Safety, Feasibility, and Advantages of Oral Microbiota Transplantation: The First Clinical Case.

The pilot clinical study presented demonstrates the possibility, safety, and effectiveness of oral microbiota transplantation from a healthy donor to a patient with neuroblastoma to prevent chemotherapy-induced oral mucositis. A 6-month-old patient with a diagnosis of retroperitoneal neuroblastoma was treated according to the NB 2004 protocol. Due to the development of severe oral mucositis, it was decided to perform oral microbiota transplantation. During the next 3 chemotherapy cycles and conditioning regimen before autologous hematopoietic cell transplantation (auto-HCT), the patient was repeatedly injected per os with donor saliva from her healthy mother. Oral microbiota transplantation was shown to effectively prevent the development of oral mucositis after chemotherapy, and only grade 1 oral mucositis developed after auto-HCT. In all loci of the oral cavity, there was a decreased abundance of bacteria from the Staphylococcaceae, Micrococcaceae, and Xanthomonadaceae families. Conversely, there was an increase in the relative abundance of Streptococcaceae and certain other bacterial taxa. In conclusion, the transplantation of maternal saliva in this patient prevented severe mucositis and was accompanied by a compositional change of the patient's oral microbiota. No adverse events due to the transplantation of maternal saliva were noted.

Efficacy and safety of nivolumab combined with brentuximab vedotin after nivolumab monotherapy failure in patients with relapsed and refractory classic Hodgkin lymphoma.

OBJECTIVES: Therapy of patients with relapsed and refractory classic Hodgkin lymphoma (r/r cHL) after PD-1 inhibitors failure remains an unresolved issue. The aim of this study was to evaluate the efficacy and safety of the combination of nivolumab with brentuximab vedotin (Nivo + BV) after nivolumab monotherapy failure. METHODS: This study retrospectively analyzed 21 patients with r/r cHL who were treated with the combination of Nivo + BV after Nivo failure. The response was evaluated by PET-CT scan according to the LYRIC criteria. Adverse events (AEs) were assessed according to NCI CTCAE v.4.03. RESULTS: Median follow-up was 19 (9-47) months. The ORR was 57%. The median OS was not reached, 24 month OS was 80% (95% CI 50-93%). Median PFS was 12 months with 24 month PFS of 31% (95% CI 12-53%). Any grade AEs were observed in 12 patients (63%), 3-4 grade AEs in 2 patients (10%). Allogeneic hematopoietic stem cell transplantation (allo-HSCT) after Nivo + BV was performed in 8 (38%) patients. The median time between Nivo + BV and allo-HSCT was 8 (5-21) months. CONCLUSIONS: Combination of Nivo + BV in r/r cHL after nivolumab monotherapy failure is potentially an effective and safe approach.

Evaluating the Effect of 3'-UTR Variants in DICER1 and DROSHA on Their Tissue-Specific Expression by miRNA Target Prediction.

Untranslated gene regions (UTRs) play an important role in controlling gene expression. 3'-UTRs are primarily targeted by microRNA (miRNA) molecules that form complex gene regulatory networks. Cancer genomes are replete with non-coding mutations, many of which are connected to changes in tumor gene expression that accompany the development of cancer and are associated with resistance to therapy. Therefore, variants that occurred in 3'-UTR under cancer progression should be analysed to predict their phenotypic effect on gene expression, e.g., by evaluating their impact on miRNA target sites. Here, we analyze 3'-UTR variants in DICER1 and DROSHA genes in the context of myelodysplastic syndrome (MDS) development. The key features of this analysis include an assessment of both "canonical" and "non-canonical" types of mRNA-miRNA binding and tissue-specific profiling of miRNA interactions with wild-type and mutated genes. As a result, we obtained a list of DICER1 and DROSHA variants likely altering the miRNA sites and, therefore, potentially leading to the observed tissue-specific gene downregulation. All identified variants have low population frequency consistent with their potential association with pathology progression.

Nivolumab discontinuation and retreatment in patients with relapsed or refractory Hodgkin lymphoma.

Immune checkpoint inhibitors (ICI) have demonstrated high therapeutic efficacy in relapsed or refractory classical Hodgkin lymphoma (r/r cHL). Nevertheless, despite the accumulated data, the question of the ICI therapy duration and efficacy of nivolumab retreatment remains unresolved. In this retrospective study, in a cohort of 23 adult patients with r/r cHL who discontinued nivolumab in complete response (CR), the possibility of durable remission achievement (2-year PFS was 55.1%) was demonstrated. Retreatment with nivolumab has demonstrated efficacy with high overall response rate (ORR) and CR (67% and 33.3% respectively). At the final analysis, all patients were alive with median PFS of 16.5 months. Grade 3-4 adverse events (AEs) were reported in 36% of patients, and there was no deterioration in terms of nivolumab retreatment-associated complications.

Presence of viremia during febrile neutropenic episodes in patients undergoing chemotherapy for malignant neoplasms.

The importance of viral infections as a leading cause of morbidity and mortality is well documented in severely immunosuppressed patients undergoing allogeneic stem cell transplantation. By contrast, viral infections generally receive less attention in patients with malignant disorders undergoing chemotherapy, where the onset of neutropenic fever is mostly associated with bacterial or fungal infections, and screening for viral infections is not routinely performed. To address the occurrence of invasive viral infections in a clinical setting commonly associated with less pronounced immunosuppression, we have prospectively screened 237 febrile neutropenic episodes in pediatric (n = 77) and adult (n = 69) patients undergoing intensive chemotherapy, primarily for treatment of acute leukemia. Serial peripheral blood specimens were tested by RQ-PCR assays for the presence and quantity of the clinically relevant viruses CMV, EBV, HHV6 and HAdV, commonly reactivated in highly immunocompromised patients. Viremia was documented in 36 (15%) episodes investigated, including the detection of HHV6 (n = 14), EBV (n = 15), CMV (n = 6), or HAdV (n = 1). While low or intermediate levels of viremia (<104 virus copies/mL) were commonly associated with bacterial or fungal co-infection, viremia at higher levels (>104 copies/mL) was documented in patients without evidence for other infections, raising the possibility that at least in some instances the onset of fever may have been attributable to the virus detected. The observations suggest that viral infections, potentially resulting from reactivation, might also play a clinically relevant role in patients receiving chemotherapy for treatment of malignant neoplasms, and routine screening for viremia in this clinical setting might be warranted.

Long-term impact of fecal transplantation in healthy volunteers.

BACKGROUND: Fecal microbiota transplantation (FMT) has been recently approved by FDA for the treatment of refractory recurrent clostridial colitis (rCDI). Success of FTM in treatment of rCDI led to a number of studies investigating the effectiveness of its application in the other gastrointestinal diseases. However, in the majority of studies the effects of FMT were evaluated on the patients with initially altered microbiota. The aim of our study was to estimate effects of FMT on the gut microbiota composition in healthy volunteers and to monitor its long-term outcomes. RESULTS: We have performed a combined analysis of three healthy volunteers before and after capsule FMT by evaluating their general condition, adverse clinical effects, changes of basic laboratory parameters, and several immune markers. Intestinal microbiota samples were evaluated by 16S rRNA gene and shotgun sequencing. The data analysis demonstrated profound shift towards the donor microbiota taxonomic composition in all volunteers. Following FMT, all the volunteers exhibited gut colonization with donor gut bacteria and persistence of this effect for almost ∼1 year of observation. Transient changes of immune parameters were consistent with suppression of T-cell cytotoxicity. FMT was well tolerated with mild gastrointestinal adverse events, however, one volunteer developed a systemic inflammatory response syndrome. CONCLUSIONS: The FMT leads to significant long-term changes of the gut microbiota in healthy volunteers with the shift towards donor microbiota composition and represents a relatively safe procedure to the recipients without long-term adverse events.

Risk-adapted GVHD prophylaxis with post-transplantation cyclophosphamide in adults after related, unrelated, and haploidentical transplantations.

INTRODUCTION: Although a number of studies were published on the efficacy of post-transplantation cyclophosphamide (PTCy) for graft-versus-host disease (GVHD) prophylaxis, no large studies prospectively evaluated this strategy in related, unrelated, and haploidentical grafts. METHODS: In this study, GVHD prophylaxis for 57 matched bone marrow (MBM) grafts consisted of single-agent PTCy, for 88 matched PBSC grafts (MPBSC) consisted of PTCy, tacrolimus, and mycophenolate mofetil (MMF) 30 mg/kg, and for 55 mismatched grafts (MMGs) consisted of PTCy, tacrolimus and MMF 45 mg/kg. RESULTS: The study met the primary endpoint to demonstrate equivalent rates of acute GVHD grade II-IV (11%, 17%,19%, P = .46), III-IV (7%, 2%, 6%, P = .41), and moderate and severe chronic GVHD (22%, 11%, 15%, P = .23). There was also no differences in non-relapse mortality (11% vs 15% vs 17%, P = .75), overall survival (63% vs 71% vs 56%, P = .72), event-free-survival (51% vs 66% vs 48%, P = .32) for MBM, MPBSC, and MMG groups, respectively. Toxicity was comparable between groups except higher incidence of nephrotoxicity in combination arms (P = .0005) and higher incidence of graft failures in MMG group (P = .004). CONCLUSION: The suggested risk-adapted PTCy-based prophylaxis is feasible and is associated with low GVHD incidence and mortality in all types of grafts. The study was registered on clinicaltrials.gov (NCT02294552).

Graft-versus-Host Disease Prophylaxis in Unrelated Peripheral Blood Stem Cell Transplantation with Post-Transplantation Cyclophosphamide, Tacrolimus, and Mycophenolate Mofetil.

Clinical efficacy of post-transplantation cyclophosphamide (PTCy) as graft-versus-host disease (GVHD) prophylaxis has been demonstrated in haploidentical and HLA-matched bone marrow but not in unrelated peripheral blood stem cell (PBSC) transplantations. Also, no direct comparisons have been published with current standard of care, combination of antithymocyte globulin (ATG), calcineurin inhibitors, and either methotrexate or mycophenolate mofetil (MMF). Eighty-six adult patients (median age 34 years; range, 18 to 59) with acute myeloblastic and lymphoblastic leukemia underwent unrelated PBSC transplantation with PTCy, tacrolimus, and MMF as GVHD prophylaxis in the single-center trial (clinicaltrial.govNCT02294552). The control group comprised 125 consecutive historical control patients who received ATG, tacrolimus, and methotrexate or MMF. Cumulative incidences of grades II to IV acute (19% versus 45%, P = .0003), grades III to IV acute (4% versus 27%, P < .0001), and chronic GVHD (16% versus 65%, P < .0001) were significantly lower in the PTCy compared with the ATG group. PTCy-based prophylaxis was associated with reduced incidence of nonrelapse mortality (16% versus 36%, P = .005; HR, .55; 95% CI, .34 to .89) and improved overall survival (69% versus 40%, P = .0007; HR, .43; 95% CI, .26 to .70), event-free survival (65% versus 38%, P = .0006; HR, .49; 95% CI, .31 to .78), and GVHD relapse-free survival (52% versus 12%, P < .0001). PTCy-based prophylaxis also had a better safety profile compared with ATG with reduced incidence of veno-occlusive disease, cytomegalovirus reactivation, invasive mycosis, and reduced severity of mucositis. In this study we demonstrated that PTCy in combination with tacrolimus and MMF is a safe and effective GVHD prophylaxis for unrelated PBSC transplantation. Although there are several limitations of the historical control approach, this study suggests the superiority of a PTCy-based approach over an ATG-based prophylaxis.

Time-Dependent Shifts in Intestinal Bacteriome, Klebsiella Colonization and Incidence of Antibiotic-Resistance Genes after Allogeneic Hematopoietic Stem Cell Transplantation.

Dose-intensive cytostatic therapy and antibiotic treatment in allogeneic hematopoietic stem cell transplantation (allo-HSCT) cause severe abnormalities in a composition of gut microbiota as well as the emergence of antibiotic resistance. The data on the longitudinal recovery of major bacterial phyla and the expansion of genes associated with antibiotic resistance are limited. We collected regular stool samples during the first year after allo-HSCT from 12 adult patients with oncohematological disorders after allo-HSCT and performed 16SrRNA sequencing, multiplex PCR, conventional bacteriology and CHROMagar testing. We observed a decline in Shannon microbiota diversity index as early as day 0 of allo-HSCT (p = 0.034) before any administration of antibiotics, which persisted up to 1 year after transplantation, when the Shannon index returned to pre-transplant levels (p = 0.91). The study confirmed the previously shown decline in Bacillota (Firmicutes) genera and the expansion of E. coli/Shigella, Klebsiella and Enterococci. The recovery of Firmicutes was slower than that of other phyla and occurred only a year post-transplant. A positive correlation was observed between the expansion of E. coli/Shigella genera and blaKPC, blaCTX-M-1 and blaTEM (p < 0.001), Klebsiella spp. and blaOXA-48-like, blaNDM, blaCTX-M-1, blaTEM, and blaSHV (p < 0.001), Pseudomonas spp. and blaNDM (p = 0.002), Enterococcus spp. and blaOXA-48-like, blaNDM, blaCTX-M-1, blaSHV (p < 0.01). The correlation was observed between the expansion of Enterobacterales and and carbapenemase-positive CHROMagar samples (p < 0.001). Samples positive for carbapenem-resitant bacteria were at their maximum levels on day +30, and were gradually diminishing one year after allo-HSCT. From day +30 to +60, all isolated K. pneumoniae strains in fecal samples proved to be resistant to the main antibiotic groups (carbapenems, aminoglycosides, fluoroquinolones, third-generation cephalosporins). One year after HSCT, we documented the spontaneous decolonization of K. pneumoniae. The sensitivity of molecular biology techniques in the search for total and antibiotic-resistant Klebsiella seems to be superior to common bacteriological cultures. Future studies should be focused on searching for novel approaches to the efficient reconstitution and/or maintenance of strictly anaerobic microbiota in oncological patients.

Prevalence of fungal DNAemia mediated by putatively non-pathogenic fungi in immunocompromised patients with febrile neutropenia: a prospective cohort study.

Invasive fungal disease (IFD) presents a life-threatening condition in immunocompromised patients, thus often prompting empirical administration of antifungal treatment, without adequate mycological evidence. Over the past years, wide use of antifungal prophylaxis resulted in decreased occurrence of IFD but has contributed to changes in the spectrum of fungal pathogens, revealing the occurrence of previously rare fungal genera causing breakthrough infections. The expanding spectrum of clinically relevant fungal pathogens required the implementation of screening approaches permitting broad rather than targeted fungus detection to support timely onset of pre-emptive antifungal treatment. To address this diagnostically important aspect in a prospective setting, we analyzed 935 serial peripheral blood (PB) samples from 195 pediatric and adult patients at high risk for IFD, involving individuals displaying febrile neutropenia during treatment of hematological malignancies or following allogeneic hematopoietic stem cell transplantation. Two different panfungal-PCR-screening methods combined with ensuing fungal genus identification by Sanger sequencing were employed. In the great majority of PB-specimens displaying fungal DNAemia, the findings were transient and revealed fungi commonly regarded as non-pathogenic or rarely pathogenic even in the highly immunocompromised patient setting. Hence, to adequately exploit the diagnostic potential of panfungal-PCR approaches for detecting IFD, particularly if caused by hitherto rarely observed fungal pathogens, it is necessary to confirm the findings by repeated testing and to identify the fungal genus present by ensuing analysis. If applied appropriately, panfungal-PCR-screening can help prevent unnecessary empirical therapy, and conversely, contribute to timely employment of effective pre-emptive antifungal treatment strategies.

Level of vascular endothelial growth factor predicts both relapse and nonrelapse mortality after allogeneic hematopoietic stem cell transplantation.

Although the prognostic significance of vascular endothelial growth factor (VEGF) has been researched extensively in patients with hematologic malignancies undergoing chemotherapy, its role in allogeneic hematopoietic stem cell transplantation (HSCT) requires further investigation. The present study evaluated the associations between VEGF level and relapse rate and early complications after HSCT. VEGF levels were analyzed in 91 consecutive patients before the start of conditioning, on day 0, on the day of engraftment, and on the day of diagnosis of veno-occlusive disease (VOD). Compared with a normal level, an elevated high VEGF-A level before conditioning was associated with an increased 2-year relapse rate (55% versus 24%, P = .003; hazard ratio [HR], 3.25; 95% confidence interval [CI], 1.49 to 7.08) and decreased event-free survival (20% versus 44%; P = .022; HR, 2.03; 95% CI, 1.11 to 3.72). No association was found between VEGF level and the incidence of acute GVHD (P > .05). In patients with VOD, VEGF-A level was elevated on day 0 and on the day of VOD diagnosis (P < .05). A low VEGF-A level on day 0 was associated with reduced nonrelapse mortality (14% versus 35%; P = .048; HR, 0.32; 95% CI, 0.10 to 0.99). Our results indicate that a high VEGF-A level before HSCT increases the risk of relapse, and a high level after conditioning is associated with increased risks of early complications and nonrelapse mortality.

A pilot study of implication of machine learning for relapse prediction after allogeneic stem cell transplantation in adults with Ph-positive acute lymphoblastic leukemia.

The posttransplant relapse in Ph-positive ALL increases the risk of death. There is an unmet need for instruments to predict the risk of relapse and plan prophylaxis. In this study, we analyzed posttransplant data by machine learning algorithms. Seventy-four Ph-positive ALL patients with a median age of 30 (range 18-55) years who previously underwent allo-HSCT, were retrospectively enrolled. Ninety-three percent of patients received prophylactic/preemptive TKIs after allo-HSCT. The values of the BCR::ABL1 level at serial assessments and over variables were collected in specified intervals after allo-HSCT. They were used to model relapse risk with several machine-learning approaches. GBM proved superior to the other algorithms and provided a maximal AUC score of 0.91. BCR::ABL1 level before and after allo-HSCT, prediction moment, and chronic GvHD had the highest value in the model. It was shown that after Day + 100, both error rates do not exceed 22%, while before D + 100, the model fails to make accurate predictions. As a result, we determined BCR::ABL1 levels at which the relapse risk remains low. Thus, the current BCR::ABL1 level less than 0.06% in patients with chronic GvHD predicts low risk of relapse. At the same time, patients without chronic GVHD after allo-HSCT should be classified as high risk with any level of BCR::ABL1. GBM model with posttransplant laboratory values of BCR::ABL1 provides a high prediction of relapse after allo-HSCT in the era of TKIs prophylaxis. Validation of this approach is warranted.

ATR-FTIR spectrum analysis of plasma samples for rapid identification of recovered COVID-19 individuals.

The development of fast, cheap and reliable methods to determine seroconversion against infectious agents is of great practical importance. In the context of the COVID-19 pandemic, an important issue is to study the rate of formation of the immune layer in the population of different regions, as well as the study of the formation of post-vaccination immunity in individuals after vaccination. Currently, the main method for this kind of research is enzyme immunoassay (ELISA, enzyme-linked immunosorbent assay). This technique is sufficiently sensitive and specific, but it requires significant time and material costs. We investigated the applicability of attenuated total reflection (ATR) Fourier transform infrared (FTIR) spectroscopy associated with machine learning in blood plasma to detect seroconversion against SARS-CoV-2. The study included samples of 60 patients. Clear spectral differences in plasma samples from recovered COVID-19 patients and conditionally healthy donors were identified using multivariate and statistical analysis. The results showed that ATR-FTIR spectroscopy, combined with principal components analysis (PCA) and linear discriminant analysis (LDA) or artificial neural network (ANN), made it possible to efficiently identify specimens from recovered COVID-19 patients. We built classification models based on PCA associated with LDA and ANN. Our analysis led to 87% accuracy for PCA-LDA model and 91% accuracy for ANN, respectively. Based on this proof-of-concept study, we believe this method could offer a simple, label-free, cost-effective tool for detecting seroconversion against SARS-CoV-2. This approach could be used as an alternative to ELISA.

Towards Understanding the Pathogenicity of DROSHA Mutations in Oncohematology.

Myelodysplastic syndrome (MDS) refers to a heterogeneous group of closely related clonal hematopoietic disorders, which are characterized by accumulation of somatic mutations. The acquired mutation burden is suggested to define the pathway and consequent phenotype of the pathology. Recent studies have called attention to the role of miRNA biogenesis genes in MDS progression; in particular, the mutational pressure of the DROSHA gene was determined. Therefore, this highlights the importance of studying the impact of all collected missense mutations found within the DROSHA gene in oncohematology that might affect the functionality of the protein. In this study, the selected mutations were extensively examined by computational screening, and the most deleterious were subjected to a further molecular dynamic simulation in order to uncover the molecular mechanism of the structural damage to the protein altering its biological function. The most significant effect was found for variants I625K, L1047S, and H1170D, presumably affecting the endonuclease activity of DROSHA. Such alterations arisen during MDS progression should be taken into consideration as evoking certain clinical traits in the malignifying clonal evolution.

A Study of Safety and Efficacy of Nivolumab and Bendamustine (NB) in Patients With Relapsed/Refractory Hodgkin Lymphoma After Nivolumab Monotherapy Failure.

This single-center prospective clinical trial evaluated the combination of nivolumab plus bendamustine (NB) as a salvage regimen in classical Hodgkin lymphoma patients after failure of nivolumab monotherapy. A total of 30 patients received nivolumab (3 mg/kg) on D1,14 and bendamustine (90 mg/m2) on D1, 2 of a 28-day cycle for up to 3 cycles. The ORR was 87% with 57% CR, 30% PR. With median follow-up of 25 months, the estimated 2-year OS was 96,7% (95% CI, 90.2%-100%), PFS was 23,3% (95% CI, 8.2%-38.4%) median PFS was 10.2 months (95% CI, 7.7-14.2 months) with median DOR 6.6 months (95% CI 3.9-11.6 months). Ten patients (33.3%) experienced grade 3 to 4 AE during therapy. Infections were most common AEs of the combined therapy. NB was a highly efficient salvage regimen in relapsed/refractory cHL with a manageable toxicity profile and modest potential for achievement of long-term remission. Registered at www.clinicaltrials.gov (#NCT0334365).

High prevalence of CD3, NK, and NKT cells in the graft predicts adverse outcome after matched-related and unrelated transplantations with post transplantation cyclophosphamide.

The predictive value of graft composition and plasma biomarkers on the outcome of allogeneic HSCT is well known for conventional GVHD prophylaxis based on calcineurin inhibitors with or without antithymocyte globulin. Currently, there is limited data whether these results could be translated to post transplantation cyclophosphamide (PTCy). The prospective extension cohort of NCT02294552 trial enrolled 79 adult patients with acute leukemia in CR. Twenty-six received matched-related bone marrow (BM) grafts with single-agent PTCy and 53 received unrelated peripheral blood stem cell graft (PBSC) with PTCy, tacrolimus, and MMF. The grafts were studied by the flow cytometry, and plasma samples were analyzed by ELISA. In the cluster and major component analysis, we determined that transplantation from donors with high content of CD3, NKT, and CD16-CD56 + subpopulations in the PBSC grafts was associated with poor immunological recovery and compromised event-free survival (50% vs. 80%, HR 2.93, p = 0.015) both due to increased relapse incidence and non-relapse mortality. The significant independent predictor of moderate and severe chronic GVHD was the high prevalence of and iNKT, Vß11, and double-positive cells in the PBSC grafts from young donors (HR 2.75, p = 0.0483). No patterns could be identified for BM grafts and for plasma biomarkers.

A Phase 2 Study of Nivolumab Using a Fixed Dose of 40 mg (Nivo40) in Patients With Relapsed/Refractory Hodgkin Lymphoma.

The introduction of nivolumab has changed the landscape of relapsed/refractory classical Hodgkin lymphoma (r/r cHL) treatment. Despite its clinical importance, this therapy may remain inaccessible for a significant number of patients worldwide, especially in low-income countries, due to its high cost. The results of pharmacokinetic analysis and clinical observations suggest the potential efficacy of low dose nivolumab in r/r cHL patients. The aim of this trial was to assess the efficacy and safety of nivolumab at a fixed dose of 40 mg in patients with r/r cHL. The study included 30 patients with r/r cHL, treated with 40 mg nivolumab every 2 weeks. The median dose of nivolumab per kilogram bodyweight was 0.59 mg/kg (0.4-1 mg/kg). Median follow up was 19.2 months (range 12.7-25.4). The objective response rate was 70%, with 13 (43.3%) patients achieving a complete response. Median PFS was 18.4 months (95% CI, 11.3 to 18.5 months) with 18-month PFS of 53.6% (95% CI, 32%-71%). At the time of analysis, 96.7% of patients were alive with a median OS not reached. Severe (grade 3-5) adverse events were observed in 4 patients (13.3%). Nivolumab in a fixed dose of 40 mg was efficient in patients with r/r cHL, independent from dose per kg bodyweight. The results of this study are in good agreement with previously reported data and create a rationale for further studies aimed to define the optimal dosing regimen of nivolumab for the treatment of r/r cHL. Registered at www.clinicaltrials.gov (NCT03343665).

Clinical and morphological practices in the diagnosis of transplant-associated microangiopathy: a study on behalf of Transplant Complications Working Party of the EBMT.

Transplant-associated thrombotic microangiopathy (TA-TMA) is a life-threatening complication of allogeneic hematopoietic stem cell transplantation (HSCT). This study evaluated clinical and morphological practices of TA-TMA diagnosis in EBMT centers. Two questionnaires, one for transplant physician and one for morphologist, and also a set of electronic blood slides from 10 patients with TA-TMA and 10 control patients with various erythrocyte abnormalities, were implemented for evaluation. Seventeen EBMT centers participated in the study. Regarding criteria used for TA-TMA diagnosis, centers reported as follows: 41% of centers used the International Working Group (IWG) criteria, 41% used "overall TA-TMA" criteria and 18% used physician's decision. The threshold of schistocytes to establish TA-TMA diagnosis in the participating centers was significantly associated with morphological results of test cases evaluations (p = 0.002). The mean number of schistocytes reported from blood slide analyses were 4.3 ± 4.5% for TA-TMA cases (range 0-19.6%, coefficient of variation (CV) 0.7) and 1.3 ± 1.6% for control cases (range 0-8.3%, CV 0.8). Half of the centers reported schistocyte levels below 4% for 7/10 TA-TMA cases. The intracenter variability was low, indicating differences in the institutional practices of morphological evaluation. In conclusion, the survey identified the need for the standardization of TA-TMA morphological diagnosis.

Clinical Correlates and Prognostic Significance of IL-8, sIL-2R, and Immunoglobulin-Free Light Chain Levels in Patients with Myelofibrosis.

BACKGROUND: Chronic myeloproliferative neoplasms are characterized by clonal hematopoiesis and persistent inflammatory reaction. In this study, the clinical significance and prognostic impact of several inflammatory markers were evaluated in patients with BCR/ABL-negative myeloproliferative malignancies. METHODS: Serum levels of interleukin-8 (IL-8) and lymphoid-associated activation markers - soluble interleukin-2 receptor (sIL-2R) and immunoglobulin-free light chains (FLC) - were evaluated in patients with primary myelofibrosis (MF), post-polycythemia vera MF, and post-essential thrombocythemia MF, and compared with the levels in healthy donors. RESULTS: In 57 MF patients, sIL-2R excess correlated with transfusion-dependent anemia (p = 0.03) and splenomegaly (p = 0.02). There were no statistically significant correlations between sIL-2R and IL-8 levels, but the plasma concentration of κ-FLC positively correlated with the IL-8 level (p = 0.027). In univariate analysis, increased levels of IL-8 (p = 0.016) and sIL-2R (p = 0.010) significantly reduced 1-year overall survival. Only elevated sIL-2R rate retained significance (p = 0.02) in multivariate analysis when Dynamic International Prognostic Scoring System plus (DIPSSplus) risk stratification was added. CONCLUSION: We observed an association between FLC and proinflammatory cytokine hyperexpression. Serum cytokine levels and FLC might be a promising approach to predicting and monitoring treatment response in MF patients.

Mesenchymal Stem Cell Magnetization: Magnetic Multilayer Microcapsule Uptake, Toxicity, Impact on Functional Properties, and Perspectives for Magnetic Delivery.

Mesenchymal stem cells (MSCs) are widely used in cell therapy due to their convenience, multiline differentiation potential, reproducible protocols, and biological properties. The potential of MSCs to impregnate magnetic microcapsules and their possible influence on cell function and ability to response to magnetic field have been explored. Interestingly, the cells suspended in media show much higher ability in internalization of microcapsules, then MSCs adhere into the surface. There is no significant effect of microcapsules on cell toxicity compared with other cell line-capsule internalization reported in literature. Due to internalization of magnetic capsules by the cells, such cell engineering platform is responsive to external magnetic field, which allows to manipulate MSC migration. Magnetically sorted MSCs are capable to differentiation as confirmed by their conversion to adipogenic and osteogenic cells using standard protocols. There is a minor effect of capsule internalization on cell adhesion, though MSCs are still able to form spheroid made by dozen of thousand MSCs. This work demonstrates the potential of use of microcapsule impregnated MSCs to carry internalized micron-sized vesicles and being navigated with external magnetic signaling.