Antibiotic-induced loss of gut microbiome metabolic output correlates with clinical responses to CAR T-cell therapy.

Antibiotic-induced microbiome dysbiosis is widespread in oncology, adversely affecting outcomes and side effects of various cancer treatments, including immune checkpoint inhibitors and chimeric antigen receptor T (CAR-T) cell therapies. In this study, we observed that prior exposure to broad-spectrum ABX with extended anaerobic coverage like piperacillin-tazobactam and meropenem was associated with worsened anti-CD19 CAR-T therapy survival outcomes in large B-cell lymphoma patients (n=422), compared to other ABX classes. In a discovery subset of these patients (n=67), we found that the use of these ABX was in turn associated with substantial dysbiosis of gut microbiome function, resulting in significant alterations of the gut and blood metabolome, including microbial effectors such as short-chain fatty acids (SCFAs) and other anionic metabolites, findings that were largely reproduced in an external validation cohort (n=58). Broader evaluation of circulating microbial metabolites revealed reductions in indole and cresol derivatives, as well as trimethylamine N-oxide, in patients who received ABX treatment (discovery n=40, validation n=28). These findings were recapitulated in an immune-competent CAR-T mouse model, where meropenem-induced dysbiosis led to a systemic dysmetabolome and decreased murine anti-CD19 CAR-T efficacy. Furthermore, we demonstrate that SCFAs can enhance the metabolic fitness of CAR-T cells, leading to improved tumor killing capacity. Together, these results suggest that broad-spectrum ABX deplete metabolically active commensals whose metabolites are essential for enhancing CAR-T efficacy, shedding light on the intricate relationship between ABX exposure, microbiome function and their impact on CAR-T cell efficacy. This highlights the potential for modulating the microbiome to augment CAR-T immunotherapy.

Comprehensive antitumor immune response boosted by dual inhibition of SUMOylation and MEK in MYC-expressing KRAS-mutant cancers.

Precision medicine has drastically changed cancer treatment strategies including KRAS-mutant cancers which have been undruggable for decades. While intrinsic or acquired treatment resistance remains unresolved in many cases, epigenome-targeted therapy may be an option to overcome. We recently discovered the effectiveness of blocking small ubiquitin-like modifier (SUMO) signaling cascade (SUMOylation) in MYC-expressing KRAS-mutant cancer cells using a SUMO-activating enzyme E inhibitor TAK-981 that results in SUMOylation inhibition. Interestingly, TAK-981 promoted the degradation of MYC via the ubiquitin-proteasome system. Moreover, combination therapy with TAK-981 and MEK inhibitor trametinib remarkably regressed xenografted KRAS-mutant tumors by accumulating DNA damage and inducing apoptosis. Whereas our recent study revealed immune-independent antitumor efficacy, we evaluated the immune responses of cancer cells and immune cells in this study. We found that TAK-981-induced MYC downregulation promoted the activation of STING followed by Stat1 and MHC class I in KRAS-mutant cancer cells. Activation of dendritic cells or T cells treated with TAK-981 was also verified by upregulated activation markers in dendritic cells or skew-toward effector-like phenotypes in T cells. Furthermore, the enhanced immune-dependent antitumor efficacy of the combination therapy with TAK-981 and trametinib was confirmed by infiltration of immune cells into tumor tissues and immunodepleting-test using immunodepleting antibodies in syngeneic immunocompetent mouse models. Together with our recent study and here, the findings support that combination inhibition of SUMOylation and MEK comprehensively conquers MYC-expressing KRAS-mutant cancers by both immune-dependent and immune-independent antitumor responses.

Dual inhibition of SUMOylation and MEK conquers MYC-expressing KRAS-mutant cancers by accumulating DNA damage.

BACKGROUND: KRAS mutations frequently occur in cancers, particularly pancreatic ductal adenocarcinoma, colorectal cancer, and non-small cell lung cancer. Although KRASG12C inhibitors have recently been approved, effective precision therapies have not yet been established for all KRAS-mutant cancers. Many treatments for KRAS-mutant cancers, including epigenome-targeted drugs, are currently under investigation. Small ubiquitin-like modifier (SUMO) proteins are a family of small proteins covalently attached to and detached from other proteins in cells via the processes called SUMOylation and de-SUMOylation. We assessed whether SUMOylation inhibition was effective in KRAS-mutant cancer cells. METHODS: The efficacy of the first-in-class SUMO-activating enzyme E inhibitor TAK-981 (subasumstat) was assessed in multiple human and mouse KRAS-mutated cancer cell lines. A gene expression assay using a TaqMan array was used to identify biomarkers of TAK-981 efficacy. The biological roles of SUMOylation inhibition and subsequent regulatory mechanisms were investigated using immunoblot analysis, immunofluorescence assays, and mouse models. RESULTS: We discovered that TAK-981 downregulated the expression of the currently undruggable MYC and effectively suppressed the growth of MYC-expressing KRAS-mutant cancers across different tissue types. Moreover, TAK-981-resistant cells were sensitized to SUMOylation inhibition via MYC-overexpression. TAK-981 induced proteasomal degradation of MYC by altering the balance between SUMOylation and ubiquitination and promoting the binding of MYC and Fbxw7, a key factor in the ubiquitin-proteasome system. The efficacy of TAK-981 monotherapy in immunocompetent and immunodeficient mouse models using a mouse-derived CMT167 cell line was significant but modest. Since MAPK inhibition of the KRAS downstream pathway is crucial in KRAS-mutant cancer, we expected that co-inhibition of SUMOylation and MEK might be a good option. Surprisingly, combination treatment with TAK-981 and trametinib dramatically induced apoptosis in multiple cell lines and gene-engineered mouse-derived organoids. Moreover, combination therapy resulted in long-term tumor regression in mouse models using cell lines of different tissue types. Finally, we revealed that combination therapy complementally inhibited Rad51 and BRCA1 and accumulated DNA damage. CONCLUSIONS: We found that MYC downregulation occurred via SUMOylation inhibition in KRAS-mutant cancer cells. Our findings indicate that dual inhibition of SUMOylation and MEK may be a promising treatment for MYC-expressing KRAS-mutant cancers by enhancing DNA damage accumulation.

JAK2/mTOR Inhibition Fails to Prevent Acute GVHD Despite Reduced Th1/Th17 cells: Final Phase II Trial Results.

Our phase I graft-versus-host disease (GVHD) prevention trial of JAK2 inhibitor, pacritinib, (recommended phase II dose: 100mg po BID day 0 to +70) plus sirolimus and tacrolimus (PAC/SIR/TAC) demonstrated the regimen was safe and free of pan-JAK myelosuppression after allogeneic hematopoietic cell transplantation (alloHCT). PAC inhibits IL-6 receptor activity and pathogenic Th1/Th17 differentiation in preclinical models and the phase I trial. Herein we report on our completed phase II trial of PAC/SIR/TAC after 8/8-HLA matched alloHCT. This single-arm phase II trial (NCT02891603) was powered to determine if PAC/SIR/TAC suppressed %pSTAT3+ CD4+ T cells at day +21 (primary endpoint: %pSTAT3+ CD4+ T cells ≤ 35%) and estimated grade II-IV acute GVHD by day +100. The impact of PAC/SIR/TAC on T cell subsets, CD28 (pS6 and pH3ser10), and IL-2 receptor (pSTAT5) signal transduction was also evaluated. Eligible patients (n=28) received alloHCT for hematologic malignancies or myeloproliferative neoplasms. Reduced or myeloablative intensity conditioning was permitted. PAC/SIR/TAC met the primary endpoint, reducing %pSTAT3+ CD4+ T cells to 9.62% at day +21. Th1/Th17 cells were decreased at day +21, increasing the ratio of Tregs to Th1 and Th17 cells with PAC/SIR/TAC at RP2D PAC compared to dose level 1 PAC. The cumulative incidence of grade II-IV acute GVHD by day +100 with PAC/SIR/TAC was similar to historic SIR/TAC values (46 v 43%). While PAC/SIR/TAC suppressed pSTAT3 and Th1/Th17 cells, the regimen did not improve acute GVHD prevention.

Biosimilars and access to biologic therapy in immune-mediated diseases.

BACKGROUND: The rise of biologic agents has been a major breakthrough in treating immune-mediated inflammatory diseases (IMIDs). However, their high cost underscores the need for strategies to optimize treatment efficiency. Biosimilars offer cost-effective alternatives to biologics. This study aimed to assess biosimilar drug availability's impact on biologic therapy access for IMIDs. RESEARCH DESIGN AND METHODS: A retrospective observational study in 15 Spanish hospitals analyzed IMID patients (arthropathies, inflammatory bowel disease and psoriasis) initiating biologic therapy with originator or biosimilar drugs (infliximab, etanercept, adalimumab). Time to availability and initiation of biologic therapy were assessed. RESULTS: 267 patients were included, with 58.4% starting on biosimilars. The mean time to availability of the biologic drugs in the hospitals was 15.9 ± 6.7 months, (20.0 ± 12.4 for originator and 11.8 ± 5.2 for biosimilars). Mean time to biologic treatment was 7.7 ± 9.0 years (8.6 ± 8.9 for originators and 7.0 ± 9.0 for biosimilars). Showing statistically significant differences among conditions. CONCLUSION: The emergence of biosimilar drugs has enhanced market competition and accelerated their adoption into hospitals' therapeutic regimens over original reference drugs. This has significantly improved access to biologic therapy for patients with IMIDs, evidenced by a notable 1.6-year reduction in access time for biosimilar drugs.

Baseline Serum Inflammatory Proteins Predict Poor CAR T Outcomes in Diffuse Large B-cell Lymphoma.

A subset of patients with diffuse large B-cell lymphoma (DLBCL) treated with CD19 chimeric antigen receptor (CAR) T-cell therapy have poor clinical outcomes. We report serum proteins associated with severe immune-mediated toxicities and inferior clinical responses in 146 patients with DLBCL treated with axicabtagene ciloleucel. We develop a simple stratification based on pre-lymphodepletion C reactive protein (CRP) and ferritin to classify patients into low-, intermediate-, and high-risk groups. We observe that patients in the high-risk category were more likely to develop grade ≥3 toxicities and had inferior overall and progression-free survival. We sought to validate our findings with two independent international cohorts demonstrating that patients classified as low-risk have excellent efficacy and safety outcomes. Based on routine and readily available laboratory tests that can be obtained prior to lymphodepleting chemotherapy, this simple risk stratification can inform patient selection for CAR T-cell therapy. SIGNIFICANCE: CAR T-cell therapy has changed the treatment paradigm for patients with relapsed/refractory hematologic malignancies. Despite encouraging efficacy, a subset of patients have poor clinical outcomes. We show that a simple clinically applicable model using pre-lymphodepletion CRP and ferritin can identify patients at high risk of poor outcomes. This article is featured in Selected Articles from This Issue, p. 80.

Agreement between the perception of colon cleansing reported by patients and colon cleansing assessed by a validated colon cleansing scale.

AIMS: Patients' perception of their cleansing quality can guide strategies to improve cleansing during colonoscopy. There are no studies assessing the agreement between the quality of cleansing perceived by patients and cleansing quality assessed during colonoscopy using validated bowel preparation scales. The main aim of this study was to compare the cleansing quality reported by patients with the quality during colonoscopy using the Boston Bowel Preparation Scale (BBPS). PATIENTS AND METHODS: Consecutive patients referred to an outpatient colonoscopy were included. Four drawings representing different degrees of cleansing were designed. Patients chose the drawing that most resembled the last stool. The predictive ability of the patient's perception and agreement between the patient's perception and the BBPS were calculated. A BBPS score of <2 points in any segment was considered inadequate. RESULTS: Six hundred and thirty-three patients were included (age: 62.8±13.7 years, male: 53.4%). Overall, 107 patients (16.9%) had inadequate cleansing during colonoscopy, and in 12.2% of cases, the patient's perception was poor. The patient's perception compared to the quality of cleanliness during colonoscopy presented a positive and negative predictive value of 54.6% and 88.3%, respectively. The agreement between patient perception and the BBPS was significant (P<0.001), although fair (k=0.37). The results were similar in a validation cohort of 378 patients (k=0.41). CONCLUSIONS: The cleanliness perceived by the patient and the quality of cleanliness using a validated scale were correlated, although fair. However, this measure satisfactorily identified patients with adequate preparation. Cleansing rescue strategies may target patients who self-report improper cleaning. Registration number of the trial: NCT03830489.

Design and validation of an artificial intelligence system to detect the quality of colon cleansing before colonoscopy.

BACKGROUND AND AIMS: Patients' perception of their bowel cleansing quality may guide rescue cleansing strategies before colonoscopy. The main aim of this study was to train and validate a convolutional neural network (CNN) for classifying rectal effluent during bowel preparation intake as "adequate" or "inadequate" cleansing before colonoscopy. PATIENTS AND METHODS: Patients referred for outpatient colonoscopy were asked to provide images of their rectal effluent during the bowel preparation process. The images were categorized as adequate or inadequate cleansing based on a predefined 4-picture quality scale. A total of 1203 images were collected from 660 patients. The initial dataset (799 images), was split into a training set (80%) and a validation set (20%). The second dataset (404 images) was used to develop a second test of the CNN accuracy. Afterward, CNN prediction was prospectively compared with the Boston Bowel Preparation Scale (BBPS) in 200 additional patients who provided a picture of their last rectal effluent. RESULTS: On the initial dataset, a global accuracy of 97.49%, a sensitivity of 98.17% and a specificity of 96.66% were obtained using the CNN model. On the second dataset, an accuracy of 95%, a sensitivity of 99.60% and a specificity of 87.41% were obtained. The results from the CNN model were significantly associated with those from the BBPS (P<0.001), and 77.78% of the patients with poor bowel preparation were correctly classified. CONCLUSION: The designed CNN is capable of classifying "adequate cleansing" and "inadequate cleansing" images with high accuracy.

Bispecific CD33/CD123 targeted chimeric antigen receptor T cells for the treatment of acute myeloid leukemia.

CD33 and CD123 are expressed on the surface of human acute myeloid leukemia blasts and other noncancerous tissues such as hematopoietic stem cells. On-target off-tumor toxicities may limit chimeric antigen receptor T cell therapies that target both CD33 and CD123. To overcome this limitation, we developed bispecific human CD33/CD123 chimeric antigen receptor (CAR) T cells with an "AND" logic gate. We produced novel CD33 and CD123 scFvs from monoclonal antibodies that bound CD33 and CD123 and activated T cells. Screening of CD33 and CD123 CAR T cells for cytotoxicity, cytokine production, and proliferation was performed, and we selected scFvs for CD33/CD123 bispecific CARs. The bispecific CARs split 4-1BB co-stimulation on one scFv and CD3ζ on the other. In vitro testing of cytokine secretion and cytotoxicity resulted in selecting bispecific CAR 1 construct for in vivo analysis. The CD33/CD123 bispecific CAR T cells were able to control acute myeloid leukemia (AML) in a xenograft AML mouse model similar to monospecific CD33 and CD123 CAR T cells while showing no on-target off-tumor effects. Based on our findings, human CD33/CD123 bispecific CAR T cells are a promising cell-based approach to prevent AML and support clinical investigation.

Severe hematotoxicity after CD19 CAR-T therapy is associated with suppressive immune dysregulation and limited CAR-T expansion.

Prolonged cytopenias after chimeric antigen receptor (CAR) T cell therapy are a significant clinical problem and the underlying pathophysiology remains poorly understood. Here, we investigated how (CAR) T cell expansion dynamics and serum proteomics affect neutrophil recovery phenotypes after CD19-directed CAR T cell therapy. Survival favored patients with "intermittent" neutrophil recovery (e.g., recurrent neutrophil dips) compared to either "quick" or "aplastic" recovery. While intermittent patients displayed increased CAR T cell expansion, aplastic patients exhibited an unfavorable relationship between expansion and tumor burden. Proteomics of patient serum collected at baseline and in the first month after CAR-T therapy revealed higher markers of endothelial dysfunction, inflammatory cytokines, macrophage activation, and T cell suppression in the aplastic phenotype group. Prolonged neutrophil aplasia thus occurs in patients with systemic immune dysregulation at baseline with subsequently impaired CAR-T expansion and myeloid-related inflammatory changes. The association between neutrophil recovery and survival outcomes highlights critical interactions between host hematopoiesis and the immune state stimulated by CAR-T infusion.

Cardiac and inflammatory biomarker differences in adverse cardiac events after chimeric antigen receptor T-Cell therapy: an exploratory study.

BACKGROUND: Chimeric antigen receptor T- Cell (CAR-T) immunotherapy has been a breakthrough treatment for various hematological malignancies. However, cardiotoxicities such as new-onset heart failure, arrhythmia, acute coronary syndrome and cardiovascular death occur in 10-15% of patients treated with CAR-T. This study aims to investigate the changes in cardiac and inflammatory biomarkers in CAR-T therapy to determine the role of pro-inflammatory cytokines. METHODS: In this observational study, ninety consecutive patients treated with CAR-T underwent baseline cardiac investigation with electrocardiogram (ECG), transthoracic echocardiogram (TTE), troponin-I, and B-type natriuretic peptide (BNP). Follow-up ECG, troponin-I and BNP were obtained five days post- CAR-T. In a subset of patients (N = 53), serum inflammatory cytokines interleukin (IL)-2, IL-6, IL-15, interferon (IFN)-γ, tumor necrosis factor (TNF)-α, granulocyte-macrophage colony-stimulating factor (GM-CSF), and angiopoietin 1 & 2 were tested serially, including baseline and daily during hospitalization. Adverse cardiac events were defined as new-onset cardiomyopathy/heart failure, acute coronary syndrome, arrhythmia and cardiovascular death. RESULTS: Eleven patients (12%) had adverse cardiac events (one with new-onset cardiomyopathy and ten with new-onset atrial fibrillation). Adverse cardiac events appear to have occurred among patients with advanced age (77 vs. 66 years; p = 0.002), higher baseline creatinine (0.9 vs. 0.7 mg/dL; 0.007) and higher left atrial volume index (23.9 vs. 16.9mL/m2; p = 0.042). Day 5 BNP levels (125 vs. 63pg/mL; p = 0.019), but not troponin-I, were higher in patients with adverse cardiac events, compared to those without. The maximum levels of IL-6 (3855.0 vs. 254.0 pg/mL; p = 0.021), IFN-γ (474.0 vs. 48.8pg/mL; p = 0.006) and IL-15 (70.2 vs. 39.2pg/mL; p = 0.026) were also higher in the adverse cardiac events group. However, cardiac and inflammatory biomarker levels were not associated with cardiac events. Patients who developed cardiac events did not exhibit worse survival compared to patients without cardiac events (Log-rank p = 0.200). CONCLUSION: Adverse cardiac events, predominantly atrial fibrillation, occur commonly after CAR-T (12%). The changes in serial inflammatory cytokine after CAR-T in the setting of adverse cardiac events suggests pro-inflammation as a pathophysiology and require further investigation for their role in adverse cardiac events. TWEET BRIEF HANDLE: CAR-T related Cardiotoxicity has elevated cardiac and inflammatory biomarkers. #CARTCell #CardioOnc #CardioImmunology.

Outcomes of CD19-Directed Chimeric Antigen Receptor T Cell Therapy for Transformed Nonfollicular Lymphoma.

CD19-directed chimeric antigen receptor (CAR) T cell (CAR-T) therapy with axicabtagene ciloleucel (axi-cel) or tisagenlecleucel (tisa-cel) are approved for the treatment of relapsed or refractory large B cell lymphoma (LBCL), including de novo diffuse LBCL (DLBCL), primary mediastinal B cell lymphoma (PMBCL), and transformed follicular lymphoma (tFL). Transformed nonfollicular lymphomas (tNFLs), including transformed marginal zone lymphoma (tMZL) and transformed chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) were not included in their respective pivotal studies. This study was conducted to evaluate the outcomes of axi-cel and tisa-cel in tNFL patients, including those who received ibrutinib concomitantly through apheresis, lymphodepletion, and CAR-T infusion. This single-center retrospective study included all patients with tCLL/SLL, tMZL, tFL, and DLBCL/PMBCL treated with CAR-T therapy outside of a clinical trial setting from November 2017 to May 2021 at Moffitt Cancer Center, Tampa, Florida. We analyzed and compared outcomes in patients with tCLL/SLL or tMZL and patients with DLBCL/tFL. The study included 134 patients who received a total of 136 CAR-T treatments (111 with axi-cel and 25 with tisa-cel). Ninety patients had de novo DLBCL/PMBCL, 23 had tFL, and 21 had tNFL (12 with tMZL and 9 with tCLL/SLL). The overall response and complete response rates were 66.7% and 55.6%, respectively, for tCLL/SLL and 92.9% and 71.4% for tMZL. The overall response and complete response rates were not different between tNFL and DLBCL/tFL (P = .92 and .81, respectively). At a median follow-up of 21.3 months, the median progression-free survival (PFS) for tCLL/SLL was 5.4 months (95% confidence interval [CI], .8 month to not assessable [NA]); for tMZL, the median PFS was not reached (NR) (95% CI, 2.3 months to NA); and for DLBCL/tFL, the median PFS was 14.3 months (95% CI, 5.6 months to NA) (P = .58). The estimated 1-year PFS rate was 29.6% (95% CI, 5.2% to 60.7%) for tCLL/SLL, 50.0% (95% CI, 22.9% to 72.2%) for tMZL, 42.7% (95% CI, 22.4% to 61.6%) for tNFL, and 53.0% (95% CI, 42.3% to 62.5%) for DLBCL/tFL. The median overall survival was NR (95% CI, 9.2 months to NA) for tCLL/SLL, 27.1 months (95% CI, 8.5 months to NA) for tMZL, and NR (95% CI, 17.4 months to NA) for DLBCL/tFL (P = .79). Compared to the DLBCL/tFL cohort, tNFL patients were more likely to develop immune effector cell-associated neurologic syndrome (ICANS) and to receive tocilizumab (P = .04 and .01, respectively, after controlling for CAR-T product) and with a possibly higher incidence of grade ≥3 cytokine release syndrome (CRS) (P = .07). Two patients in the tNFL cohort died of treatment-related toxicity after receiving axi-cel. Six tNFL patients received ibrutinib concurrently with tisa-cel, with 1 case of grade ≥3 CRS/ICANS that rapidly resolved and no other severe toxicities. Our case series supports the use of CD19 CAR-T therapy in relapsed/refractory tCLL/SLL and tMZL. The concurrent use of ibrutinib and tisa-cel in tNFL was associated with manageable toxicity in tNFL.

A non-antibiotic-disrupted gut microbiome is associated with clinical responses to CD19-CAR-T cell cancer immunotherapy.

Increasing evidence suggests that the gut microbiome may modulate the efficacy of cancer immunotherapy. In a B cell lymphoma patient cohort from five centers in Germany and the United States (Germany, n = 66; United States, n = 106; total, n = 172), we demonstrate that wide-spectrum antibiotics treatment ('high-risk antibiotics') prior to CD19-targeted chimeric antigen receptor (CAR)-T cell therapy is associated with adverse outcomes, but this effect is likely to be confounded by an increased pretreatment tumor burden and systemic inflammation in patients pretreated with high-risk antibiotics. To resolve this confounding effect and gain insights into antibiotics-masked microbiome signals impacting CAR-T efficacy, we focused on the high-risk antibiotics non-exposed patient population. Indeed, in these patients, significant correlations were noted between pre-CAR-T infusion Bifidobacterium longum and microbiome-encoded peptidoglycan biosynthesis, and CAR-T treatment-associated 6-month survival or lymphoma progression. Furthermore, predictive pre-CAR-T treatment microbiome-based machine learning algorithms trained on the high-risk antibiotics non-exposed German cohort and validated by the respective US cohort robustly segregated long-term responders from non-responders. Bacteroides, Ruminococcus, Eubacterium and Akkermansia were most important in determining CAR-T responsiveness, with Akkermansia also being associated with pre-infusion peripheral T cell levels in these patients. Collectively, we identify conserved microbiome features across clinical and geographical variations, which may enable cross-cohort microbiome-based predictions of outcomes in CAR-T cell immunotherapy.

CYAD-01, an autologous NKG2D-based CAR T-cell therapy, in relapsed or refractory acute myeloid leukaemia and myelodysplastic syndromes or multiple myeloma (THINK): haematological cohorts of the dose escalation segment of a phase 1 trial.

BACKGROUND: CYAD-01 is an autologous chimeric antigen receptor (CAR) T-cell product based on the natural killer (NK) group 2D (NKG2D) receptor, which binds eight ligands that are overexpressed in a wide range of haematological malignancies but are largely absent on non-neoplastic cells. Initial clinical evaluation of a single infusion of CYAD-01 at a low dose in patients with relapsed or refractory acute myeloid leukaemia, myelodysplastic syndromes, and multiple myeloma supported the feasibility of the approach and prompted further evaluation of CYAD-01. The aim of the present study was to determine the safety and recommended phase 2 dosing of CYAD-01 administered without preconditioning or bridging chemotherapy. METHODS: The multicentre THINK study was an open-label, dose-escalation, phase 1 study for patients with relapsed or refractory acute myeloid leukaemia, myelodysplastic syndromes, or multiple myeloma, after at least one previous line of therapy. Patients were recruited from five hospitals in the USA and Belgium. The dose-escalation segment evaluated three dose levels: 3 × 108 (dose level one), 1 × 109 (dose level two), and 3 × 109 (dose level three) cells per infusion with a 3 + 3 Fibonacci study design using a schedule of three infusions at 2-week intervals followed by potential consolidation treatment consisting of three additional infusions. The occurrence of dose-limiting toxicities post-CYAD-01 infusion was assessed as the primary endpoint in the total treated patient population. The trial was registered with ClinicalTrials.gov, NCT03018405, and EudraCT, 2016-003312-12, and has been completed. FINDINGS: Between Feb 6, 2017, and Oct 9, 2018, 25 patients were registered in the haematological dose-escalation segment. Seven patients had manufacturing failure for insufficient yield and two had screening failure. 16 patients were treated with CYAD-01 (three with multiple myeloma and three with acute myeloid leukaemia at dose level one; three with acute myeloid leukaemia at dose level two; and six with acute myeloid leukaemia and one with myelodysplastic syndromes at dose level three). Median follow-up was 118 days (IQR 46-180). Seven patients (44%) had grade 3 or 4 treatment-related adverse events. In total, five patients (31%) had grade 3 or 4 cytokine release syndrome across all dose levels. One dose-limiting toxicity of cytokine release syndrome was reported at dose level three. No treatment-related deaths occurred, and the maximum tolerated dose was not reached. Three (25%) of 12 evaluable patients with relapsed or refractory acute myeloid leukaemia or myelodysplastic syndromes had an objective response. Among responders, two patients with acute myeloid leukaemia proceeded to allogeneic haematopoietic stem-cell transplantation (HSCT) after CYAD-01 treatment, with durable ongoing remissions (5 and 61 months). INTERPRETATION: Treatment with a multiple CYAD-01 infusion schedule without preconditioning is well tolerated and shows anti-leukaemic activity, although without durability outside of patients bridged to allogeneic HSCT. These phase 1 data support the proof-of-concept of targeting NKG2D ligands by CAR T-cell therapy. Further clinical studies with NKG2D-based CAR T-cells are warranted, potentially via combinatorial antigen targeted approaches, to improve anti-tumour activity. FUNDING: Celyad Oncology.

Human Effectors of Acute and Chronic GVHD Overexpress CD83 and Predict Mortality.

PURPOSE: Acute and chronic GVHD remain major causes of transplant-related morbidity and mortality (TRM) after allogeneic hematopoietic cell transplantation (alloHCT). We have shown CD83 chimeric antigen receptor (CAR) T cells prevent GVHD and kill myeloid leukemia cell lines. In this pilot study, we investigate CD83 expression on GVHD effector cells, correlate these discoveries with clinical outcomes, and evaluate critical therapeutic implications for transplant recipients. EXPERIMENTAL DESIGN: CD83 expression was evaluated among circulating CD4+ T cells, B-cell subsets, T follicular helper (Tfh) cells, and monocytes from patients with/without acute or chronic GVHD (n = 48 for each group), respectively. CD83 expression was correlated with survival, TRM, and relapse after alloHCT. Differential effects of GVHD therapies on CD83 expression was determined. RESULTS: CD83 overexpression on CD4+ T cells correlates with reduced survival and increased TRM. Increased CD83+ B cells and Tfh cells, but not monocytes, are associated with poor posttransplant survival. CD83 CAR T eliminate autoreactive CD83+ B cells isolated from patients with chronic GVHD, without B-cell aplasia as observed with CD19 CAR T. We demonstrate robust CD83 antigen density on human acute myeloid leukemia (AML), and confirm potent antileukemic activity of CD83 CAR T in vivo, without observed myeloablation. CONCLUSIONS: CD83 is a promising diagnostic marker of GVHD and warrants further investigation as a therapeutic target of both GVHD and AML relapse after alloHCT.

Efficacy and Safety of Erenumab, Galcanezumab, and Fremanezumab in the Treatment of Drug-Resistant Chronic Migraine: Experience in Real Clinical Practice.

BACKGROUND: Due to the recent introduction of new biologic drugs for chronic migraine, a global evaluation in real clinical practice is necessary. OBJECTIVE: The objective was to evaluate the effectiveness and safety in real clinical practice of drugs targeting the calcitonin gene-related peptide receptor (CGRPr) in patients with chronic migraine. METHODS: Single-center, restrospective study (2019-2022), including patients with chronic migraine treated with erenumab, galcanezumab, or fremanezumab. Effectiveness variables were recorded, namely, number of migraine headache days per month (MHD), Migraine Disability Assessment Scale (MIDAS) score, and Headache Impact Test-6 (HIT-6) score, assessing changes at week 12, 24 from baseline. Toxicity was recorded following the Common Terminology Criteria for Adverse Events (CTCAE) v5.0 criteria. RESULTS: In all, 104 patients were included (46.2% erenumab, 41.3% galcanezumab, 12.5% fremanezumab). A reduction in MHD, MIDAS, and HIT-6 was achieved at weeks 12 and 24 with erenumab (p75% at week 24 than those intensified; P = 0.041). There was no difference in efficacy (P = 0.154) or improvement in quality of life (P = 0.783, P = 0.150), but there was greater toxicity (P < 0.001) among nonresponders with erenumab 70 mg versus erenumab 140 mg. CONCLUSIONS: The results confirm the effectiveness and safety of anticalcitonin gene-related peptide (CGRP) drugs in real clinical practice. However, the study shows little benefit from erenumab intensification, with similar effectiveness and worse tolerability than the standard dose.

Large Scale Ex Vivo Expansion of γδ T cells Using Artificial Antigen-presenting Cells.

Higher γδ T cell counts in patients with malignancies are associated with better survival. However, γδ T cells are rare in the blood and functionally impaired in patients with malignancies. Promising results are reported on the treatment of various malignancies with in vivo expansion of autologous γδ T cells using zoledronic acid (zol) and interleukin-2 (IL-2). Here we demonstrated that zol and IL-2, in combination with a novel genetically engineered K-562 CD3scFv/CD137L/CD28scFv/IL15RA quadruplet artificial antigen-presenting cell (aAPC), efficiently expand allogeneic donor-derived γδ T cells using a Good Manufacturing Practice (GMP) compliant protocol sufficient to achieve cell doses for future clinical use. We achieved a 633-fold expansion of γδ T cells after day 10 of coculture with aAPC, which exhibited central (47%) and effector (43%) memory phenotypes. In addition, >90% of the expanded γδ T cells expressed NKG2D, although they have low cell surface expression of PD1 and LAG3 inhibitory checkpoint receptors. In vitro real-time cytotoxicity analysis showed that expanded γδ T cells were effective in killing target cells. Our results demonstrate that large-scale ex vivo expansion of donor-derived γδ T cells in a GMP-like setting can be achieved with the use of quadruplet aAPC and zol/IL-2 for clinical application.

Impact of carbon dioxide values after aortic impingement on postoperative morbimortality in pediatric patients undergoing cardiac surgery / Impacto de los valores de bióxido de carbono post-pinzamiento aórtico sobre la morbimortalidad postoperatoria en pacientes pediátricos sometidos a cirugía cardíaca

Abstract: Introduction: Inducing hypocapnia is a common practice during pediatric general anesthesia, even though it has not shown clear benefits. Objective: To compare the impact of carbon dioxide values after aortic impingement (< 32.7 vs ≥ 32.7 mmHg) on postoperative morbimortality among pediatric patients undergoing cardiac surgery. Material and methods: A case-control study included 90 pediatric patients undergoing cardiac surgery with cardiopulmonary bypass. The study cases consisted of 45 patients who died within 30 days of the postoperative period. Cases and controls were individually matched (1:1 ratio). Descriptive and inferential statistics (Mann-Whitney's U, Student's t and χ2 tests) were used to analyze the results. A p < 0.05 was considered significant. A univariate analysis was also carried out. The strength of association between morbimortality and carbon dioxide values after aortic impingement was determined using the odds ratio. The data were processed using SPSS v-24.0. Results: The group with carbon dioxide values of < 32.7 mmHg after aortic impingement was associated with greater morbidity (OR 24.75; 95% CI 4.92-124.32) and mortality (OR 22.47; 95% CI 4.85-10.17) at 30 days. Conclusion: Pediatric patients undergoing cardiac surgery with carbon dioxide values of < 32.7 mmHg after aortic impingement showed higher postoperative morbimortality than those with carbon dioxide values of ≥ 32.7 mmHg.

Resumen: Introducción: La hipocapnia es una práctica común durante la anestesia general pediátrica; sin embargo, a lo largo del tiempo no ha mostrado beneficios bien definidos. Objetivo: Comparar el impacto del bióxido de carbono post-pinzamiento aórtico (< 32.7 vs ≥ 32.7 mmHg) sobre la morbimortalidad postoperatoria en los pacientes pediátricos sometidos a cirugía cardíaca. Material y métodos: Se realizó un estudio de casos y controles que incluyó 90 pacientes pediátricos sometidos a cirugía cardíaca con derivación cardiopulmonar. Se consideraron casos 45 pacientes que fallecieron dentro de los 30 días del postoperatorio. Los controles fueron pareados en relación 1:1. Para su análisis se realizó estadística descriptiva e inferencial con U de Mann-Whitney, t de Student y χ2 según fue el caso. Una p < 0.05 fue significativa. Se realizó un análisis univariado. La fuerza de asociación entre la morbimortalidad y los valores de bióxido de carbono post-pinzamiento aórtico se obtuvo mediante el odds ratio. Los datos fueron procesados mediante SPSS v-24.0. Resultados: El grupo con valores de bióxido de carbono post-pinzamiento aórtico < 32.7 mmHg se asoció con una mayor morbilidad a los 30 días (OR 24.75; IC del 95% 4.92-124.32) y mortalidad (OR 22.47; IC del 95% 4.85-10.17). Conclusión: Los pacientes pediátricos sometidos a cirugía cardíaca con valores de bióxido de carbono post-pinzamiento aórtico < 32.7 mmHg tienen mayor morbimortalidad postoperatoria que los que tienen valores ≥ 32.7 mmHg.