The antiviral response triggered by the cGAS/STING pathway is subverted by the foot-and-mouth disease virus proteases.

Propagation of viruses requires interaction with host factors in infected cells and repression of innate immune responses triggered by the host viral sensors. Cytosolic DNA sensing pathway of cyclic GMP-AMP synthase (cGAS) and stimulator of interferon genes (STING) is a major component of the antiviral response to DNA viruses, also known to play a relevant role in response to infection by RNA viruses, including foot-and-mouth disease virus (FMDV). Here, we provide supporting evidence of cGAS degradation in swine cells during FMDV infection and show that the two virally encoded proteases, Leader (Lpro) and 3Cpro, target cGAS for cleavage to dampen the cGAS/STING-dependent antiviral response. The specific target sequence sites on swine cGAS were identified as Q140/T141 for the FMDV 3Cpro and the KVKNNLKRQ motif at residues 322-330 for Lpro. Treatment of swine cells with inhibitors of the cGAS/STING pathway or depletion of cGAS promoted viral infection, while overexpression of a mutant cGAS defective for cGAMP synthesis, unlike wild type cGAS, failed to reduce FMDV replication. Our findings reveal a new mechanism of RNA viral antagonism of the cGAS-STING innate immune sensing pathway, based on the redundant degradation of cGAS through the concomitant proteolytic activities of two proteases encoded by an RNA virus, further proving the key role of cGAS in restricting FMDV infection.

Cytomegalovirus Reactivations in Allogeneic Hematopoietic Stem Cell Transplantation from HLA-Matched and Haploidentical Donors with Post-Transplantation Cyclophosphamide.

Cytomegalovirus (CMV) reactivations cause significant morbidity in allogeneic hematopoietic stem cell transplantation (HSCT) recipients. Graft-versus-host disease (GVHD) prophylaxis with post-transplantation cyclophosphamide (PTCy) is associated with an increased risk of CMV infections. Data are limited comparing HSCT with PTCy performed from matched sibling donors (MSDs), matched unrelated donors (MUDs), and haploidentical (Haplo) donors. In the present study, we aimed to characterize CMV reactivation and recurrence in patients with hematologic malignancies undergoing HSCT from MSD, MUD, and Haplo donors using PTCy as GVHD prophylaxis in the pre-letermovir era. We also analyzed risk factors of CMV reactivation, including GVHD as a time-dependent variable, on the incidence and mortality associated with CMV infections. We analyzed CMV reactivation in patients undergoing HSCT from 160 MSDs, 124 MUDs, and 82 Haplo donors from a single institution. Uniform GVHD prophylaxis with PTCy, sirolimus, and mycophenolate mofetil was given irrespective of donor type. Overall, 46% of patients had at least 1 CMV reactivation. The 1-year cumulative incidence of CMV infection was 39% for MSD, 44% for MUD, and 62% for Haplo donors (P < .001), with 96% of reactivations occurring before day +100. Multivariate analysis identified factors associated with the first CMV reactivation, including Haplo donor, positive recipient CMV serology, older patient age, and grade II-IV acute GVHD. The 1-year cumulative incidence of second reactivation from HSCT was 13%. Recipient CMV seropositivity, older patient age, and grade II-IV acute GVHD, but not type of donor, were identified as adverse factors for second CMV reactivation in multivariate analysis. The 1-year cumulative incidence of a third reactivation post HSCT was 4.4%. Ten cases of CMV disease were recorded, with no attributable deaths. Nevertheless, the risk for nonrelapse mortality was greater for patients who experienced CMV reactivation in multivariate time-dependent Cox model analysis. CMV reactivation is frequent in HSCT with PTCy in patients not receiving letermovir prophylaxis. Identified risk factors include the use of a Haplo donor, recipient CMV seropositivity, and grade II-IV acute GVHD. The prevalence of recurrent CMV reactivations is a noteworthy issue, especially after acute GVHD, warranting trials of secondary prophylaxis strategies.

A simple agent-based hybrid model to simulate the biophysics of glioblastoma multiforme cells and the concomitant evolution of the oxygen field.

BACKGROUND AND OBJECTIVES: Glioblastoma multiforme (GBM) is one of the most aggressive cancers of the central nervous system. It is characterized by a high mitotic activity and an infiltrative ability of the glioma cells, neovascularization and necrosis. GBM evolution entails the continuous interplay between heterogeneous cell populations, chemotaxis, and physical cues through different scales. In this work, an agent-based hybrid model is proposed to simulate the coupling of the multiscale biological events involved in the GBM invasion, specifically the individual and collective migration of GBM cells and the concurrent evolution of the oxygen field and phenotypic plasticity. An asset of the formulation is that it is conceptually and computationally simple but allows to reproduce the complexity and the progression of the GBM micro-environment at cell and tissue scales simultaneously. METHODS: The migration is reproduced as the result of the interaction between every single cell and its micro-environment. The behavior of each individual cell is formulated through genotypic variables whereas the cell micro-environment is modeled in terms of the oxygen concentration and the cell density surrounding each cell. The collective behavior is formulated at a cellular scale through a flocking model. The phenotypic plasticity of the cells is induced by the micro-environment conditions, considering five phenotypes. RESULTS: The model has been contrasted by benchmark problems and experimental tests showing the ability to reproduce different scenarios of glioma cell migration. In all cases, the individual and collective cell migration and the coupled evolution of both the oxygen field and phenotypic plasticity have been properly simulated. This simple formulation allows to mimic the formation of relevant hallmarks of glioblastoma multiforme, such as the necrotic cores, and to reproduce experimental evidences related to the mitotic activity in pseudopalisades. CONCLUSIONS: In the collective migration, the survival of the clusters prevails at the expense of cell mitosis, regardless of the size of the groups, which delays the formation of necrotic foci and reduces the rate of oxygen consumption.

PCR-Based Strategy for Introducing CRISPR/Cas9 Machinery into Hematopoietic Cell Lines.

Acute myeloid leukemia is a complex heterogeneous disease characterized by the clonal expansion of undifferentiated myeloid precursors. Due to the difficulty in the transfection of blood cells, several hematological models have recently been developed with CRISPR/Cas9, using viral vectors. In this study, we developed an alternative strategy in order to generate CRISPR constructs by fusion PCR, which any lab equipped with basic equipment can implement. Our PCR-generated constructs were easily introduced into hard-to-transfect leukemic cells, and their function was dually validated with the addition of MYBL2 and IDH2 genes into HEK293 cells. We then successfully modified the MYBL2 gene and introduced the R172 mutation into the IDH2 gene within NB4 and HL60 cells that constitutively expressed the Cas9 nuclease. The efficiency of mutation introduction with our methodology was similar to that of ribonucleoprotein strategies, and no off-target events were detected. Overall, our strategy represents a valid and intuitive alternative for introducing desired mutations into hard-to-transfect leukemic cells without viral transduction.

Reduced Incidence of Neurologic Complications after Allogeneic Hematopoietic Stem Cell Transplantation with Calcineurin-Free Graft-versus-Host Disease Prophylaxis.

Calcineurin inhibitors (CNIs), including cyclosporine and tacrolimus, are frequently associated with neurologic complications after allogeneic hematopoietic stem cell transplantation (HSCT). However, there is a lack of studies comparing the incidence and characteristics of neurologic complications in patients undergoing HSCT based on CNI-free or CNI-based GVHD prophylaxis. This retrospective single-center study analyzed the neurologic complications in 2 cohorts of patients undergoing HSCT with either CNI-based GVHD prophylaxis (n = 523) or CNI-free prophylaxis with post-transplantation cyclophosphamide, sirolimus, and mycophenolate mofetil (n = 371). The latter cohort included older patients and received more reduced-intensity conditioning and transplants from matched unrelated and haploidentical donors. The 2-year cumulative incidence of neurologic complications was significantly lower in the CNI-free cohort (6.9% versus 11.9%; P = .016), and GVHD prophylaxis was the sole statistically significant variable in multivariate analysis (hazard ratio, 2.2; 95% confidence interval [CI], .25 to 3.13; P = .0017). The distribution of neurologic types was similar in the 2 cohorts, with encephalopathy the most prevalent complication, except for headaches and myopathy, which decreased equally from 15% in the CNI-based cohort to 4% in the CNI-free cohort. Neurologic complications had negative impacts on mortality and survival rates, with a significantly higher 2-year cumulative incidence of nonrelapse mortality (NRM) (44% [95% CI, 34% to 54%] versus 16% [95% CI, 13% to 18%]; P < .0001) and inferior overall survival (66% [95% CI, 62% to 69%] versus 46% [95% CI, 37% to 58%]; P < .0001) in patients with neurologic complications. This study suggests that CNI-free GVHD prophylaxis with post-transplantation cyclophosphamide, sirolimus, and mycophenolate mofetil may reduce not only the incidence of GVHD incidence, but also the rates of neurologic complications and NRM, leading to improved survival outcomes in patients undergoing HSCT.

Transfusion Burden in Allogeneic Hematopoietic Stem Cell Transplantation over Time: Experience from a Single Institution.

INTRODUCTION: Transfusion plays a main role in supportive treatment for patients who receive an allogeneic hematopoietic stem cell transplantation (HSCT). In this study, we compare the transfusion requirements of patients undergoing different modalities of HSCT according to different time periods. The objective is to assess the evolution of HSCT transfusion requirements over time, from a single institution. METHODS: The clinical charts and transfusion records of patients who underwent HSCT of different modalities at La Fe University Hospital during a twelve-year period were reviewed (2009-2020). For analysis, we divided the overall time into three periods: 1 from 2009 to 2012, 2 from 2013 to 2016 and 3 from 2017 to 2020. The study included 855 consecutive adult HSCT: 358 HLA-matched related donors (MRD), 134 HLA-matched unrelated donors (MUD), 223 umbilical cord blood transplantation (UCBT) and 140 haploidentical transplants (Haplo-HSCT). RESULTS: There were no significant differences in RBC and PLT requirements or transfusion independence among the three time periods for MUD and Haplo-HSCT. However, the transfusion burden increased significantly for MRD HSCT during the 2017-2020 period. CONCLUSION: despite HSCT modalities having evolved and changed over time, overall transfusion requirements have not significantly decreased and continue to be a cornerstone of transplantation-supportive care.

GAM: General Auxetic Metamaterial with Tunable 3D Auxetic Behavior Using the Same Unit Cell Boundary Connectivity.

Research on auxetic metamaterials is important due to their high performance against impact loadings and their usefulness in actuators, among other applications. These metamaterials offer a negative Poisson's ratio at the macro level. However, usual auxetic metamaterials face challenges in (1) grading the effect, (2) coupling and combining auxetic metamaterials with non-auxetic materials due to boundary compatibility, (3) obtaining the same auxetic behavior in all directions in the transverse plane, and (4) adapting the regular geometry to the component design boundary and shape. The goal of this paper is to present a novel, recently patented tunable 3D metamaterial created to reproduce a wide spectrum of 3D auxetic and non-auxetic Poisson's ratios and Young's moduli. This wide range is obtained using the same basic unit cell geometry and boundary connections with neighboring cells, facilitating designs using functionally graded metamaterials as only the connectivity and position of the cell's internal nodes are modified. Based on simple spatial triangularization, the metamaterial is easily scalable and better accommodates spatial curvatures or boundaries by changing the locations of nodes and lengths of bars.

Impact of Post-Transplantation Cyclophosphamide on Transfusion Requirements in HLA-Matched Sibling Peripheral Blood Stem Cell Transplantation.

Post-transplantation cyclophosphamide (PTCy)-based graft-versus-host disease (GVHD) prophylaxis is being increasingly used in allogeneic hematopoietic stem cell transplantation (allo-HSCT) from HLA-matched related donors (MRDs); however, information regarding the transfusion needs in this setting is lacking. This study compared RBC and platelet units transfused and time to transfusion independence according to the GVHD prophylaxis regimen in MRD HSCT. We performed a matched-pair analysis comparing the transfusion requirements and the clinical outcomes of patients who underwent MRD peripheral blood HSCT using PTCy between January 2017 and June 2021 (n = 100) with historical MRD HSCTs using standard cyclosporine A (CsA)-based prophylaxis (n = 100). Neutrophil engraftment was significantly delayed in the PTCy group compared with the CsA group (16 days versus 13 days; P = .003). PTCy was associated with increased RBC (median, 5 units versus 4 units; P = .04) and platelet (median, 6 units versus 3 units; P = .01) transfusion requirements during the first 30 days after transplantation. The proportion of patients requiring platelet transfusion during days 31 to 90 after transplantation was also higher in the PTCy group (55% versus 25%; P < .0001). In multivariate analysis, PTCy was associated with delayed RBC and platelet transfusion independence (hazard ratio, .61 [P = .007] and .51 [P < .0001], respectively). The cumulative incidence (CuI) of BK polyomavirus-associated hemorrhagic cystitis grade ≥2 at 100 days was higher in the PTCy group (34% versus 12%; P < .0001); however, the PTCy group had lower rates of grade II-IV acute GVHD (100-day CuI, 57% versus 23%; P < .0001) and moderate to severe chronic GVHD (1-year CuI, 49% versus 28%; P = .003), as well as better 2-year overall survival (74% versus 56%; P = .01). Our study shows that although PTCy increases the transfusion burden in MRD HSCT, it is associated with a low incidence of severe GVHD and with encouraging survival outcomes.

Glomerulonephritis Secondary to Acute Promyelocytic Leukemia that Resolved after Induction Therapy.

The association of glomerulonephritis and malignant hematological disease is uncommon, but well known in chronic leukemias, lymphomas, and monoclonal gammopathies. However, only a few cases of glomerulonephritis and acute myeloid leukemia have been reported in the literature. We describe the first case of a genetically diagnosed acute promyelocytic leukemia presenting with nephrotic range proteinuria that resolved with induction therapy with ATRA and ATO and performed a comprehensive review of the literature.

Integrated flow cytometry and sequencing to reconstruct evolutionary patterns from dysplasia to acute myeloid leukemia.

Clonal evolution in acute myeloid leukemia (AML) originates long before diagnosis and is a dynamic process that may affect survival. However, it remains uninvestigated during routine diagnostic workups. We hypothesized that the mutational status of bone marrow dysplastic cells and leukemic blasts, analyzed at the onset of AML using integrated multidimensional flow cytometry (MFC) immunophenotyping and fluorescence-activated cell sorting (FACS) with next-generation sequencing (NGS), could reconstruct leukemogenesis. Dysplastic cells were detected by MFC in 285 of 348 (82%) newly diagnosed patients with AML. Presence of dysplasia according to MFC and World Health Organization criteria had no prognostic value in older adults. NGS of dysplastic cells and blasts isolated at diagnosis identified 3 evolutionary patterns: stable (n = 12 of 21), branching (n = 4 of 21), and clonal evolution (n = 5 of 21). In patients achieving complete response (CR), integrated MFC and FACS with NGS showed persistent measurable residual disease (MRD) in phenotypically normal cell types, as well as the acquisition of genetic traits associated with treatment resistance. Furthermore, whole-exome sequencing of dysplastic and leukemic cells at diagnosis and of MRD uncovered different clonal involvement in dysplastic myelo-erythropoiesis, leukemic transformation, and chemoresistance. Altogether, we showed that it is possible to reconstruct leukemogenesis in ∼80% of patients with newly diagnosed AML, using techniques other than single-cell multiomics.

Post-transplant cyclophosphamide and sirolimus based graft-versus-host disease prophylaxis after allogeneic stem cell transplantation for acute myeloid leukemia.

Post-transplant cyclophosphamide (PTCy) has emerged as a promising graft-versus-host disease (GvHD) prophylaxis in allogeneic hematopoietic stem cell transplantation (allo-HSCT). However, no studies have reported the efficacy of a GvHD prophylaxis based on PTCy with sirolimus (Sir-PTCy) in patients with acute myeloid leukemia (AML). In this retrospective study, we analyze the use of sirolimus in combination with PTCy, with or without mycophenolate mofetil (MMF), on 242 consecutive adult patients with AML undergoing a myeloablative first allo-HSCT from different donor types, in three European centers between January 2017 and December 2020. Seventy-seven (32%) patients received allo-HSCT from HLA-matched sibling donor, 101 (42%) from HLA-matched and mismatched unrelated donor, and 64 (26%) from haploidentical donor. Except for neutrophil and platelet engraftment, which was slower in the haploidentical cohort, no significant differences were observed in major transplant outcomes according to donor type in univariate and multivariate analysis. GvHD prophylaxis with Sir-PTCy, with or without MMF, is safe and effective in patients with AML undergoing myeloablative allo-HSCT, resulting in low rates of transplant-related mortality, relapse/progression, and acute and chronic GvHD in all donor settings.

Use of Venetoclax in Patients with Relapsed or Refractory Acute Myeloid Leukemia: The PETHEMA Registry Experience.

The effectiveness of venetoclax (VEN) in relapsed or refractory acute myeloid leukemia (RR-AML) has not been well established. This retrospective, multicenter, observational database studied the effectiveness of VEN in a cohort of 51 RR-AML patients and evaluated for predictors of response and overall survival (OS). The median age was 68 years, most were at high risk, 61% received ≥2 therapies for AML, 49% had received hypomethylating agents, and ECOG was ≥2 in 52%. Complete remission (CR) rate, including CR with incomplete hematological recovery (CRi), was 12.4%. Additionally, 10.4% experienced partial response (PR). The CR/CRi was higher in combination with azacitidine (AZA; 17.9%) than with decitabine (DEC; 6.7%) and low-dose cytarabine (LDAC; 0%). Mutated NPM1 was associated with increased CR/CRi. Median OS was 104 days (95% CI: 56-151). For the combination with AZA, DEC, and LDAC, median OS was 120 days, 104 days, and 69 days, respectively; p = 0.875. Treatment response and ECOG 0 influenced OS in a multivariate model. A total of 28% of patients required interruption of VEN because of toxicity. Our real-life series describes a marginal probability of CR/CRi and poor OS after VEN-based salvage. Patients included had very poor-risk features and were heavily pretreated. The small percentage of responders did not reach the median OS.

Characteristics, clinical outcomes, and risk factors of SARS-COV-2 infection in adult acute myeloid leukemia patients: experience of the PETHEMA group.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection produces higher morbidity and mortality in hematological malignancies, but evidence in acute myeloid leukemia (AML) is scarce. A multicenter observational study was conducted to determine the clinical outcomes and assess the impact of therapeutic approaches in adult AML patients with SARS-CoV-2 infection in the first wave (March-May 2020). Overall, 108 patients were included: 51.9% with active leukemia and 70.4% under therapeutic schedules for AML. Signs and symptoms of SARS-CoV-2 were present in 96.3% of patients and 82.4% received specific treatment for SARS-CoV-2. The mortality rate was 43.5% and was correlated with age, gender, active leukemia, dyspnea, severe SARS-CoV-2, intensive care measures, neutrophil count, and D-dimer levels. A protective effect was found with azithromycin, lopinavir/ritonavir, and normal liver enzyme levels. During the SARS-CoV-2 first wave, our findings suggested an increased mortality in AML in a short period. SARS-CoV-2 management could be guided by risk factors in AML patients.

Correction: Liquori et al. Acute Promyelocytic Leukemia: A Constellation of Molecular Events around a Single PML-RARA Fusion Gene. Cancers 2020, 12, 624.

The authors wish to make the following corrections to this paper [...].

The Mutational Landscape of Acute Myeloid Leukaemia Predicts Responses and Outcomes in Elderly Patients from the PETHEMA-FLUGAZA Phase 3 Clinical Trial.

We sought to predict treatment responses and outcomes in older patients with newly diagnosed acute myeloid leukemia (AML) from our FLUGAZA phase III clinical trial (PETHEMA group) based on mutational status, comparing azacytidine (AZA) with fludarabine plus low-dose cytarabine (FLUGA). Mutational profiling using a custom 43-gene next-generation sequencing panel revealed differences in profiles between older and younger patients, and several prognostic markers that were useful in young patients were ineffective in older patients. We examined the associations between variables and overall responses at the end of the third cycle. Patients with mutated DNMT3A or EZH2 were shown to benefit from azacytidine in the treatment-adjusted subgroup analysis. An analysis of the associations with tumor burden using variant allele frequency (VAF) quantification showed that a higher overall response was associated with an increase in TET2 VAF (odds ratio (OR), 1.014; p = 0.030) and lower TP53 VAF (OR, 0.981; p = 0.003). In the treatment-adjusted multivariate survival analyses, only the NRAS (hazard ratio (HR), 1.9, p = 0.005) and TP53 (HR, 2.6, p = 9.8 × 10-7) variants were associated with shorter overall survival (OS), whereas only mutated BCOR (HR, 3.6, p = 0.0003) was associated with a shorter relapse-free survival (RFS). Subgroup analyses of OS according to biological and genomic characteristics showed that patients with low-intermediate cytogenetic risk (HR, 1.51, p = 0.045) and mutated NRAS (HR, 3.66, p = 0.047) benefited from azacytidine therapy. In the subgroup analyses, patients with mutated TP53 (HR, 4.71, p = 0.009) showed a better RFS in the azacytidine arm. In conclusion, differential mutational profiling might anticipate the outcomes of first-line treatment choices (AZA or FLUGA) in older patients with AML. The study is registered at ClinicalTrials.gov as NCT02319135.

Long-Term Outcomes After Autologous Versus Allogeneic Stem Cell Transplantation in Molecularly-Stratified Patients With Intermediate Cytogenetic Risk Acute Myeloid Leukemia: A PETHEMA Study.

Acute myeloid leukemia (AML) with intermediate risk cytogenetics (IRcyto) comprises a variety of biological entities with distinct mutational landscapes that translate into differential risks of relapse and prognosis. Optimal postremission therapy choice in this heterogeneous patient population is currently unsettled. In the current study, we compared outcomes in IRcyto AML recipients of autologous (autoSCT) (n = 312) or allogeneic stem cell transplantation (alloSCT) (n = 279) in first complete remission (CR1). Molecular risk was defined based on CEBPA, NPM1, and FLT3-ITD mutational status, per European LeukemiaNet 2017 criteria. Five-year overall survival (OS) in patients with favorable molecular risk (FRmol) was 62% (95% confidence interval [CI], 50-72) after autoSCT and 66% (95% CI, 41-83) after matched sibling donor (MSD) alloSCT (P = .68). For patients of intermediate molecular risk (IRmol), MSD alloSCT was associated with lower cumulative incidence of relapse (P < .001), as well as with increased nonrelapse mortality (P = .01), as compared to autoSCT. The 5-year OS was 47% (95% CI, 34-58) after autoSCT and 70% (95% CI, 59-79) after MSD alloSCT (P = .02) in this patient subgroup. In a propensity-score matched IRmol subcohort (n = 106), MSD alloSCT was associated with superior leukemia-free survival (hazard ratio [HR] 0.33, P = .004) and increased OS in patients alive 1 year after transplantation (HR 0.20, P = .004). These results indicate that, within IRcyto AML in CR1, autoSCT may be a valid option for FRmol patients, whereas MSD alloSCT should be the preferred postremission strategy in IRmol patients.

Influence of polymorphisms in anthracyclines metabolism genes in the standard induction chemotherapy of acute myeloid leukemia.

OBJECTIVES: Genetic variability in anthracycline metabolism could modify the response and safety of acute myeloid leukemia (AML) induction. METHODS: Polymorphisms in genes that encodes enzymes of anthracyclines metabolic pathway (CBR3: rs1056892, rs8133052, NQO1: rs1800566, NQO2: rs1143684, NOS3: rs1799983, rs2070744) were evaluated in 225 adult de novo AML patients. RESULTS: The variant CBR3 rs8133052 was associated with lower hepatotoxicity (P = 0.028). Wild-type genotype of NQO2 rs1143684 was related to higher complete remission (P = 0.014), and the variant allele with greater gastrointestinal toxicity (P = 0.024). However, the variant genotype of NQO1 rs1800566 was associated with mucositis (P = 0.018), but heterozygous genotype showed less gastrointestinal toxicity (P = 0.028) and thrombocytopenia (P = 0.009). Protective effects against nephrotoxicity and thrombocytopenia were reported with variant NOS3 rs1799983 (P = 0.006, P = 0.014), whereas carriers of NOS3 rs2070744 showed higher hepatotoxicity and thrombocytopenia (P = 0.017, P = 0.013). CONCLUSIONS: This study supports the influence of genetic variability of idarubicin metabolizing could be critical in predicting anthracycline-induced toxicities.

Adoptive transfer of ex vivo expanded SARS-CoV-2-specific cytotoxic lymphocytes: A viable strategy for COVID-19 immunosuppressed patients?

Cellular and humoral response to acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections is on focus of research. We evaluate herein the feasibility of expanding virus-specific T cells (VST) against SARS-CoV-2 ex vivo through a standard protocol proven effective for other viruses. The experiment was performed in three different donors' scenarios: (a) SARS-CoV-2 asymptomatic infection/negative serology, (b) SARS-CoV-2 symptomatic infection/positive serology, and (c) no history of SARS-CoV-2 infection/negative serology. We were able to obtain an expanded VST product from donors 1 and 2 (1.6x and 1.8x increase of baseline VST count, respectively) consisting in CD3 + cells (80.3% and 62.7%, respectively) with CD4 + dominance (60% in both donors). Higher numbers of VST were obtained from the donor 2 as compared to donor 1. T-cell clonality test showed oligoclonal reproducible peaks on a polyclonal background for both donors. In contrast, VST could be neither expanded nor primed in a donor without evidence of prior infection. This proof-of-concept study supports the feasibility of expanding ex vivo SARS-CoV-2-specific VST from blood of convalescent donors. The results raise the question of whether the selection of seropositive donors may be a strategy to obtain cell lines enriched in their SARS-CoV-2-specificity for future adoptive transfer to immunosuppressed patients.

Central Nervous System Involvement in Epstein-Barr Virus-Related Post-Transplant Lymphoproliferative Disorders after Allogeneic Hematopoietic Stem Cell Transplantation.

Central nervous system (CNS) involvement in Epstein-Barr virus-related post-transplant lymphoproliferative disorders (EBV-PTLDs) after allogeneic hematopoietic stem cell transplantation (allo-HSCT) is poorly defined. We analyzed the incidence, clinical and pathological characteristics, and impact on outcomes of EBV-PTLDs with CNS involvement (CNS-PTLDs) in 1009 consecutive adult patients undergoing allo-HSCT at a single-center institution. Four hundred eighty-two patients received matched sibling donor (MSD) transplants, 388 umbilical cord blood transplants (UCBTs), 56 matched unrelated donor (MUD) transplants, and 83 haploidentical transplants. We detected 25 cases of biopsy-proven EBV-PTLDs. Of these, nine patients (36%) had CNS-PTLDs: six after UCBT (67%), one after MSD transplantation (11%), one after MUD transplantation (11%), and one after haploidentical transplantation (11%). The 5-year cumulative incidence risk of CNS-PTLDs was 0.9%. Median time from transplant to CNS-PTLDs was 187 days, and all patients had neurological symptoms at diagnosis. Six out of the nine cases (67%) occurred with systemic involvement, and three cases (33%) had isolated CNS involvement. The most frequent histological subtype was monomorphic EBV-PTLD, and laboratory characteristics were similar to EBV-PTLDs without CNS involvement. We observed statistical differences in the rate of positive EBV DNA detection in plasma between isolated CNS-PTLDs (detection in one out of three, 33%) and the rest of the EBV-PTLDs (100%) (P = .01). Treatment strategies included chemotherapy, radiotherapy, and T cell therapy. However, seven out of nine patients died due to progression of the CNS-PTLDs at a median time of 17 days (range, 8 to 163) from diagnosis. The 5-years overall survival in patients who developed CNS-PTLDs was 22% (95% confidence interval [CI], 7% to 75%) and 5-year treatment-related mortality was 78% (95% CI, 51% to 100%), with no statistically significant differences between CNS-PTLDs and the rest of the EBV-PTLDs. In conclusion, despite advances in EBV monitoring and treatment strategies, CNS-PTLDs remain an uncommon but serious complication after allo-HSCT, with very poor prognosis.