CAR affinity modulates the sensitivity of CAR-T cells to PD-1/PD-L1-mediated inhibition.

Chimeric antigen receptor (CAR)-T cell therapy for solid tumors faces significant hurdles, including T-cell inhibition mediated by the PD-1/PD-L1 axis. The effects of disrupting this pathway on T-cells are being actively explored and controversial outcomes have been reported. Here, we hypothesize that CAR-antigen affinity may be a key factor modulating T-cell susceptibility towards the PD-1/PD-L1 axis. We systematically interrogate CAR-T cells targeting HER2 with either low (LA) or high affinity (HA) in various preclinical models. Our results reveal an increased sensitivity of LA CAR-T cells to PD-L1-mediated inhibition when compared to their HA counterparts by using in vitro models of tumor cell lines and supported lipid bilayers modified to display varying PD-L1 densities. CRISPR/Cas9-mediated knockout (KO) of PD-1 enhances LA CAR-T cell cytokine secretion and polyfunctionality in vitro and antitumor effect in vivo and results in the downregulation of gene signatures related to T-cell exhaustion. By contrast, HA CAR-T cell features remain unaffected following PD-1 KO. This behavior holds true for CD28 and ICOS but not 4-1BB co-stimulated CAR-T cells, which are less sensitive to PD-L1 inhibition albeit targeting the antigen with LA. Our findings may inform CAR-T therapies involving disruption of PD-1/PD-L1 pathway tailored in particular for effective treatment of solid tumors.

Biomarkers of Efficacy and Safety of the Academic BCMA-CART ARI0002h for the Treatment of Refractory Multiple Myeloma.

PURPOSE: B-cell maturation antigen (BCMA)-chimeric antigen receptor T-cells (CART) improve results obtained with conventional therapy in the treatment of relapsed/refractory multiple myeloma. However, the high demand and expensive costs associated with CART therapy might prove unsustainable for health systems. Academic CARTs could potentially overcome these issues. Moreover, response biomarkers and resistance mechanisms need to be identified and addressed to improve efficacy and patient selection. Here, we present clinical and ancillary results of the 60 patients treated with the academic BCMA-CART, ARI0002h, in the CARTBCMA-HCB-01 trial. PATIENTS AND METHODS: We collected apheresis, final product, peripheral blood and bone marrow samples before and after infusion. We assessed BCMA, T-cell subsets, CART kinetics and antibodies, B-cell aplasia, cytokines, and measurable residual disease by next-generation flow cytometry, and correlated these to clinical outcomes. RESULTS: At cut-off date March 17, 2023, with a median follow-up of 23.1 months (95% CI, 9.2-37.1), overall response rate in the first 3 months was 95% [95% confidence interval (CI), 89.5-100]; cytokine release syndrome (CRS) was observed in 90% of patients (5% grades ≥3) and grade 1 immune effector cell-associated neurotoxicity syndrome was reported in 2 patients (3%). Median progression-free survival was 15.8 months (95% CI, 11.5-22.4). Surface BCMA was not predictive of response or survival, but soluble BCMA correlated with worse clinical outcomes and CRS severity. Activation marker HLA-DR in the apheresis was associated with longer progression-free survival and increased exhaustion markers correlated with poorer outcomes. ARI0002h kinetics and loss of B-cell aplasia were not predictive of relapse. CONCLUSIONS: Despite deep and sustained responses achieved with ARI0002h, we identified several biomarkers that correlate with poor outcomes.

EASIX and cardiac adverse events after allogeneic hematopoietic cell transplantation.

This study investigates the interaction between endothelial activation, indirectly measured using EASIX, and the probability of presenting cardiac adverse events (CAE) during the first year after allo-HCT. The 437 consecutive adults undergoing PB allo-HCT from 2012 and 2021 were included. EASIX was retrospectively calculated before and during the first 6 months after allo-HCT and transformed to log2-base to conduct the statistical analysis. The median age was 53, 46 (10.5%) patients had previous history of cardiac disease, MAC allo-HCTs were performed in 186 (42.6%) patients, and PTCY was administered in 242 (55.5%). The 1-year incidence of CAE was 12.6% (n = 55). The most prevalent cardiac events were heart failure and arrhythmias, 32.7% and 23.6% respectively, and the day +100 mortality rate of these patients was 40.5%. During the first 6 months after allo-HCT, EASIX trends were significantly higher in patients who developed CAE. Regression analyses confirmed that higher log2-EASIX values were predictors for higher risk for CAE during the first year after allo-HCT. This analysis identifies a significant association between higher endothelial activation, indirectly measured using EASIX, and higher risk for cardiac toxicity diagnosed during the first year after allo-HCT and extends the applicability of EASIX for identifying patients at risk for CAE.

Endothelial Activation and Stress Index in adults undergoing allogeneic hematopoietic cell transplantation with post-transplant cyclophosphamide-based prophylaxis.

BACKGROUND AIMS: Post-transplant cyclophosphamide (PTCY)-based prophylaxis is becoming widespread for allogeneic hematopoietic cell transplantation (allo-HCT) performed independently of the selected donor source. In parallel, use of the Endothelial Activation and Stress Index (EASIX)-considered a surrogate parameter of endothelial activation-for predicting patient outcomes and clinical complications is gaining popularity in the allo-HCT setting. METHODS: We first investigated whether the dynamics of EASIX after allo-HCT differ between patients receiving PTCY and patients receiving other prophylaxis. We then investigated whether the predictive capacity of EASIX persists in PTCY-based allo-HCT. A total of 328 patients transplanted between 2014 and 2020 were included, and 201 (61.2%) received PTCY. RESULTS: EASIX trends differed significantly between the groups. Compared with patients receiving other prophylaxis, patients receiving PTCY had lower EASIX on day 0 and higher values between day 7 and day 100. In patients receiving PTCY, higher EASIX correlated significantly with higher non-relapse mortality (NRM) and lower overall survival (OS) when measured before and during the first 180 days after allo-HCT. In addition, higher EASIX scores measured at specific time points were predictors of veno-occlusive disease (VOD), transplant-associated thrombotic microangiopathy (TA-TMA) and grade 2-4 acute graft-versus-host disease (aGVHD) risk. CONCLUSIONS: This study demonstrates how EASIX trends vary during the first 180 days after allo-HCT in patients receiving PTCY and those not receiving PTCY and validates the utility of this index for predicting NRM, OS and risk of VOD, TA-TMA and grade 2-4 aGVHD in patients receiving PTCY.

The academic point-of-care anti-CD19 chimeric antigen receptor T-cell product varnimcabtagene autoleucel (ARI-0001 cells) shows efficacy and safety in the treatment of relapsed/refractory B-cell non-Hodgkin lymphoma.

Varnimcabtagene autoleucel (var-cel) is an academic anti-CD19 chimeric antigen receptor (CAR) product used for the treatment of non-Hodgkin lymphoma (NHL) in the CART19-BE-01 trial. Here we report updated outcomes of patients with NHL treated with var-cel. B-cell recovery was compared with patients with acute lymphoblastic leukaemia (ALL). Forty-five patients with NHL were treated. Cytokine release syndrome (any grade) occurred in 84% of patients (4% grade ≥3) and neurotoxicity in 7% (2% grade ≥3). The objective response rate was 73% at Day +100, and the 3-year duration of response was 56%. The 3-year progression-free and overall survival were 40% and 52% respectively. High lactate dehydrogenase was the only covariate with an impact on progression-free survival. The 3-year incidence of B-cell recovery was lower in patients with NHL compared to ALL (25% vs. 60%). In conclusion, in patients with NHL, the toxicity of var-cel was manageable, while B-cell recovery was significantly prolonged compared to ALL. This trial was registered as NCT03144583.

Post-Transplantation Cyclophosphamide and Tacrolimus for Graft-versus-Host Disease Prevention after Allogeneic Hematopoietic Cell Transplantation from HLA-Matched Donors Has More Advantages Than Limitations.

This study compared the efficacy of graft-versus-host disease (GVHD) prophylaxis with post-transplantation cyclophosphamide (PTCy) and tacrolimus (Tac) versus other regimens in 272 adults undergoing peripheral blood (PB) allogeneic hematopoietic cell transplantation (allo-HCT) from HLA-matched donors. Of these 272 patients, 95 (34.9%) received PTCy/Tac. The times to neutrophil and platelet engraftment were longer in the PTCy/Tac group (20 days versus 16 days for neutrophils and 19 days versus 12 days for platelets). The day +30 cumulative incidence (CuI) of bacterial bloodstream infection was higher in the PTCy/Tac group (43.2% versus 13.0%; P < .001). The CuIs of grade II-IV and grade III-IV acute GVHD (aGVHD) at day +180 were 14.7% and 4.2%, and the CuI of moderate/severe cGVHD at 2 years was 2.4% in the PTCy/Tac group and 41.8% (hazard ratio [HR], .29; P < .001), 15.8%, (HR, .24; P = .007), and 47.0% (HR, .05; P < .001), respectively, in the no-PTCy group. The duration of immunosuppression was shorter in patients receiving PTCy/Tac (6.2 months versus 9.0 months; P < .001). PTCy/Tac patients had higher OS (2 years: 74.3% versus 60.9%; HR, .54; P = .012), lower NRM (2 years: 8.6% versus 15.8%; HR, .54; P = .11), comparable CuI of relapse (2 years: 26.0% versus 24.4%; HR, 1.03; P = .89), and higher GRFS (2 years: 59.1% versus 16.7%; HR, .32; P < .001). Using PTCy/Tac in HLA-matched PB allo-HCT improved transplantation outcomes at out institution compared with previous prophylactic regimens, including a higher probability of survival despite more delayed engraftment and a higher rate of bacterial infection.

Infection epidemiology in relation to different therapy phases in patients with haematological malignancies receiving CAR T-cell therapy.

BACKGROUND: We described the real-life epidemiology and causes of infections on the different therapy phases in patients undergoing chimeric antigen receptor (CAR) T-cells directed towards CD19+ or BCMA+ cells. METHODS: All consecutive patients receiving CAR T-cell therapy at our institution were prospectively followed-up. We performed various comparative analyses of all patients and subgroups with and without infections. RESULTS: Ninety-one adults mainly received CAR T-cell therapy for acute leukaemia (53%) and lymphoma (33%). We documented a total of 77 infections in 47 (52%) patients, 37 (48%) during the initial neutropenic phase and 40 (52%) during the non-neutropenic phase. Infections during the neutropenic phase were mainly due to bacterial (29, 78%): catheter infections (11 [38%] cases), endogenous source (5 [17%]), and Clostridioides difficile (5 [17%]). Patients receiving corticosteroids after CAR T-cell therapy had a higher risk of endogenous infection (100% vs. 16%; p = .006). During the non-neutropenic phase, bacterial infections remained very frequent (24, 60%), mainly with catheter source (8, 33%). Respiratory tract infections were common (17, 43%). CONCLUSIONS: Infections after CAR T-cell therapy were frequent. During the neutropenic phase, it is essential to prevent nosocomial infections and balance the use of antibiotics to lower endogenous bacteraemia and Clostridial infection rates.

At-Home Foscarnet Administration in Patients with Cytomegalovirus Infection Post-Allogeneic Stem Cell Transplantation: A Unicentric, Safe, and Feasible Program.

Cytomegalovirus (CMV) infection is a relevant cause of morbimortality in patients receiving allogeneic stem cell transplantation (allo-HCT). Foscarnet (FCN) is an effective drug against CMV administered intravenously and usually on an inpatient basis. The Home Care Unit (HCU) for hematologic patients at our hospital designed an at-home FCN administration model to avoid the hospitalization of patients requiring FCN treatment. This study analyzes whether the at-home administration of FCN is as safe and effective as its hospital administration. We collected and compared demographic, clinical, analytical, and economic data of patients with CMV infection post-allo-HCT who received FCN in the hospital (n = 16, 17 episodes) vs. at-home (n = 67, 88 episodes). The proportions of patients with cured CMV infections were comparable between the two groups (65.9% vs. 76.5%, p = 0.395). The median duration of FCN treatment was 15 (interquartile range [IQR] 9-23) and 14 (IQR 11-19) days in the HCU and inpatient cohorts, respectively (p = 0.692). There were no significant differences in the FCN toxicities between groups except for hypocalcemia (26.1% vs. 58.8%, p = 0.007), which was more prevalent in the inpatient cohort. A significant cost-effectiveness was found in the HCU cohort, with a median savings per episode of EUR 5270. It may be concluded that home administration of FCN is a safe, effective, and cost-efficient therapeutic option for patients with CMV infection and disease.

Results of a phase 1/2 study of cemdisiran in healthy subjects and patients with paroxysmal nocturnal hemoglobinuria.

Complement dysregulation underpins the physiopathology of paroxysmal nocturnal hemoglobinuria (PNH). Cemdisiran, an RNA interference investigational treatment, silences complement component 5 (C5) expression in the liver. Previously reported results showed sustained reduction in C5 levels following cemdisiran monotherapy, with >90% reduction in patients with PNH. This phase 1/2 study evaluated single (Part A, n = 32; 50-900 mg) or multiple (Part B, n = 24; 100-600 mg) ascending doses of cemdisiran or placebo (double-blind, randomized 3:1) in healthy adults, or cemdisiran in patients with PNH who were naive to, or receiving, eculizumab (Part C, n = 6; 200 or 400 mg weekly; open-label). The primary objective was to assess the safety and tolerability of cemdisiran. Other assessments included change in complement activity, lactate dehydrogenase levels, and inhibition of hemolysis following cemdisiran treatment. Cemdisiran was generally well tolerated in this study. Overall, 75%, 89%, and 100% of subjects in Parts A, B, and C, respectively, experienced ≥1 non-serious adverse event (AE). Most events were Grade 1 or 2 in severity and the most common AEs included nasopharyngitis and headache. Cemdisiran elicited robust, sustained reductions in the complement activity in healthy adults and patients with PNH. In Part C, exploratory analyses showed that cemdisiran monotherapy was insufficient to prevent hemolysis in patients with PNH as measured by serum lactate dehydrogenase levels. Cemdisiran and eculizumab combination therapy reduced the dose of eculizumab required to provide adequate control of intravascular hemolysis. These results demonstrate a potential benefit of cemdisiran coadministration in patients who are inadequate responders to eculizumab alone.

Fractionated initial infusion and booster dose of ARI0002h, a humanised, BCMA-directed CAR T-cell therapy, for patients with relapsed or refractory multiple myeloma (CARTBCMA-HCB-01): a single-arm, multicentre, academic pilot study.

BACKGROUND: Chimeric antigen receptor (CAR) T-cell therapy is a promising option for patients with heavily treated multiple myeloma. Point-of-care manufacturing can increase the availability of these treatments worldwide. We aimed to assess the safety and activity of ARI0002h, a BCMA-targeted CAR T-cell therapy developed by academia, in patients with relapsed or refractory multiple myeloma. METHODS: CARTBCMA-HCB-01 is a single-arm, multicentre study done in five academic centres in Spain. Eligible patients had relapsed or refractory multiple myeloma and were aged 18-75 years; with an Eastern Cooperative Oncology Group performance status of 0-2; two or more previous lines of therapy including a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 antibody; refractoriness to the last line of therapy; and measurable disease according to the International Myeloma Working Group criteria. Patients received an initial fractionated infusion of 3 × 106 CAR T cells per kg bodyweight in three aliquots (0·3, 0·9, and 1·8 × 106 CAR-positive cells per kg intravenously on days 0, 3, and 7) and a non-fractionated booster dose of up to 3 × 106 CAR T cells per kg bodyweight, at least 100 days after the first infusion. The primary endpoints were overall response rate 100 days after first infusion and the proportion of patients developing cytokine-release syndrome or neurotoxic events in the first 30 days after receiving treatment. Here, we present an interim analysis of the ongoing trial; enrolment has ended. This study is registered with ClinicalTrials.gov, NCT04309981, and EudraCT, 2019-001472-11. FINDINGS: Between June 2, 2020, and Feb 24, 2021, 44 patients were assessed for eligibility, of whom 35 (80%) were enrolled. 30 (86%) of 35 patients received ARI0002h (median age 61 years [IQR 53-65], 12 [40%] were female, and 18 [60%] were male). At the planned interim analysis (cutoff date Oct 20, 2021), with a median follow-up of 12·1 months (IQR 9·1-13·5), overall response during the first 100 days from infusion was 100%, including 24 (80%) of 30 patients with a very good partial response or better (15 [50%] with complete response, nine [30%] with very good partial response, and six [20%] with partial response). Cytokine-release syndrome was observed in 24 (80%) of 30 patients (all grade 1-2). No cases of neurotoxic events were observed. Persistent grade 3-4 cytopenias were observed in 20 (67%) patients. Infections were reported in 20 (67%) patients. Three patients died: one because of progression, one because of a head injury, and one due to COVID-19. INTERPRETATION: ARI0002h administered in a fractioned manner with a booster dose after 3 months can provide deep and sustained responses in patients with relapsed or refractory multiple myeloma, with a low toxicity, especially in terms of neurological events, and with the possibility of a point-of-care approach. FUNDING: Instituto de Salud Carlos III (co-funded by the EU), Fundación La Caixa, and Fundació Bosch i Aymerich.

Characterization of the endotheliopathy, innate-immune activation and hemostatic imbalance underlying CAR-T cell toxicities: laboratory tools for an early and differential diagnosis.

BACKGROUND: Chimeric antigen receptor (CAR)-T cell-based immunotherapy constitutes a revolutionary advance for treatment of relapsed/refractory hematological malignancies. Nevertheless, cytokine release and immune effector cell-associated neurotoxicity syndromes are life-threatening toxicities in which the endothelium could be a pathophysiological substrate. Furthermore, differential diagnosis from sepsis, highly incident in these patients, is challenging. Suitable laboratory tools could be determinant for their appropriate management. METHODS: Sixty-two patients treated with CAR-T cell immunotherapy for hematological malignancies (n=46 with CD19-positive diseases, n=16 with multiple myeloma) were included. Plasma samples were obtained: before CAR-T cell infusion (baseline); after 24-48 hours; at suspicion of any toxicity onset and 24-48 hours after immunomodulatory treatment. Biomarkers of endothelial dysfunction (soluble vascular cell adhesion molecule 1 (sVCAM-1), soluble TNF receptor 1 (sTNFRI), thrombomodulin (TM), soluble suppression of tumorigenesis-2 factor (ST2), angiopoietin-2 (Ang-2)), innate immunity activation (neutrophil extracellular traps (NETs), soluble C5b-9 (sC5b-9)) and hemostasis/fibrinolysis (von Willebrand Factor antigen (VWF:Ag), ADAMTS-13 (A13), α2-antiplasmin (α2-AP), plasminogen activator inhibitor-1 antigen (PAI-1 Ag)) were measured and compared with those in cohorts of patients with sepsis and healthy donors. RESULTS: Patients who developed CAR-T cell toxicities presented increased levels of sVCAM-1, sTNFRI and ST2 at the clinical onset versus postinfusion values. Twenty-four hours after infusion, ST2 levels were good predictors of any CAR-T cell toxicity, and combination of ST2, Ang-2 and NETs differentiated patients requiring intensive care unit admission from those with milder clinical presentations. Association of Ang-2, NETs, sC5b-9, VWF:Ag and PAI-1 Ag showed excellent discrimination between severe CAR-T cell toxicities and sepsis. CONCLUSIONS: This study provides relevant contributions to the current knowledge of the CAR-T cell toxicities pathophysiology. Markers of endotheliopathy, innate immunity activation and hemostatic imbalance appear as potential laboratory tools for their prediction, severity and differential diagnosis.

Effect of CD34+ Cell Dose on the Outcomes of Allogeneic Stem Cell Transplantation with Post-Transplantation Cyclophosphamide.

The impact of infused CD34+ cell dose on outcomes after allogeneic hematopoietic stem cell transplantation (alloHSCT) using standard graft-versus-host disease (GVHD) prophylaxis remains controversial. Information on this subject is scarce for alloHSCT using high-dose post-transplantation cyclophosphamide (PTCy). We aimed to assess the effect of CD34+ cell dose in peripheral blood stem cell (PBSC) grafts on the outcome of alloHSCT using PTCy-based GVHD prophylaxis. To do so, we conducted a single-center retrospective analysis of 221 consecutive adult patients who underwent PTCy alloHSCT from HLA-matched sibling donors (MSDs; n = 22), HLA-matched unrelated donors (MUDs; n = 83), mismatched unrelated donors (MMUDs; n = 73), and haploidentical donors (n = 43). Based on the binary partitioning method, 5 × 106/kg was used as the optimal cutoff for CD34+ cell dose. According to our institutional protocol, the maximum CD34+ cell dose was capped at 8 × 106/kg. The study cohort was divided into 2 groups based on CD34+ cell dose: high dose (>5 to 8 × 106/kg) and low dose (≤5 × 106/kg). Patients receiving high-dose CD34+-containing grafts had significantly shorter median times to neutrophil engraftment and platelet engraftment compared to those who received low-dose CD34+ (19 days versus 21 days [P = .002] and 16 days versus 22 days [P = .04], respectively). There were no differences between the high-dose and low-dose groups in the cumulative incidence of day +100 acute GVHD (grade II-IV: 25% versus 23% [P = .7]; grade III-IV: 5% versus 4% [P = .4], respectively) or 2-year chronic GVHD (moderate/severe GVHD: 9% versus 6%; P = .5). There was no impact of CD34+ cell dose on survival outcomes with the use of MSDs, MUDs, or MMUDs. Recipients of haploidentical alloHSCT using low-dose CD34+ cells had significantly worse overall survival (hazard ratio [HR], 6.01; P = .004) and relapse-free survival (HR, 4.57; P = .004). In recipients of PBSC PTCy alloHSCT, infused CD34+ cell doses >5 to 8 × 106/kg were associated with faster neutrophil and platelet engraftment, independent of donor type. Our study suggests an impact of CD34+ cell dose on survival outcomes only with haploidentical donors, for whom the administration of a CD34+ cell dose ≤5 × 106/kg significantly decreased survival outcomes.

Incidence, risk factors, and impact of early cardiac toxicity after allogeneic hematopoietic cell transplant.

This study investigates early cardiac events (ECEs) occurring during the first 180 days after allogeneic hematopoietic cell transplant (allo-HCT) in 416 adults receiving posttransplant cyclophosphamide (PTCY) (n = 258) or not receiving PTCY (n = 158). Total body irradiation (TBI) was given to 133 (31.9%) patients, of whom 111 (83.4%) received TBI combined with PTCY. The day +180 cumulative incidence function (CIF) of ECEs was 8.4%, with heart failure (n = 13) and pericardial complications (n = 11) being the most prevalent complications. The incidence of ECEs was higher in patients receiving PTCY, and receiving TBI. ECEs were more prevalent in haploidentical HCTs than in matched sibling donor, 10/10 HLA-matched unrelated donor, and 9/10 HLA-mismatched unrelated donor allo-HCTs. As for the ECE risk from the combination of PTCY and TBI, the multivariate analysis reported that patients receiving PTCY without TBI, TBI without PTCY, and TBI with PTCY were at higher risk for ECEs compared with patients receiving neither PTCY nor TBI. Pre-existing cardiac morbidity predicted ECEs. However, using high-dose CY-containing preparative regimens did not increase the risk for cardiac toxicity at +180 days after allo-HCT. ECEs were associated with higher nonrelapse mortality and lower overall survival. Considering that PTCY and TBI were predictors for ECEs, and the impact of this complication on transplant mortality, the implementation of cardiac monitoring plans could be appropriate in patients receiving these medications.

Evaluation of cell collection efficiency in non-mobilized adult donors for autologous chimeric antigen receptor T-cell manufacturing.

BACKGROUND AND OBJECTIVES: Data about collection efficiency 1 (CE1), which takes into account blood cell counts before and after collection, thus providing a more accurate estimate, in the collection of autologous T lymphocytes by apheresis for chimeric antigen receptor (CAR) T-cells remain scarce. We evaluated donor- and procedure-related characteristics that might influence the CE1 of lymphocytes. MATERIALS AND METHODS: We retrospectively reviewed all mononuclear cell (MNC) collections) performed for CAR T-cell manufacturing in our institution from May 2017 to June 2021 in adult patients. Age, gender, weight, total blood volume (TBV), prior haematopoietic cell transplant, diagnosis, days between last treatment and apheresis, pre-collection cell counts, duration of apheresis, TBV processed, vascular access, inlet flow and device type were analysed as potential factors affecting CE1 of lymphocytes. RESULTS: A total of 127 autologous MNC collections were performed on 118 patients diagnosed with acute lymphoblastic leukaemia (n = 53, 45%), non-Hodgkin lymphoma (n = 40, 34%), multiple myeloma (n = 19, 16%), and chronic lymphocytic leukaemia (n = 6, 5%). The median CE1 of lymphocytes was 47% (interquartile range: 32%-65%). In multiple regression analysis, Amicus device was associated with higher CE1 of lymphocytes (p = 0.01) and lower CE1 of platelets (p < 0.01) when compared with Optia device. CONCLUSION: The knowledge of the MNC and lymphocyte CE1 of each apheresis device used to collect cells for CAR T therapy, together with the goal of the number of cells required, is essential to define the volume to be processed and to ensure the success of the collection.

Impact of Early Intrapatient Variability of Tacrolimus Concentrations on the Risk of Graft-Versus-Host Disease after Allogeneic Stem Cell Transplantation Using High-Dose Post-Transplant Cyclophosphamide.

Tacrolimus (Tac) is a pivotal immunosuppressant agent used to prevent graft-versus-host disease (GVHD) after allogeneic stem cell transplantation (alloHSCT). Tac is characterized by a narrow therapeutic window and a high inter-patient and intra-patient pharmacokinetic variability (IPV). Although high IPV of Tac concentrations has been associated with adverse post-transplant outcomes following solid organ transplantation, the effects of Tac IPV on alloHSCT recipients have not been determined. Tac IPV was therefore retrospectively evaluated in 128 alloHSCT recipients receiving high-dose post-transplant cyclophosphamide (PTCy) and the effects of Tac IPV on the occurrence of acute GVHD (aGVHD) were analyzed. Tac IPV was calculated from pre-dose concentrations (C0) measured during the first month after Tac initiation. The cumulative rates of grades II-IV and grades III-IV aGVHD at day +100 were 22.7% and 7%, respectively. Higher Tac IPV was associated with a greater risk of developing GVHD, with patients having IPV > 50th percentile having significantly higher rates of grades II-IV (34.9% vs. 10.8%; hazard ratio [HR] 3.858, p < 0.001) and grades III-IV (12.7% vs. 1.5%; HR 9.69, p = 0.033) aGVHD than patients having IPV ≤ 50th percentile. Similarly, patients with IPV > 75th percentile had higher rates of grades II-IV (41.9% vs. 16.5%; HR 3.30, p < 0.001) and grades III-IV (16.1% vs. 4.1%; HR 4.99, p = 0.012) aGVHD than patients with IPV ≤ 75th percentile. Multivariate analyses showed that high Tac IPV (>50th percentile) was an independent risk factor for grades II-IV (HR 2.99, p = 0.018) and grades III-IV (HR 9.12, p = 0.047) aGVHD. Determination of Tac IPV soon after alloHSCT could be useful in identifying patients at greater risk of aGVHD.

CAR density influences antitumoral efficacy of BCMA CAR T cells and correlates with clinical outcome.

Identification of new markers associated with long-term efficacy in patients treated with CAR T cells is a current medical need, particularly in diseases such as multiple myeloma. In this study, we address the impact of CAR density on the functionality of BCMA CAR T cells. Functional and transcriptional studies demonstrate that CAR T cells with high expression of the CAR construct show an increased tonic signaling with up-regulation of exhaustion markers and increased in vitro cytotoxicity but a decrease in in vivo BM infiltration. Characterization of gene regulatory networks using scRNA-seq identified regulons associated to activation and exhaustion up-regulated in CARHigh T cells, providing mechanistic insights behind differential functionality of these cells. Last, we demonstrate that patients treated with CAR T cell products enriched in CARHigh T cells show a significantly worse clinical response in several hematological malignancies. In summary, our work demonstrates that CAR density plays an important role in CAR T activity with notable impact on clinical response.

Bacterial Bloodstream Infections in Patients Undergoing Allogeneic Hematopoietic Cell Transplantation With Post-Transplantation Cyclophosphamide.

This study investigates the incidence and predictors for bacterial bloodstream infection (BSI) in 330 adults undergoing allo-HCT, and explores the effect of post-transplantation cyclophosphamide (PTCY) on the probability of presenting this complication. All patients received levofloxacin during the aplastic phase. Only the first episode of BSI was counted as an event. Patients were classified into 2 groups: PTCY-based (n = 200) versus other prophylaxis (n = 130). One hundred twenty-four patients were diagnosed with a first episode of BSI, most of them during the first 30 days (70.2%). Proportions of BSIs caused by Gram-positive bacteria were comparable to those caused by Gram-negative bacteria (48.3% versus 45.9%). The cumulative incidence of BSI was higher in patients receiving PTCY than in those receiving other prophylaxis (days 30 and 100: 35.0% and 37.0% versus 13.1% and 18.5%, P < .001). At day 30, the likelihood of BSI was 2.41 (P = .012) times higher in the PTCY group than in the non-PTCY group. The 30-day mortality rate in all patients with BSI was 8.0%, lower (P = .002) in the PTCY group (2.3%) than in the non-PTCY group (21.6%). Finally, the overall survival of patients receiving PTCY and diagnosed with BSI was similar to that of patients without presenting this complication. © 2023 American Society for Blood and Marrow Transplantation. Published by Elsevier Inc. All rights reserved.

T cells isolated from G-CSF-treated multiple myeloma patients are suitable for the generation of BCMA-directed CAR-T cells.

Autologous cell immunotherapy using B cell maturation antigen (BCMA)-targeted chimeric antigen receptor (CAR)-T cells is an effective novel treatment for multiple myeloma (MM). This therapy has only been used for relapsed and refractory patients, at which stage the endogenous T cells used to produce the CAR-T cells are affected by the immunosuppressive nature of advanced MM and/or side effects of previous therapies. An alternative pool of "fitter" T cells is found in leukocytoapheresis products that are routinely collected to obtain hematopoietic progenitor cells for autologous stem cell transplantation (ASCT) early in the treatment of MM. However, to mobilize the progenitor cells, patients are dosed with granulocyte colony-stimulating factor (G-CSF), which is reported to adversely affect T cell proliferation, function, and differentiation. Here, we aimed to first establish whether G-CSF treatment negatively influences T cell phenotype and to ascertain whether previous exposure of T cells to G-CSF is deleterious for anti-BCMA CAR-T cells. We observed that G-CSF had a minimal impact on T cell phenotype when added in vitro or administered to patients. Moreover, we found that CAR-T cell fitness and anti-tumor activity were unaffected when generated from G-CSF-exposed T cells. Overall, we showed that ASCT apheresis products are a suitable source of T cells for anti-BCMA CAR-T cell manufacture.