Therapy For Hematologic Malignancies Research Articles

PARP inhibition leads to synthetic lethality with key splicing-factor mutations in myelodysplastic syndromes.

Splicing factors are frequently mutated in patients with myelodysplastic syndromes and acute myeloid leukaemia. Recent studies have revealed convergent molecular defects caused by splicing factor mutations, among which R-loop dysregulation and resultant genome instability are suggested as contributing factors to disease progression. On the other hand, understanding how mutant cells survive upon aberrant R-loop formation and genome instability is essential for developing novel therapeutics. The immunoprecipitation was performed to identify R-loops in association with PARP1/poly-ADP-ribosylation. The western blot, immunofluorescence, and flow cytometry assays were used to test the cell viability, cell cycle arrest, apoptosis, and ATM activation in mutant cells following the treatment of the PARP inhibitor. The Srsf2(P95H) knock-in murine hematopoietic cells and MLL-AF9 transformed leukaemia model were generated to investigate the potential of the PARP inhibitor as a therapy for haematological malignancies. The disease-causing mutations in SRSF2 activate PARP and elevate the overall poly-ADP-ribosylation levels of proteins in response to R-loop dysregulation. In accordance, mutant cells are more vulnerable to the PARP inhibitors in comparison to the wild-type counterpart. Notably, the synthetic lethality was further validated in the Srsf2(P95H) knock-in murine hematopoietic cell and MLL-AF9 leukaemia model. Our findings suggest that mutant cells antagonise the genome threat caused by R-loop disruption by PARP activation, thus making PARP targeting a promising therapeutic strategy for myeloid cancers with mutations in SRSF2.

Identification of novel GSPT1 degraders by virtual screening and bioassay

GSPT1 plays crucial physiological functions, such as terminating protein translation, overexpressed in various tumors. It is a promising anti-tumor target, but is also considered as an “undruggable” protein. Recent studies have found that a class of small molecules can degrade GSPT1 through the “molecular glue” mechanism with strong antitumor activity, which is expected to become a new therapy for hematological malignancies. Currently available GSPT1 degraders are mostly derived from the scaffold of immunomodulatory imide drug (IMiD), thus more active compounds with novel structure remain to be found. In this work, using computer-assisted multi-round virtual screening and bioassay, we identified a non-IMiD acylhydrazone compound, AN5782, which can reduce the protein level of GPST1 and obviously inhibit the proliferation of tumor cells. Some analogs were obtained by a substructure search of AN5782. The structure-activity relationship analysis revealed possible interactions between these compounds and CRBN-GSPT1. Further biological mechanistic studies showed that AN5777 decreased GSPT1 remarkably through the ubiquitin-proteasome system, and its effective cytotoxicity was CRBN- and GSPT1-dependent. Furthermore, AN5777 displayed good antiproliferative activities against U937 and OCI-AML-2 cells, and dose-dependently induced G1 phase arrest and apoptosis. The structure found in this work could be good start for antitumor drug development.

Phase 1 study of CAR-37 T cells in patients with relapsed or refractory CD37+ lymphoid malignancies

We report on the first-in-human clinical trial using chimeric antigen receptor (CAR) T-cells targeting CD37, an antigen highly expressed in B- and T-cell malignancies (clinicaltrials.gov NCT04136275). Five patients with relapsed or refractory CD37+ lymphoid malignancies were enrolled and infused with autologous CAR-37 T-cells. CAR-37 T-cells expanded in the peripheral blood of all patients and, at peak, comprised >94% of the total lymphocytes in 4/5 patients. Tumor responses were observed in 4/5 patients, with 3 complete responses, 1 mixed response, and 1 patient whose disease progressed rapidly and with relative loss of CD37 expression. Three patients experienced prolonged and severe pancytopenia, and in two of these patients, efforts to ablate CAR-37 T-cells (which were engineered to co-express truncated EGFR) with cetuximab, were unsuccessful. Hematopoiesis was restored in these two patients following allogeneic hematopoietic stem cell transplantation. No other severe, non-hematopoietic toxicities occurred. We investigated the mechanisms of profound pancytopenia and did not observe activation of CAR-37 T-cells in response to hematopoietic stem cells in vitro or hematotoxicity in humanized models. Patients with pancytopenia had sustained high levels of IL-18, with low levels of IL-18 binding protein in their peripheral blood. IL-18 levels were significantly higher in CAR-37-treated patients relative to both cytopenic and non-cytopenic cohorts of CAR-19-treated cohorts of patients. In conclusion, CAR-37 T-cells exhibited anti-tumor activity, with significant CAR expansion and cytokine production. CAR-37 T-cells may be an effective therapy in hematologic malignancies as a bridge to hematopoietic stem cell transplant.

Efficacy and safety of chimeric antigen receptor T-cell (CAR-T) therapy in hematologic malignancies: a living systematic review (protocol)

Objective To determine the efficacy and safety of CAR-T therapy in the treatment of patients with hematologic malignancies, in comparison with other current therapies. Design A living systematic review Methods We will include randomized trials evaluating the effect of CAR-T therapy versus other active treatments, hematopoietic stem cell transplantation, best supportive care or any other intervention in patients with hematologic malignancies. Non-randomized primary studies will be searched in case we found no direct evidence from randomized controlled trials. Two reviewers will independently screen each study for eligibility, extract data, and assess the risk of bias. Efficacy measures will include overall survival rate, overall response rate, complete response/remission (CR) rate, partial response/remission (PR) rate, relapse from CR, progression-free survival, and time from CAR-T infusion to transplantation. Safety measures will include serious adverse events, the incidence of cytokine release syndrome, graft-versus-host disease, neurotoxicity, and total adverse events. Quality of life will also be assessed. Meta-analyses will be carried out to summarize the results. We will apply the GRADE approach to assess the certainty of the evidence for each outcome. A living, web-based version of this review will be openly available until there is solid evidence to respond to the review objective. We will resubmit it for publication every time the conclusions change or whenever there are substantial updates.

Current progress in chimeric antigen receptor T‐cell therapy for malignant tumors

AbstractIn the realm of malignant tumor treatment, particularly regarding hematologic malignancies, chimeric antigen receptor T‐cell (CAR‐T) immunotherapy has witnessed remarkable advancements in recent years. This approach involves genetically modifying and engineering a patient's T‐cells ex vivo to express a specific CAR, known as CAR‐T cells. When these modified cells are reintroduced into the patient, they can specifically recognize target antigens and exhibit highly efficient cytotoxicity against cells expressing these antigens, making them suitable for the treatment of malignant tumors. CD19, which is expressed on the surface of B lymphocytes at different stages of differentiation, has been identified as a suitable target for the treatment of most B‐cell lymphomas. CAR‐T cells targeting CD19 have demonstrated excellent specificity, cytotoxicity, and persistence in both in vitro and clinical trials, showing tremendous potential for application. However, identifying appropriate targets for CAR‐T therapy in solid tumors remains a challenge, leading to limited advancements in this area. This review discusses the mechanisms, applications, limitations, and prospects of CAR‐T therapy in hematologic malignancies and solid tumors, aiming to provide directions for future research in this field.

Osteosarcoma Arising as a Secondary Malignancy following Treatment for Hematologic Cancer: A Report of 33 Affected Patients from the Cooperative Osteosarcoma Study Group (COSS).

Osteosarcoma may arise as a secondary cancer following leukemias or lymphomas. We intended to increase the knowledge about such rare events. We searched the Cooperative Osteosarcoma Study Group's database for individuals who developed their osteosarcoma following a previous hematological malignancy. The presentation and treatment of both malignancies was investigated, and additional neoplasms were noted. Outcomes after osteosarcoma were analyzed and potential prognostic factors were searched for. A total of 33 eligible patients were identified (male: 23, female: 10; median age: 12.9 years at diagnosis of hematological cancer; 20 lymphomas, 13 leukemias). A cancer predisposition syndrome was evident in one patient only. The hematological cancers had been treated by radiotherapy in 28 (1 unknown) and chemotherapy in 26 cases, including bone-marrow transplantation in 9. The secondary bone sarcomas (high-grade central 27, periosteal 2, extra-osseous 2, undifferentiated pleomorphic sarcoma of bone 2) arose after a median lag-time of 9.4 years, when patients were a median of 19.1 years old. Tumors were considered radiation-related in 26 cases (1 unknown). Osteosarcoma-sites were in the extremities (19), trunk (12), or head and neck (2). Metastases at diagnosis affected eight patients. Information on osteosarcoma therapy was available for 31 cases. All of these received chemotherapy. Local therapy involved surgery in 27 patients, with a good response reported for 9/18 eligible patients. Local radiotherapy was given to three patients. The median follow-up was 3.9 (0.3-12.0) years after bone tumor diagnosis. During this period, 21 patients had developed events as defined, and 15 had died, resulting in 5-year event-free and overall survival rates of 40% (standard error: 9%) and 56% (10%), respectively. There were multiple instances of additional neoplasms. Several factors were found to be of prognostic value (p < 0.05) for event-free (osteosarcoma site in the extremities) or overall (achievement of a surgical osteosarcoma-remission, receiving chemotherapy for the hematologic malignancy) survival. We were able to prove radiation therapy for hematological malignancies to be the predominant risk factor for later osteosarcomas. A resulting overrepresentation of axial and a tendency towards additional neoplasms affects prognosis. Still, selected patients may become long-term survivors with appropriate therapies, which is an argument against therapeutic negligence.

Emerging Innate Immune Cells in Cancer Immunotherapy: Promises and Challenges.

Immune checkpoint inhibitor (ICI)-based therapy has made an unprecedented impact on survival benefit for a subset of cancer patients; however, only a subset of cancer patients is benefiting from ICI therapy if all cancer types are considered. With the advanced understanding of interactions of immune effector cell types and tumors, cell-based therapies are emerging as alternatives to patients who could not benefit from ICI therapy. Pioneering work of chimeric antigen receptor T (CAR-T) therapy for hematological malignancies has brought encouragement to a broad range of development for cellular-based cancer immunotherapy, both innate immune cell-based therapies and T-cell-based therapies. Innate immune cells are important cell types due to their rapid response, versatile function, superior safety profiles being demonstrated in early clinical development, and being able to utilize multiple allogeneic cell sources. Efforts on engineering innate immune cells and exploring their therapeutic potential are rapidly emerging. Some of the therapies, such as CD19 CAR natural killer (CAR-NK) cell-based therapy, have demonstrated comparable early efficacy with CD19 CAR-T cells. These studies underscore the significance of developing innate immune cells for cancer therapy. In this review, we focus on the current development of emerging NK cells, γδ T cells, and macrophages. We also present our views on potential challenges and perspectives to overcome these challenges.

The effect of antifungal treatments on bronchoalveolar lavage galactomannan for the diagnosis of invasive fungal infections in patients with hematological malignancy: Single-center experience

Background Invasive fungal infections (IFI) mainly caused by aspergillus species are one of the leading causes of death in patients undergoing immunosuppressive therapy for hematological malignancies. The presence of galactomannan in bronchoalveolar lavage fluid (BAL GM) is an important diagnostic marker. Some of the factors affecting the BAL GM are still unknown. Antifungal treatment administered before or after BAL also affect BAL GM results. Methods To investigate the effect of BAL GM timing on the diagnosis of IFE, 100 patients receiving immunosuppressive therapy with hematological malignancy at the Uludağ University Faculty of Medicine, Hematology Department, and underwent BAL over a 3-year period with the suspicion of IFE as well as 127 BAL procedures of these patients were examined. Results There were 70 patients who started antifungal therapy before BAL and 30 patients who did not. BAL GM was found positive in 33 (47.1%) of the 70 patients who received antifungal therapy compared to 22 (73.3%) of the 30 patients who did not receive antifungal therapy. There was a significant difference between the two groups in terms of BAL GM positivity (p = 0.016). Subsequently, 127 BAL procedures of these 100 patients were evaluated. When the second, third, and subsequent BAL procedures of the same patients were included in the study, BAL GM was positive in 41 (46.6%) of the 88 procedures in patients who received treatment before BAL and in 25 (64.1%) of the 39 procedures in patients who did not receive treatment before BAL. The rate of BAL GM positivity did not differ between groups (p = 0.068). Conclusions The balance between reducing the risk by initiating early antifungal therapy and maximizing the diagnostic value of BAL GM should be evaluated individually for each patient.

Chimeric Antigen Receptor (CAR) T-Cell Therapy in Hematologic Malignancies: Clinical Implications and Limitations.

Chimeric antigen receptor (CAR) T-cell therapy has become a powerful treatment option in B-cell and plasma cell malignancies, and many patients have benefited from its use. To date, six CAR T-cell products have been approved by the FDA and EMA, and many more are being developed and investigated in clinical trials. The whole field of adoptive cell transfer has experienced an unbelievable development process, and we are now at the edge of a new era of immune therapies that will have its impact beyond hematologic malignancies. Areas of interest are, e.g., solid oncology, autoimmune diseases, infectious diseases, and others. Although much has been achieved so far, there is still a huge effort needed to overcome significant challenges and difficulties. We are witnessing a rapid expansion of knowledge, induced by new biomedical technologies and CAR designs. The era of CAR T-cell therapy has just begun, and new products will widen the therapeutic landscape in the future. This review provides a comprehensive overview of the clinical applications of CAR T-cells, focusing on the approved products and emphasizing their benefits but also indicating limitations and challenges.

2022–2023 Drug Updates in Hematologic Malignancies

At JADPRO Live 2023 in Orlando, Rebecca L. Rezac, PharmD, BCOP, summarized key information on US Food and Drug Administration approvals from late 2022 to late 2023. Dr. Rezac described indications, mechanisms of action, and monitoring and management of side effects of new therapies for hematologic malignancies.

Tocilizumab administration in cytokine release syndrome is associated with hypofibrinogenemia after chimeric antigen receptor T-cell therapy for hematologic malignancies.

Chimeric antigen receptor (CAR-) T cell therapy causes serious side effects including cytokine release syndrome (CRS). CRS-related coagulopathy is associated with hypofibrinogenemia that is thus far considered the result of disseminated intravascular coagulation (DIC) and liver dysfunction. We investigated incidence and risk factors for hypofibrinogenemia in 41 consecutive adult patients with hematologic malignancies (median age 69 years, range 38-83 years) receiving CAR-T cell therapy between 01/2020 and 05/2023 at the University Medical Center Regensburg. CRS occurred in 93% of patients and was accompanied by hypofibrinogenemia already from CRS grade 1. Yet, DIC and liver dysfunction mainly occurred in severe CRS (≥ grade 3). After an initial increase during CRS, fibrinogen levels dropped after administration of tocilizumab in a dose dependent manner (r = -0.44, p = 0.004). In contrast, patients who did not receive tocilizumab had increased fibrinogen levels. Logistic regression analysis identified tocilizumab as an independent risk factor for hypofibrinogenemia (odds ratio = 486, p < 0.001). We thus hypothesize that fibrinogen synthesis in CRS is upregulated in an interleukin-6-dependent acute phase reaction compensating for CRS-induced consumption of coagulation factors. Tocilizumab inhibits fibrinogen upregulation resulting in prolonged hypofibrinogenemia. These observations provide novel insights into the pathophysiology of hypofibrinogenemia following CAR-T cell therapy and emphasize the need for close fibrinogen monitoring after tocilizumab treatment of CRS.

Abstract 5092: Development of a priming approach to enhance antigen-directed therapies in hematological malignancies and solid tumors

Abstract Clinical success of antigen-directed therapies in cancer has been limited by heterogeneous antigen expression and a narrow therapeutic window. In many patients, failure of antigen-directed therapies such as CAR T-cell therapies, antibody-drug conjugates (ADC) or radioligand therapies (RLT) has less to do with a lack of potency or specificity of the agent and more to do with variable and suboptimal antigen expression in the patient. We are developing combination therapies that introduce a priming agent designed to optimize antigen expression in the patient and improve the therapeutic window of existing antigen-directed therapies. Our high-throughput platform utilizes machine learning and automation to scale functional combinatorial screening for small molecule priming agents using cell lines and patient samples. Using this platform, we are identifying priming therapies for CD123, CD33, CD22, and CD19 in hematological malignancies including acute myeloid leukemia (AML), and HER2, HER3, and TROP2 in solid tumors including breast cancer.For our CD123 program, we have identified priming agents that selectively upregulate CD123 expression ≥2-fold on AML cells without increasing expression on normal myeloid progenitor cells, which also express CD123. AML cells treated with a CD123 priming agent in combination with a CD123 ADC showed greater tumor-cell killing compared to the ADC alone. In comparison, normal myeloid progenitor cells from healthy bone marrow treated with the same CD123 priming-ADC combination treatment did not show an increase in ADC-mediated cell killing. From our mechanistic studies, we have identified a somatic mutation associated with CD123 upregulation that may lead to selective upregulation on cancer cells and not on normal cells. Additionally, this marker can be used for patient selection. We have also observed CD123 upregulation in vivo using mouse models of AML and are currently conducting additional in vivo proof of concept studies to demonstrate that CD123 priming therapy enhances efficacy.There are two distinct advantages of a priming approach paired with an antigen-directed therapy, which include: (1) improving the durability of patient responses and (2) expanding the patient population. Our priming combination approach has the potential to overcome antigen heterogeneity and thereby improve the clinical success of antigen-directed therapies. Citation Format: Kathryn E. Vanderlaag, Anukriti Dhar, Molly Volkman, Erik Werner, Transon Nguyen, Kamran Ali. Development of a priming approach to enhance antigen-directed therapies in hematological malignancies and solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5092.

Abstract 1096: Enhancing effector function in T cell adoptive cell therapy in murine models of pediatric sarcomas

Abstract New treatment approaches for high-risk pediatric sarcomas are desperately needed. Immunotherapy advances with adoptive cell therapy in hematologic malignancies have not yet translated to pediatric solid tumors including sarcomas. Hallmarks of the immune microenvironment in pediatric sarcomas include the presence of immunosuppressive cytokines and immune cells and a lack of potent T cell activation. We are seeking ways to overcome these barriers to effective adoptive cell therapy in pediatric sarcoma. We are exploring introduction of a novel synthetic co-receptor, CD8a:MyD88, that can augment tumor-specific T cell effector function and potentially improve outcomes in murine models of sarcoma. Tumor-antigen specific T cells or tumor-infiltrating T cell populations are transduced ex vivo to express the CD8a:MyD88 synthetic co-receptor resulting in MyD88 signaling when T cells encounter their cognate MHC I/epitope complex. We have studied the effects of CD8a:MyD88 expression on cytokine production and lytic activity against murine sarcoma cell lines in vitro. Initial experiments utilized the OT1 T cell model (specific to the model antigen chicken ovalbumin) which we introduced into our murine sarcoma cell lines (KPSarc pleomorphic sarcoma and M3-9M rhabdomyosarcoma). Using GFP as a marker of T cell transduction, the production of effector cytokine interferon gamma was measured following T cell activation compared with GFP-negative non-transduced T cells. CD8a:MyD88 expressing T cells expressed higher levels of interferon gamma at basal levels and following antigen-specific activation when co-cultured with Ova-expressing sarcoma cells. Improved tumor-specific lytic function in CD8a:MyD88 expressing T cells was also observed in co-culture experiments compared to untransduced or control vector- transduced T cells. We further hypothesized that skewing T cell differentiation toward an effector phenotype with the addition of interleukin-12 during T cell expansion would increase the anti-tumor effect of the CD8a:MyD88 expressing T cells. Co-culture with our murine sarcoma cell lines showed increased tumor-lytic function in T cells primed with IL-12. We have ongoing experiments studying the in vivo effects of the CD8a:MyD88 expressing T cells in model antigen-expressing murine sarcoma models. In addition, the ability of the CD8a:MyD88 co-receptor to augment the effector function of polyclonal sarcoma infiltrating T cells is being explored. These results will further define the obstacles and potential solutions to successful adoptive T cell therapy for pediatric sarcomas. Citation Format: Jessica P. Sanchez, Eduardo Davila, Brian H. Ladle. Enhancing effector function in T cell adoptive cell therapy in murine models of pediatric sarcomas [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1096.

Abstract 44: A novel, codon-based method to modulate off-tumor toxicity of chimeric antigen receptor (CAR) T cells

Abstract Chimeric antigen receptor (CAR) T cell therapies for hematologic malignancies, while promising and efficacious, can also result in a variety of adverse effects. Of note, on-target, off-tumor toxicity is one adverse effect that continues to limit the safety and efficacy of CAR-T cells and has even led to life-threatening or fatal outcomes in patients (Morgan et al. 2010.). To address this, we have developed a unique, codon-based approach by replacing strategically selected codons within CAR non-antigen binding domains with their rare synonymous isoforms. Hoekema et al. (1987) reported the ability of synonymous rare codons to alter gene and protein expression in other model systems. In CAR-T cells, we aim to use this novel, codon-based approach to fine tune and modulate CAR-T cell expression and function to elicit full on-tumor activity while lessening off-tumor toxicity. In our approach, we replaced specific endogenous codons within the non-antigen binding domains of Hu8F4-CAR, a TCR-like 2nd generation CAR developed by our group which targets the leukemia-associated antigen PR1/HLA-A2 (Ma et al. 2016.). Briefly, we created variants of Hu8F4-CAR containing rare codon substitutions in two subtypes: intracellular-based (type 1) and combined extracellular- and intracellular-based (type 2). Each variant subtype (type 1 or 2) had either between 6 and 30 or 27 and 57 rare codons introduced, respectively. Expression profiling and functional assessment of the Hu8F4-CAR variants was assessed in the Jurkat mutant cell line (J.RT3-T3.5) or human primary healthy peripheral blood mononuclear cells (PBMCs) that were enriched for T cells, respectively. Expression profiling results show that surface expression is reduced in J.RT3-T3.5 cells when assessed by flow cytometry by 1-1.5-fold compared to unmodified Hu8F4-CAR. When evaluated by immunoblotting, novel lower molecular weight bands emerge only in rare codon-modified variants, suggesting that rare codons may have altered translational or post-translational processing leading to truncated CAR protein isoforms (30, 45, or 60kDa vs 73kDa for unmodified Hu8F4-CAR), which may contribute to reduced cell surface CAR expression. Furthermore, we have conducted in vitro functional testing of the rare codon-modified variants and show reduced activity against non-malignant off-tumor targets that express PR1 and/or HLA-A2 including healthy donor PBMCs, mobilized hematopoietic cells, and bone marrow. Importantly, in addition to reduced off-tumor activity against healthy myeloid cells, rare codon-modified variants preserve their activity against on-tumor target cells, including the U937-A2+ cell line and primary acute myeloid leukemia. Altogether, these results suggest that our approach can modulate CAR expression and functionality creating a novel methodology to reduce the likelihood of off-tumor adverse effects typically associated with CAR therapies. Citation Format: Rolando A. Vedia, Sijie Lu, Anne V. Philips, Anna Sergeeva, Lisa S. St John, Karen Clise-Dwyer, Gheath Alatrash, Jeffrey J. Molldrem. A novel, codon-based method to modulate off-tumor toxicity of chimeric antigen receptor (CAR) T cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 44.

Abstract 5230: A modified FcγRI expressing T cell (SolidT) exhibits profound anti-tumor activity followed by maturation into effector memory t cells that can be reactivated to kill a secondary tumor

Abstract We previously described a subset of cytotoxic FcγRI (CD64)-expressing T cells in tumors and inflamed tissues (PMID 31449054). We subsequently expressed an engineered modified CD64 into T cells (“SolidT”) and demonstrated that these cells possessed greater specificity for tumor over normal tissues, reduced exhaustion, and attenuated cytokine secretion compared to other engineered cell therapies. SolidT activation correlates with surface tumor antigen density enabling the differentiation between tumor cells and normal cells by use of a specific antibody targeting tumor antigens. We previously showed that SolidT + mAb has anti-tumor activity in vitro and in vivo in several tumor models, including immunodeficient HER2 xenografts and an immunocompetent syngeneic B16-TYRP1 model (PMID 37070661). In this study, we further assessed the effect of SolidT on HER2+ tumors in a rechallenge model and extended the platform to CD20+ lymphoma models by combining SolidT with rituximab. In an in vivo experiment using HER2+, NCI-N87 cells in immunodeficient mice, several mice in the SolidT + trastuzumab group experienced complete tumor clearance. These mice were monitored until day 84, then rechallenged with NCI-N87 cells along with a new control group. After tumor establishment, mice were treated only with trastuzumab weekly for two weeks. While tumors in the control group progressed, the tumors in the rechallenged mice were completely cleared with a trastuzumab boost only, suggesting a SolidT cell memory response. Bone marrow from these rechallenged mice was analyzed and revealed the presence of effector memory SolidT cells. The SolidT model was then studied in a human, CD20+ lymphoma model in vitro and in vivo in immunodeficient mice. SolidT + rituximab was more potent in-vivo against CD20+ lymphoma cells than rituximab. In vivo, mice treated with SolidT + rituximab had significantly greater survival, lower tumor burden, and fewer sites of metastases than mice treated with rituximab alone. Bone marrow from these mice was analyzed and revealed the presence of effector memory SolidT cells in mice treated with SolidT + rituximab. We demonstrated that marrow resident SolidT cells developed an effector memory phenotype that positively correlated with outcome in HER2+ and CD20+ xenograft models, consistent with earlier data from a syngeneic B16 melanoma model. We further demonstrated that SolidT cells can be reactivated in a rechallenge model with only the addition of tumor targeting mAb, resulting in the eradication of the recurring tumors. This data further confirms that the SolidT platform can be targeted against different tumor antigens by using different tumor targeting mAbs. The SolidT platform could represent a paradigm shift in the approach to engineered cellular therapy for hematological malignancies and solid tumors. Citation Format: Shahar Dotan, Noam Pilpel, Peleg Rider, Hana Shpilt, Diana Rasoulouniriana, Yaron Carmi, James E. Wooldridge. A modified FcγRI expressing T cell (SolidT) exhibits profound anti-tumor activity followed by maturation into effector memory t cells that can be reactivated to kill a secondary tumor [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5230.

Inflammation as a driver of hematological malignancies.

Hematopoiesis is a tightly regulated process that produces all adult blood cells and immune cells from multipotent hematopoietic stem cells (HSCs). HSCs usually remain quiescent, and in the presence of external stimuli like infection or inflammation, they undergo division and differentiation as a compensatory mechanism. Normal hematopoiesis is impacted by systemic inflammation, which causes HSCs to transition from quiescence to emergency myelopoiesis. At the molecular level, inflammatory cytokine signaling molecules such as tumor necrosis factor (TNF), interferons, interleukins, and toll-like receptors can all cause HSCs to multiply directly. These cytokines actively encourage HSC activation, proliferation, and differentiation during inflammation, which results in the generation and activation of immune cells required to combat acute injury. The bone marrow niche provides numerous soluble and stromal cell signals, which are essential for maintaining normal homeostasis and output of the bone marrow cells. Inflammatory signals also impact this bone marrow microenvironment called the HSC niche to regulate the inflammatory-induced hematopoiesis. Continuous pro-inflammatory cytokine and chemokine activation can have detrimental effects on the hematopoietic system, which can lead to cancer development, HSC depletion, and bone marrow failure. Reactive oxygen species (ROS), which damage DNA and ultimately lead to the transformation of HSCs into cancerous cells, are produced due to chronic inflammation. The biological elements of the HSC niche produce pro-inflammatory cytokines that cause clonal growth and the development of leukemic stem cells (LSCs) in hematological malignancies. The processes underlying how inflammation affects hematological malignancies are still not fully understood. In this review, we emphasize the effects of inflammation on normal hematopoiesis, the part it plays in the development and progression of hematological malignancies, and potential therapeutic applications for targeting these pathways for therapy in hematological malignancies.

The Effect of Retinoic Acid on Arsenite-Transformed Malignant UROtsa Bladder Cancer Cells: In Vitro Model of Basal Muscle-Invasive Bladder Cancer.

Bladder cancer (BC) is the eighth most common cause of cancer death in the United States of America. BC is classified into non-muscle-invasive bladder cancer (NMIBC) and muscle-invasive bladder cancer (MIBC). Genetically, MIBCs are categorized into the more aggressive basal subtype or less aggressive luminal subtype. All-trans retinoic acid (tretinoin), the ligand for the RAR-RXR retinoic acid receptor, is clinically used as a differentiation therapy in hematological malignancies. This study aims to determine the effects of retinoic acid on arsenite-transformed malignant urothelial cells (UROtsa As), serving as a model for basal muscle-invasive bladder cancer. We treated three independent isolates of arsenite-transformed malignant human urothelial UROtsa cells (UROtsa As) with tretinoin for 48 h. Cell viability, proliferation, and apoptosis were analyzed using crystal violet staining and flow cytometry. mRNA and protein level analyses were performed using RT-qPCR and the Simple Western™ platform, respectively. Tretinoin was found to reduce cell proliferation and urosphere formation, as well as decrease the expression of basal markers (KRT1, KRT5, KRT6, EGFR) and increase the expression of luminal differentiation markers (GATA3, FOXA1). Mechanistically, the antiproliferative effect of tretinoin was attributed to the downregulation of c-myc. Our results suggest that targeting the retinoic acid pathway can diminish the aggressive behavior of basal muscle-invasive urothelial cancer and may enhance patient survival.

Efficacy and safety of cladribine in combination with busulfan and cyclophosphamide as an intensive conditioning regimen preceding allogeneic hematopoietic stem cell transplantation in relapsed or refractory acute myeloid leukemia

BackgroundCladribine, an analogue of deoxyadenosine, is used for therapy of hematological malignancies. Cladribine-containing regimen has been recommended as a rescue therapy for relapsed or refractory (R/R) acute myeloid leukemia (AML). Its combination with busulfan plus cyclophosphamide (BuCy), as an intensive conditioning regimen prior to allogeneic hematopoietic stem cell transplantation (allo-HSCT), requires more clinical evidence. This study aimed to explore the efficacy and safety of cladribine plus BuCy administered as an intensive conditioning regimen before allo-HSCT in R/R AML patients. MethodsTwenty-three R/R AML patients, who underwent cladribine plus BuCy intensive conditioning regimen before allo-HSCT, were retrospectively analyzed. The median (range) follow-up duration time of observation was 0.73 (0.08–2.69) years. ResultsThe median (range) returned levels of mononuclear cells were 11.5 (6.1–18.5) x 108/kg and CD34+ cells were 5.5 (3.5–9.3) x 106/kg. The median (range) time of platelet reconstitution was 13.0 (9.0–21.0) days and neutrophil reconstitution was 14.0 (11.0–26.0) days. The incidence of conditioning regimen related toxicity (CRRT) affected 69.6% of patients; all CRRT-affected patients had grade I-II symptoms, including gastrointestinal tract (39.1%), oral cavity (26.1%), liver (8.7%), and kidney (4.3%) CRRTs. The incidence of acute graft-versus-host disease (GVDH) included 30.4% among all patients with 4.3% of grade III-IV acute GVHD, and 34.8% of chronic GVHD. During the follow-up period, 4 (17.4%) patients relapsed, and 6 (26.1%) patients died (cause of death: disease relapse, n = 3; infection, n = 2; GVHD, n = 1). The 1-year and 2-year accumulating event-free survival rates were 66.3% and 53.1%, respectively. The 1-year accumulating overall survival rate was 74.7% and 2-year survival rate was 64.0%. ConclusionCladribine plus BuCy intensive conditioning regimen before allo-HSCT exhibits favorable treatment efficacy with acceptable toxicity in R/R AML patients.

Gamma Delta T Cells: A Prospective Analysis of Their Regeneration Kinetics and Their Impact on the Clinical Outcome Following Allogeneic Hematopoietic Stem Cell Transplantation

Haematopoietic stem cell transplantation (HSCT) plays an important role in the therapy of hematological malignancies. Gamma delta T cells (ᵞᵟ T cells) are a distinct lineage of T lymphocytes that might play an important role in immune recovery and could utilize a graft –versus –leukemia effect post HSCT, furthermore, higher counts of ᵞᵟ T cells could improve clinical outcome after HSCT. This work was conducted to analyze the kinetics of gamma delta T cells recovery post HSCT, and to correlate their reconstitution with different factors that may influence the clinical outcome following HSCT. In this study, 22 consecutive allogeneic hematopoietic stem cell transplantation (HSCT) recipients were analysed during the first-year post transplantation by measuring the absolute count of CD3 T cells and percentages of gamma delta T cells subsets every month for each patient using flow cytometric technique. Statistics (means, minimal, and maximal values) were used to describe patient baseline characteristics. Results were presented as mean values of CD3+, gamma delta T cells %, and p-values. Higher gamma delta T cells percentages were significantly correlated with younger patient and donor age, sex matched transplantation, leukemic diseases, un-manipulated transplants, and in patients without chronic graft versus host disease complications. Furthermore, positive correlation between CD3 T cells counts and gamma delta T cells % was also determined. Overall survival and better clinical outcome following allogeneic HSCT could be related with proper gamma delta T cells reconstitution.

The Role of Cancer-Specific Target Antigens in CAR T Cell Therapy in Hematological Malignancies

The Role of Cancer-Specific Target Antigens in CAR T Cell Therapy in Hematological Malignancies