Developing anti-factor VIII (FVIII) inhibitory antibodies (inhibitors) are a significant complication of FVIII protein replacement therapy in hemophilia A. Our previous study demonstrated that follicular helper T (TFH) cells play a critical role in FVIII inhibitor development. Follicular regulatory T (TFR) cells are a subset of forkhead box protein P3 positive (Foxp3+) T cells identified in the germinal center that can modulate TFH cell activation of B cells and antibody development. Here, we report that FVIII immunization significantly increases the TFR cells in the spleens of FVIII inhibitor-producing FVIIInull mice compared with saline-treated controls and non-inhibitor-producing animals. The TFH/TFR ratio significantly increased in FVIII inhibitor-producing mice. The emergence of TFR cells correlated with titers of FVIII inhibitors in FVIII-immunized mice. Using TFR-deficient Foxp3Cre+Bcl6fl/fl (Bcl6FC) mice, we found that the loss of TFR cells led to significantly decreased FVIII inhibitors compared with wild-type (WT) mice on FVIII immunization (24 ± 16 and 131 ± 114 Bethesda unit (BU)/mL, respectively) but not total anti-FVIII IgG levels and that TFR cells regulated IgG subclass switching and FVIII-specific B-cell responses. Interestingly, on FVIII immunization, mice with phosphatase and tensin (Pten) deficiency in Foxp3+ cells (Foxp3Cre+Ptenfl/fl), a model with augmented TFR cells, developed markedly lower FVIII inhibitor titers (8.1 ± 8.6 BU/mL) than WT controls. When CD4Cre+Bcl6fl/fl mice, a TFH- and TFR-deficient model, were immunized with FVIII, none of the animals developed FVIII inhibitors. In conclusion, FVIII immunization induces TFR cell activation and expansion. TFR cells have a dual function in regulating the development of FVIII inhibitors, and the TFH/TFR pathway is pivotal in FVIII inhibitor development in mice.

Although allogeneic hematopoietic stem cell transplantation is a leading treatment approach for myeloid malignancies, challenges in its immune biology and in treatment approaches remain. In the past decade, major advances in the knowledge of mechanisms of graft-versus-host disease (GVHD) has allowed development of new treatments both for GVHD prophylaxis and treatment. However, although successes did occur, failure did as well. Reasons for failure can be linked either to incomplete understanding of the pathophysiology of GVHD, or, in some cases, to errors in the design of clinical trials. Better GVHD prophylaxes and disease control have likely led to decreased nonrelapse mortality (NRM). However, although NRM rates have decreased, rates of relapse of the original malignancy have not significantly improved. Our current understanding of the biology of the graft-versus-leukemia (GVL) effect still lags behind that of GVHD, and treatment approaches to manipulate the GVL effect remain limited. The reasons for such a lag are numerous, but improved knowledge of the biology of hematological malignancies open the gate to new developments, providing that we can better understand the interplay between the immune system with leukemic clones. From a therapeutical perspective, much attention has been paid to the results from randomized clinical trials and from a biological perspective on recent discoveries, especially in the human setting. The objective of this perspective is to analyze what are the current challenges in the biology and treatment of GVHD and GVL and to provide a personal view on how some biological and therapeutic issues could be approached.

Disease response and progression assessment in multiple myeloma is based on various measurements of monoclonal protein (serum and urine protein electrophoresis, serum free light chain, and/or quantitative immunoglobulins). Currently, the International Myeloma Working Group consensus response criteria require 2 sequential assessments of any 1 marker made at any time before confirmation of disease progression and the institution of any new therapy. However, this can be cumbersome in clinical trials. Herein, we hypothesized that if 2 markers meet the progression criteria simultaneously, a repeat of either will not be necessary for confirmation. We retrospectively studied all sequential patients with myeloma enrolled in clinical trials at Mayo Clinic. We identified 583 episodes of confirmed progression in our study. Among the 583 progression episodes, nearly 70% (sensitivity of the simultaneous criteria) met the 2 simultaneous variable criteria at the first testing, indicating progression. Conversely, among 413 patients who met progression criteria by 2 simultaneous values, 98% (specificity of the simultaneous criteria) of patients subsequently had confirmed progression by sequential values. In summary, for patients with 2 disease burden markers meeting the simultaneous progression criteria, sequential assessment of either 1 for confirmation may not be necessary to determine disease progression.

The human genome contains regulatory DNA elements, known as enhancers, that can activate gene transcription over long chromosomal distances. Here, we showed that enhancer distance can be critical for gene silencing. We demonstrated that linear recruitment of the normally distal strong HBB enhancer to developmentally silenced embryonic HBE or fetal HBG promoters through deletion or inversion of intervening DNA sequences led to strongly reactivated expression in adult erythroid cells and ex vivo differentiated hematopoietic stem and progenitor cells. A similar observation was made for the HBA locus in which deletion-to-recruit of the distal enhancer strongly reactivated embryonic HBZ expression. Overall, our work assigned function to seemingly nonregulatory genomic segments; by providing linear separation, they may support genes to autonomously control their transcriptional response to distal enhancers.

Chronic lymphocytic leukemia (CLL) is a disease of great clinical and biologic heterogeneity; some patients are observed for years without symptoms, whereas others rapidly develop progressive disease requiring treatment. With therapy, some patients eventually develop resistant CLL or transformation to an aggressive form. Across this spectrum, patients experience immune dysfunction associated with increased risk for infection and second cancers, contributing to morbidity and mortality of the disease. The ultimate therapeutic bull's-eye for CLL is to eliminate the disease and achieve immune restoration. Disease elimination can potentially be achieved for a fraction of patients treated first line with chemoimmunotherapy (fludarabine, cyclophosphamide, and rituximab), for some patients who receive time-limited combined targeted therapy, and for some patients with relapsed/refractory CLL who undergo allogeneic stem cell transplant. Long-term immune restoration for these patients is elusive. Current targeted therapies, including Bruton tyrosine kinase and B-cell lymphoma 2 inhibitors and CD20 monoclonal antibodies used in combinations, can produce exceptional therapeutic outcomes, which are improving survival for patients who need treatment. Although clear progress has been made toward highly effective CLL management, appreciation of the full impact of these advances will require time because of the chronic nature of the disease. In addition, it is imperative to ensure global access to the targeted therapies, emphasizing the need for harmonized regulatory oversight and affordable treatment options worldwide. Here, we discuss research and collaborative strategies to refine the use of targeted agents to eliminate CLL and restore immune function for all affected individuals.

Relapsed/refractory T-cell acute lymphoblastic leukemia (ALL)/lymphoma (LBL) represent a significant unmet medical need. WU-CART-007 is a CD7-targeting, allogeneic, fratricide-resistant chimeric antigen receptor T cell product generated from healthy donor T cells. WU-CART-007 was evaluated in a phase 1/2 study with a 3+3 dose-escalation design followed by cohort expansion in relapsed/refractory T-ALL/LBL. Patients received one infusion of WU-CART-007 after standard or enhanced lymphodepleting chemotherapy. The primary objectives, to characterize safety and assess the composite complete remission rate, were met. Of 28 patients enrolled, 13 received the recommended phase 2 dose (RP2D) of 900 million cells of WU-CART-007 with enhanced lymphodepletion. The most common treatment-related adverse event was cytokine release syndrome (88.5%; 19.2% grade 3-4). Two grade 1 immune effector cell-associated neurotoxicity syndrome events (7.7%) and one grade 2 acute graft-vs-host disease event occurred (3.8%). One grade 2 immune effector cell associated HLH-like syndrome (IEC-HS) was observed. Among the 11 patients evaluable for response at the RP2D who received enhanced lymphodepleting chemotherapy, the overall response rate was 90.9% and composite complete remission rate was 72.7%. WU-CART-007 at the RP2D demonstrated a high response rate in patients with relapsed/refractory T-ALL/LBL and has the potential to provide a new treatment option. ClinicalTrials.gov registration: NCT04984356.