Loss Of CD4 Research Articles

GABA-induced monocytic reactive oxygen species impair CD4 restoration in treated HIV-1 adults.

In order to better understand why about 15% of people living with Human Immunodeficiency Virus-1 (PWH) on highly active antiretroviral therapy do not restore their CD4 count, we explored the link previously reported between glutamate plasma level and CD4 count. We recruited fourty-four adults living with HIV-1 aviremic under antiretroviral therapy. Their peripheral blood concentrations in glutamate and GABA were determined by ELISA. Flow cytometry was used to detect GABA receptor, reactive oxygen species (ROS) produced by monocytes, and programmed T cell death. DNA-dependent protein kinase (DNA-PK) and p53 phosphorylation were analyzed by western blot. DNA damage was quantified by immunofluorescence. We show that i) some virologic responders present high plasma levels of glutamate and of its derivative, GABA; ii) monocytes express the GABA receptor GABA-B1; iii) GABA-B1 stimulation induces monocytic ROS production; iv) GABA-B1-overexpressing monocytes of PWH with high plasma levels of GABA release high amount of ROS; and v) monocyte-derived ROS oxidise the DNA of CD4+ T cells, creating double-strand breaks which activate DNA-PK and p53, and finally apoptosis. The intensity of this cascade of events is inversely correlated with the slope of CD4+ T cell recovery in treated PWH. We propose that DNA damage resulting from ROS produced by GABA-activated monocytes plays a key role in impaired immune restoration. Consequently, GABA-B1 antagonists and/or ROS inhibitors might be a promising therapy for non-immunologic responders. Furthermore, the same mechanism could be involved in CD4 loss in the natural course of the infection.

CD4 expression controls epidermal stem cell balance

The balance of stem cell populations is essential for the maintenance, renewal, and repair of the mammalian epidermis. Here, we report that CD4, which is a typical marker of helper T cells, monocytes, macrophages, and dendritic cells, is also expressed on murine K5+ keratinocytes. Lineage tracing of CD4+ cells reveals that their epidermal progeny has self-renewal abilities and clonogenic potential. The progeny of CD4+ epidermal cells contributes to epidermal renewal and progressively colonizes the interfollicular epidermis and hair follicles with age, thereby developing to all epidermal lineages. Wound healing studies furthermore show that the progeny of CD4+ epidermal cells accumulates at wound sites. Finally, using CD4 knockout mice we demonstrate that CD4 expression is essential for maintaining fast-cycling epidermal stem cells during homeostasis and that CD4 loss mitigates the age-related decline in wound repair capacity. Collectively, our data support the conclusion that CD4 expression is required for long-term maintenance of the epidermal stem cell balance.

The Pathogenic Role of Expanded CD8⁺CD28null Angiogenic T Cells in ANCA-Associated Vasculitis.

Objectives: Angiogenic T cells (Tang) are crucial in promoting angiogenesis, with the loss of CD28 serving as a marker for highly differentiated and senescent T cells. This study aims to investigate the characteristics and potential roles of CD8+CD28null Tang in patients with ANCA-associated vasculitis (AAV). Methods: A cohort of AAV patients and matched healthy controls (HCs) were analyzed. Flow cytometry was used to assess the profiles of circulating CD8+CD28null Tang. In vitro functional assays were performed to evaluate the pathogenic properties of CD8+CD28null Tang. Results: CD8+CD28null Tang levels were significantly higher in the peripheral blood of AAV patients compared to HCs, and their levels were further increased in AAV patients with MPO⁺, p-ANCA⁺, or interstitial lung disease compared to their respective control groups. Additionally, there was a positive correlation between both the percentage and absolute count of CD8+CD28null Tang and the Birmingham Vasculitis Activity Score (BVAS). In patients with a good treatment response, both the percentage and absolute count of CD8+CD28null Tang were significantly reduced, and this reduction was positively correlated with the decrease in BVAS scores. In vitro studies revealed that CD8+CD28null Tang displayed enhanced chemotactic properties, induced apoptosis in human umbilical vein endothelial cells (HUVECs), and inhibited their proliferation, migration, and tube formation. Conclusions: AAV patients exhibit increased levels of circulating CD8+CD28null Tang, which can be reduced following effective treatment. Furthermore, CD8+CD28null Tang may contribute to the pathogenesis of AAV by promoting apoptosis and inhibiting the proliferation, migration, and tube formation of HUVECs.

Differentiating reactive and neoplastic gamma-delta (γδ) T-cell expansions in the peripheral blood and bone marrow.

The clinical and immunophenotypic attributes of reactive γδ T-cell expansions are less well characterized than their malignant counterparts, which can pose diagnostic challenges. This study aims to investigate the characteristics and long-term clinical outcomes of reactive γδ T-cell expansions. A retrospective review was performed to identify patients with expanded γδ T-cell population (>15% of T-cells) by flow cytometry in peripheral blood and/or bone marrow specimens over a 17-year period. The cases were divided into reactive and malignant categories and their clinical and immunophenotypic findings were compared. Clinical follow-up was performed. 97 patients were identified including 19 malignant and 78 reactive cases with a variety of underlying conditions. The median absolute γδ T-cell count and median percentage of γδ T-cells per total T-cells were significantly lower in reactive vs. malignant cases (p = 0.0001 and p < 0.00001, respectively). Reactive cases showed more frequent brighter surface CD3 expression (87.1% vs. 42.1%; p < 0.0001), no discrete loss of CD7 (0% vs. 36.9%; p < 0.0001), less frequent lack of CD5 (25.7% vs. 42.4%; p < 0.0001), and no homogeneous CD56 expression (0% vs. 31.6%; p > 0.0001) as compared with malignant cases. Upon long-term follow-up, none of the reactive cases showed clinical evidence of malignant evolution. Reactive expansions of γδ T-cells can be seen in a variety of conditions including hematologic neoplasms, autoimmune and post-transplant states, and infections. Such cases have significantly lower γδ T-cell counts and percentages and no discrete loss of CD7. Lack of CD5 on its own is not an indication of immunophenotypic aberrancy in γδ T-cells. Upon long-term clinical follow-up, such reactive expansions show no evidence of evolution to γδ T-cell malignancies.

Autoimmune-Like Mechanism in Heart Failure Enables Preventive Vaccine Therapy.

Heart failure (HF) is strongly associated with inflammation. In pressure overload (PO)-induced HF, cardiac stress triggers adaptive immunity, ablation, or inhibition, which blocks disease progression. We hypothesized that PO-HF might fulfill the often-used criteria of autoimmunity: if so, the associated adaptive immune response would be not only necessary but also sufficient to induce HF; it should also be possible to identify self-antigens driving the autoimmune response. Finally, we hypothesized that such an antigen-specific response can be manipulated to preventively reduce the severity of PO-HF in a tolerizing vaccine. We used the transfer of lymphocytes or serum from PO-HF mice into healthy recipients to assess whether the adaptive response is sufficient to induce disease. We devised a novel pipeline to identify self-antigens driving the response. We immunized healthy mice with novel antigens to assess whether they induce disease. To determine whether these antigens could be present in human patients, we sought to detect existing responses against these antigens in patients with HF. Finally, we used the antigens in an oral tolerance protocol to preventively protect mice from subsequently induced PO-HF, analyzing the results with next-generation sequencing. We found that PO-HF fulfills the criteria of an autoimmune disease, albeit partially, and identified novel cardiac self-antigens, capable of inducing cardiac dysfunction. The novel antigens in a tolerizing vaccine formulation preemptively reduced the severity of disease triggered by subsequent application of PO, via induction of effector regulatory T cells, enabling a potent reduction of PO-driven loss of systolic function, cardiac inflammation, and proinflammatory CD4+ T-cell clonal expansion. We demonstrate that PO-HF is triggered by hemodynamic stress and then sets off an autoimmune-like response against cardiac self-antigens. The antigens can be used to reduce the severity of future-onset disease, via oral tolerization, effectively acting as a protective vaccine.

Thymic and T-cell intrinsic critical roles associated with severe combined immunodeficiency and Omenn syndrome due to a heterozygous variant (G201R) in PSMB10.

Heterozygous immunoproteasome 20 S subunit beta 10 (PSMB10) mutations can cause severe combined immunodeficiency and Omenn syndrome. Hematopoietic stem cell transplantation in these patients is associated with severe complications and poor immune reconstitution, often resulting in death. We sought to perform immunologic and molecular characterization of an infant with a PSMB10 heterozygous variant. A heterozygous variant in PSMB10 (p.G201R) was identified in the index patient but not her parents. Detailed immunophenotyping and functional studies, including flow cytometry, immunoblotting, and T-cell development in artificial thymic organoids, were performed. The patient presented with severe B-, natural killer-, and T-cell lymphopenia, with a progressive increase in memory CD4+ T cells and loss of CD8+ T cells, diminished Vbeta family diversity, and abnormal IL-7 signaling. Immunoproteasome protein expression (PSMB9 and PSMB10) was markedly reduced in the patient's cells, including PBMCs, EBV-transformed B cells, and fibroblasts, the mutation likely acting in a dominant-negative fashion. The patient's CD34+ cells showed a normal early T-cell development but slightly impaired generation of CD3+TCR αβ+ cells in artificial thymic organoids, and human thymus single-cell RNA sequencing demonstrated that PSMB10 is expressed in different subsets of cortical and medullary thymic epithelial cells. Collectively, these data indicate that PSMB10 mutations affect positive selection of CD8 T cells, generation of a diverse T-cell repertoire, and negative selection of autoreactive T cells. The PSMB10 G201R variant is associated with reduced immunoproteasome expression levels that appear to play vital roles in hematopoietic and extrahematopoietic immune system development and function. PSMB10-associated thymoproteasome dysfunction leads to impaired thymopoiesis and the development of severe combined immunodeficiency and Omenn syndrome, suggesting possible benefit from thymus implantation.

Differentiating mycosis fungoides lesions from their mimickers clinically and histologically: A single tertiary center retrospective analysis in Saudi Arabia.

To identify the clinical and histological features of MF that can assist in distinguishing MF from MF-mimicking cases. Although mycosis fungoides (MF) is the most common subtype of cutaneous T-cell lymphoma, clinicopathological correlations are required to establish an accurate diagnosis, which are currently lacking. This retrospective observational study evaluated the clinical presentations, characteristics, and histological features of 56 patients with suspected MF who presented to our clinic between January 2018 and August 2022. Immunohistochemistry was performed, and the loss of CD5 and CD7T-cells and T-cell receptor rearrangement was evaluated. Overall, 34 patients were diagnosed with MF, whereas 22 were not. Clinical erythroderma, poikiloderma, and nodular presentations were more commonly associated with a histological diagnosis of MF than macular presentations. Erythema and pruritus were significantly more common in MF cases than in MF-mimicking cases (p<0.05). Epidermotropism and parakeratosis were the key histological features for diagnosing MF. Additionally, Pautrier's microabscesses correlated with the clinical presentation of plaques in MF. Loss of CD7 expression on the T-cell surface was observed even in early-stage MF cases. Our proposed diagnostic features are statistically valid and, along with those previously reported, can aid in identifying and distinguishing MF cases from MF-mimicking cases.

Retrospective Study of CD20 Expression Loss in Relapsed or Refractory B-Cell Non-Hodgkin Lymphoma.

CD20-targeted therapies are widely used in the management of B-cell lymphomas. Re-treatment with CD20-directed agents is common; however, previous research has demonstrated loss of CD20 expression at relapse in a subset of patients. In this single-center retrospective cohort of 243 patients, CD20 analysis was performed by immunohistochemistry (IHC) and/or flow cytometry at diagnosis and at relapse if a biopsy was performed. Of 109 patients with relapsed or refractory B-cell lymphoma, 59 patients with CD20-positive lymphoma at diagnosis underwent a biopsy at relapse for a total of 76 biopsies across all relapses. The rate of partial or complete CD20 expression loss was 11.9% (four patients with partial loss, three patients with complete loss). There were four cases of CD20 loss at first relapse (three IHC, one flow cytometry), two at second relapse (one IHC, one IHC and flow cytometry), and one at fifth relapse (IHC and flow cytometry). CD20 antigen escape was observed in marginal zone lymphoma, follicular lymphoma, and diffuse large B-cell lymphoma (DLBCL). All patients with CD20 expression loss previously received rituximab. Among patients with CD20 antigen escape, 85.7% had stage IV disease, and median overall survival after CD20 loss was 4 months. In the group of five patients with indolent lymphoma and CD20 expression loss, three patients (60%) had concurrent transformation to high-grade lymphoma. This study, which reinforces the importance of repeating a biopsy at relapse before implementing CD20-directed therapy, is particularly relevant given the widespread use of rituximab along with the emerging significance of CD20-targeted bispecific antibodies in the management of B-cell lymphomas.

Chronic inflammation degrades CD4 T cell immunity to prior vaccines in treated HIV infection

To date, our understanding of how HIV infection impacts vaccine-induced cellular immunity is limited. Here, we investigate inflammation, immune activation and antigen-specific T cell responses in HIV-uninfected and antiretroviral-treated HIV-infected people. Our findings highlight lower recall responses of antigen-specific CD4 T cells that correlate with high plasma cytokines levels, T cell hyperactivation and an altered composition of the T subsets enriched with more differentiated cells in the HIV-infected group. Transcriptomic analysis reveals that antigen-specific CD4 T cells of the HIV-infected group have a reduced expression of gene sets previously reported to correlate with vaccine-induced pathogen-specific protective immunity and further identifies a consistent impairment of the IFNα and IFNγ response pathways as mechanism for the functional loss of recall CD4 T cell responses in antiretroviral-treated people. Lastly, in vitro treatment with drugs that reduce inflammation results in higher memory CD4 T cell IFNγ responses. Together, our findings suggest that vaccine-induced cellular immunity may benefit from strategies to counteract inflammation in HIV infection.

Nanobody-based naturally selected CD7-targeted CAR-T therapy for acute myeloid leukemia

AbstractApproximately 30% of patients with acute myeloid leukemia (AML) express CD7 on their myeloblasts. We have previously demonstrated that single-chain variable fragment (scFv)–based “naturally selected” CD7 chimeric antigen receptor T-cell (NS7CAR-T) therapy shows significant efficacy, with a favorable safety profile in T-cell lymphoid malignancies. Here, we derived dual nanobody-based dVHH NS7CAR-Ts that have superior CD7 binding specificity, affinity to their scFv-based counterparts, and improved proliferative capability. In this phase 1 clinical trial, we evaluated the efficacy and safety of dVHH NS7CAR-Ts for patients with CD7+ refractory/relapsed AML. A cohort of 10 patients received dVHH NS7CAR-Ts across 2 dosage levels of 5 × 105/kg and 1 × 106/kg. Before enrollment, patients had undergone a median of 8 (range, 3-17) prior lines of therapy. Seven patients had prior transplants. After NS7CAR-T infusion, 7 of 10 (70%) patients achieved complete remission (CR). The median observation time was 178 days (range, 28-776). Among 7 patients who achieved CR, 3 who relapsed from prior transplants underwent a second allogeneic hematopoietic stem cell transplant. One patient remained leukemia free on day 401, and the other 2 died on day 241 and day 776, respectively, from nonrelapse-related causes. Three CR patients without consolidative allogeneic hematopoietic stem cell transplant relapsed within 90 days. All the nonresponders and relapsed patients had CD7 loss. The treatment was well tolerated, with 80% experiencing mild cytokine release syndrome and none had neurotoxicity. This trial underscores the potential promising treatment of dVHH NS7CAR-Ts in providing clinical benefits with a manageable safety profile to patients with CD7+ AML, warranting further investigation. This trial was registered at www.ClinicalTrials.gov as #NCT04938115.

ARI0003: Co-transduced CD19/BCMA dual-targeting CAR-T cells for the treatment of non-Hodgkin lymphoma

CD19 CAR-T therapy has achieved remarkable responses in relapsed/refractory non-Hodgkin lymphoma (NHL). However, challenges persist, with refractory responses or relapses after CAR-T administration linked to CD19 loss or downregulation. Given the co-expression of CD19 and BCMA in NHL, we hypothesized that dual targeting could enhance long-term efficacy. We optimized different dual-targeting approaches, including co-transduction of two lentiviral vectors, bicistronic, tandem, and loop and pool strategies, based on our academic anti-CD19 (ARI0001) and anti-BCMA (ARI0002h) CAR-T cells. Comparison with anti-CD19/CD20 or anti-CD19/CD22 dual targeting was also performed. We demonstrate that anti-CD19/BCMA CAR-T cells can be effectively generated through the co-transduction of two lentiviral vectors after optimization to minimize competition for cellular resources. Co-transduced Tcells, called ARI0003, effectively targeted NHL tumor cells with high avidity, outperforming anti-CD19 CAR-T cells and other dual-targeting approaches both invitro and invivo, particularly in low CD19 antigen density models. ARI0003 maintained effectiveness post-CD19 CAR-T treatment in xenograft models and in spheroids from relapsed CART-treated patients. ARI0003 CAR-T cells were effectively manufactured under Good Manufacturing Practice conditions, with a reduced risk of genotoxicity compared to other dual-targeting approaches. A first-in-human phase 1 clinical trial (CARTD-BG-01; this study was registered at ClinicalTrials.gov [NCT06097455]) has been initiated to evaluate the safety and efficacy of ARI0003 in NHL.

Central Memory CD4 T Cells from Persons with HIV Accumulate DNA Content Defects During Proliferative Response.

Central memory (TCM) cells are a subpopulation of CD4 T cells that sustain overall CD4 T cell counts in HIV infection. The mechanisms underlying their eventual demise, which leads to loss of CD4 T cell counts, are not known. To understand their proneness to death despite their increased movement to proliferation, we examined cell division together with possible cell accumulation in different phases of the cell cycle. Purified circulating TCM cells from untreated people living with HIV (PLWH) (n = 9) and healthy controls (n = 10) were stimulated in vitro using anti-CD3/CD28 agonistic antibodies plus IL-2 and cultured for 4 days. Cell viability, DNA content, proliferation, and cyclin A and cyclin B expression were measured. We found that PLWH TCM cells more frequently had a DNA content lower than G0/G1, compared with controls (p = .043). These cells accumulated with each division. The proportion of cells with sub-G0/G1 DNA content that were cycling (expressing cyclin A) was greater in the PLWH group (p = .003). The percentage of TCM cells expressing cyclin A+ among those in G0/G1 and was also greater in the PLWH group (p = .043), suggesting arrest before G2/M. While TCM cells from PLWH can proliferate, during this process some of them accumulate defects in DNA content that are incompatible with viability, suggesting that they could be intrinsically prone to cell cycle-dependent death. This provides a possible mechanism underlying the increased TCM cell turnover in HIV infection.

Allogeneic CD5-specific CAR-T therapy for relapsed/refractory T-ALL: a phase 1 trial.

Refractory or relapsed T cell acute lymphoblastic leukemia (r/r T-ALL) patients have poor prognoses, due to the lack of effective salvage therapies. Recently, CD7-targeting chimeric antigen receptor (CAR)-T therapies show efficacy in patients with r/r T-ALL, but relapse with CD7 loss is common. This study evaluates a CD5-gene-edited CAR-T cell therapy targeting CD5 in 19 r/r T-ALL patients, most of whom had previously failed CD7 CAR-T interventions. CAR-T products were derived from previous transplant donors (Cohort A) or newly matched donors (Cohort B). Primary endpoints were dose-limiting toxicity at 21 days and adverse events within 30 days. Secondary endpoints were responses, pharmacokinetics and severe adverse events after 30 days. A total of 16 received infusions, 10 at target dose of 1 × 106 kg-1. All encountered grade 3-4 cytopenias and one had a grade 3 infection within 30 days. All patients (100%) achieved complete remission or complete remission with incomplete blood count recovery by day 30. At a median follow-up of 14.3 months, four received transplantation; three were in remission and one died of infection. Of 12 untransplanted patients, 2 were in remission, 3 relapsed, 5 died of infection and 2 of thrombotic microangiopathy. CAR-T cells persisted and cleared CD5+ T cells. CD5- T cells, mostly CD5-gene-edited, increased but remained below normal levels. These results suggest this CD5-specific CAR-T intervention has a high remission rate for T-ALL patients. Evidence also suggests the risk of late-onset severe infection may be mitigated with consolidative transplantation. This study provides insights that could help to optimize this promising intervention. ClinicalTrials.gov registration: NCT05032599 .

Long-term Tolerance to Islet Transplantation via Targeted Reduction of beta cell-specific T cells.

Activated beta-cell reactive CD4 + and CD8 + T effector cells undergo a profound DNA-damage response which is targetable by small molecule inhibitors of the p53 and cell cycle pathways that lead to apoptosis. The use of a combination of MDM2 and WEE1 inhibitors, which termed "p53 potentiation with checkpoint abrogation" (PPCA), conferred significant therapeutic efficacy in treating mouse models of new onset T1D. Specific targeting of these T effector cells by PPCA results in a loss of inflammatory T cell subsets, notably proliferation CD4 + Th0 and Th1 subsets and CD8 + T effector memory cells, as determined by single cell RNA-seq studies with the preservation of T regulatory cells. When autologous islet grafts are given to established diabetic NOD mice, a single course of PPCA results in long-term islet graft acceptance, restoration of normoglycemia and loss of beta cell specific CD4 + and CD8 + T cells. PPCA shows promise as a potential means of estimating islet graft tolerance in T1D recipients of islet graft transplantation.

The histone methyltransferase Mixed-lineage-leukemia-1 drives T cell phenotype via Notch signaling in diabetic tissue repair.

Immune cell-mediated inflammation is important in normal tissue regeneration but can be pathologic in diabetic wounds. Limited literature exists on the role of CD4+ T cells in normal or diabetic wound repair; however, the imbalance of CD4+ Th17/Tregs has been found to promote inflammation in other diabetic tissues. Here, using human tissue and murine transgenic models, we identified that the histone methyltransferase Mixed-lineage-leukemia-1 (MLL1) directly regulates the Th17 transcription factor RORγ via an H3K4me3 mechanism and increases expression of Notch receptors and downstream Notch signaling. Furthermore, we found that Notch receptor signaling regulates CD4+ Th cell differentiation and is critical for normal wound repair, and loss of upstream Notch pathway mediators or receptors in CD4+ T cells resulted in the loss of CD4+ Th cell differentiation in wounds. In diabetes, MLL1 and Notch-receptor signaling was upregulated in wound CD4+ Th cells, driving CD4+ T cells toward the Th17 cell phenotype. Treatment of diabetic wound CD4+ T cells with a small molecule inhibitor of MLL1 (MI-2) yielded a significant reduction in CD4+ Th17 cells and IL-17A. This is the first study to our knowledge to identify the MLL1-mediated mechanisms responsible for regulating the Th17/Treg balance in normal and diabetic wounds and to define the complex role of Notch signaling in CD4+ T cells in wounds, where increased or decreased Notch signaling both result in pathologic wound repair. Therapeutic targeting of MLL1 in diabetic CD4+ Th cells may decrease pathologic inflammation through regulation of CD4+ T cell differentiation.

Tumor CD20 Loss and Poor Outcome in Relapsed/Refractory (R/R) Diffuse Large B-Cell Lymphoma (DLBCL)

Tumor CD20 Loss and Poor Outcome in Relapsed/Refractory (R/R) Diffuse Large B-Cell Lymphoma (DLBCL)

ABCL-119 Tumor CD20 Loss and Poor Outcome in Relapsed/Refractory (R/R) Diffuse Large B-Cell Lymphoma (DLBCL)

ABCL-119 Tumor CD20 Loss and Poor Outcome in Relapsed/Refractory (R/R) Diffuse Large B-Cell Lymphoma (DLBCL)

Metabolism of immune cells in septic shock in children

In septic shock the hyperimmune response and immunosuppression can occur at the same time, being defined as mixed antagonist response syndrome. Under standard conditions, mitochondria supports cellular energy metabolism through the citric cycle and the phosphorylation chain. Both NK and T cells produce an immediate effector response to inflammatory signals using the rapid production of energy in aerobic glycolysis (glucose-lactate]. When the inflammatory signal is eliminated, they return to metabolism in the Krebs cycle. Septic shock in the children was accompanied by immunoparalysis (TNFα &lt;200pg/ml), lower monocyte human antigen (HLA-DR), loss of peripheral non-naive CD4 T cells and low mitochondrial respiration. Children with septic shock had a in hospital mortality rate of 10.8-33.5%, high risk of hospitali­zation in the next 28 days and in the following months with predominantly respiratory infections. The treatment of mitochondrial dysfunction, insufficiently structured, is based on antioxidant medication, but the results await confirmation.

CD19 expression in diffuse large B-cell lymphoma (DLBCL) patient biopsies after treatment with tafasitamab.

e19050 Background: Tafasitamab (tafa) is a CD19-targeting immunotherapy that induces enhanced antibody dependent cellular cytotoxicity and phagocytosis. Tafa received accelerated FDA approval in 2020 for treating relapsed or refractory DLBCL in combination with lenalidomide in patients ineligible for autologous stem cell transplant, based on results from L-MIND, a multicenter, open-label, single-arm, phase 2 trial. FirmMIND (NCT05429268) is an ongoing identical study, further evaluating efficacy and safety of tafa in this setting. Several other CD19 targeting agents are available; however, CD19 antigen loss following treatment with chimeric antigen receptor T-cell therapy targeting CD19 (CART19) has raised concerns over CD19-targeted therapy sequencing. Loss of CD19 has not been observed to date in tafa clinical trials or practice. Methods: To evaluate CD19 expression at screening and after disease progression in firmMIND, we developed an immunohistochemistry (IHC) assay using a mAb that binds to an intracellular epitope of CD19 clone LE-CD19. To validate that CD19 expression is detectable after tafa treatment, we treated cultured SU-DHL-4 or Jurkat cells with tafa at a saturating concentration for 24 hours. After washing out unbound tafa, cells were harvested at specific timepoints. Resulting cell line blocks were assessed for CD19 expression by IHC. We also monitored CD19 trafficking in response to fluorescently labeled tafa by confocal microscopy. Results: By IHC, CD19 expression was comparable between treatment conditions, suggesting the assay can effectively measure CD19 expression in lymphoma biopsy samples post-tafa. Confocal microscopy showed a fraction of tafa-bound CD19 internalized and colocalized with a CD19 intracellular pool labeled with anti-CD19 mAb clone EPR5906; this mAb also detected CD19 that remained localized on the cell surface. The data demonstrated that CD19 persists on tumor cells after tafa treatment at levels that could be bound by other CD19-targeting therapies. As of the January 25, 2024 data cutoff, we received 6 end-of-treatment (EOT) samples from patients in firmMIND. Biopsies were collected 14-26 days after the last tafa dose; 5 of 6 samples were biopsies from lymph nodes or liver and showed ≥80% of tumor cells staining CD19+ on membranes with 3+ intensity by IHC, except 1 that was collected only 6 days after last dose and showed 50% positivity. The sixth biopsy (extranodal subcutaneous lesion) showed no CD19 expression at baseline or EOT. Conclusions: Patient biopsy analysis and preclinical data suggest that following tafa treatment, CD19 antigen is preserved on the surface of tumor cells after short and transient internalization and could be targeted by other CD19 agents, such as CART therapy. This conclusion is supported by sequential treatment with tafa followed by CART in vivo (Sakemura, 2024). Analyses are ongoing to confirm these findings.

PMB-CT01 (BAFFR-CAR T cell) therapy to examine preliminary safety and clinical responses in patients with B-cell malignancies who are ineligible for or failed CD19-directed therapy, including CD19-negative disease.

6534 Background: CD19-targeted therapies have shown remarkable efficacy in patients with B-cell malignancies. However, relapse with CD19 loss as a mechanism of tumor escape is common and represents a serious challenge. Patients who are ineligible for or have failed prior CD19-directed therapy have limited salvage options and a very poor prognosis. The B-cell activating factor receptor (BAFF-R) is functionally expressed in B-cell acute lymphoblastic leukemia (B-ALL) and non-Hodgkin lymphomas (NHL), including in patients with CD19-negative relapse (Qin et al., Sci Transl Med. 2019). As a critical regulator of B-cell function and survival, BAFF-R may be less prone to downregulation by tumors as a mechanism of antigen escape. Methods: We are conducting two phase 1 clinical trials of BAFFR-CAR T cells in patients with relapsed/refractory (r/r) B-ALL (NCT04690595) and NHL (NCT05370430). Primary endpoints are dose limiting toxicities (DLTs) and all other toxicities. Secondary endpoints include response rate, minimal residual disease (MRD) negative rate, PFS and OS. Response is evaluated using European LeukemiaNet criteria for B-ALL and Lugano 2014 for NHL. Results: As of Feb. 1st, 2024, 1 B-ALL and 3 NHL patients have completed treatment and post-treatment evaluations. Each received 50M BAFFR CAR T cells (starting dose level in both trials). The B-ALL patient had CD19/CD20/CD22-negative disease, and had received prior blinatumomab. NHL patients #1 and #2 both had CD19-expressing mantle cell lymphoma (MCL) and had received prior CD19 CAR T cells. Patient #2 had also received a CD3/CD20 bispecific antibody. NHL patient #3 had CD19/CD20-negative T cell/histiocyte-rich large B-cell lymphoma (THRBCL) and had not received prior CD19 CAR T cells. There were no DLTs, all patients had Gr. 1 cytokine release syndrome, and 2/3 NHL patients had Gr. 1 immune effector cell-associated neurotoxicity syndrome. Robust CAR T cell expansion was seen in all patients with a peak on days 12-14 post-infusion. The B-ALL and the 2 MCL patients reached MRD-negative complete response (CR) at 1 month post-treatment. The THRBCL patient had partial response (PR) at 1 month that converted to CR at 3 months. The B-ALL patient successfully transitioned to allogeneic HCT while in CR at 3 months post-infusion. Responses are ongoing in all NHL patients at 14, 10, and 8 months for patients #1, #2 and #3, respectively. Additional patients have been enrolled at the next dose level (200M CAR T cells). Results for these patients will be presented at the meeting. Conclusions: With a 100% CR rate at 3 months in the first 4 patients and durable responses at dose level 1, BAFFR-CAR T cells are a promising therapeutic option for patients with r/r B-cell malignancies who are ineligible or failed prior CD19-targeted therapy, including with CD19 antigen loss. Clinical trial information: NCT04690595 , NCT05370430 .