and antigens involved with GVL and GVHD are important to possibly be able to control and direct these events more precisely.

259: Absolute chimerism as a tool in monitoring imminent and manifest graft rejection after hematopoietic cell transplantation with nonmyeloablative conditioning

ondary to HIV. Treatment was initiated with D4T (Stavudine), 3TC (Lamivudine) and Efavirenz achieving undetectable viral load and increasing CD4 count. Patient also received Erythropoietin (EPO) and granulocyte-colony stimulating factors (G-CSF) showing an increased number of white blood cells (WBC) but continued with high transfusional requeirment. We had to stop treatment in October because of liver failure and lactic acidosis. In November we changed Stavudine for Tenofovir and reiniciated treatment. Viral load was always undetectable and CD4 count was 500 cells/ul. However megakaryocytopoiesis and erythropoiesis did not respond, requiring many transfusions. Coombs Direct Test (CDT) and Coombs Indirec Test (CIT) were positive.He showed an immunohaematologic profile with 1 autoantibody (Anti-e) and 3 alloantibodies (Anti-Jka, Anti-Lua, Anti-Cw). As he had an identical twin, he was submitted to a syngeneic BMT. Although he was heavily transfused (421 Units ), we did not want to increase immunosuppression so as not to have viral reactivation. The conditioning regimen consisted of Cy 50 mg/kg/qd x 4, and in order to lower the risk of engrafment failure, peripheral blood stem cells were used to maximize the number of donor cells infused (11 10 CD34 cells/kg). At the transplantation the patient was in high-risk. The antiretroviral treatment was not discontinued. No graft versus host disease (GVHD) profilaxis was needed. Neutrophils and platelets engrafted at day 11. After 10 months of transplantation he continues in complete haematologic remission. Serum antibodies, CDT and CIT are negative.Viral load remains undetectable (b-DNA).