Hematologic Malignancy Cell Research Articles

Safety, Tolerability, Pharmacokinetics (PK) and Preliminary Antitumor Activity of the Cyclin-Dependent Kinase-9 (CDK9) Inhibitor AZD4573 in Relapsed/Refractory Hematological Malignancies: A Phase 1 First-in-Human Study

Introduction: CDK9, a serine/threonine kinase, regulates transcription elongation by phosphorylating RNA polymerase II at serine 2 (pSer2). Transient inhibition of CDK9 modulates expression of genes with short-lived transcripts and labile proteins, providing a potential therapeutic opportunity in tumors dependent upon oncogenes fitting these criteria. AZD4573 is a highly potent and selective CDK9 inhibitor that rapidly and preferentially decreases BCL2 family anti-apoptotic proteins MCL-1, BFL-1 as well as the oncogene MYC, inducing apoptosis in a broad range of human hematological malignant cell lines. AZD4573 monotherapy has shown antitumor effects associated with transient CDK9 inhibition in leukemia and lymphoma preclinical models. This multicenter, nonrandomized, open-label, phase 1 study (NCT03263637) was conducted to assess safety, tolerability, PK, pharmacodynamics (PD) and preliminary antitumor activity of AZD4573 in patients (pts) with relapsed or refractory hematological malignancies. Methods: The study comprised two parallel dose-escalation arms using a 3+3 design. Arm A included pts with diffuse large B-cell lymphoma (DLBCL), mantle cell lymphoma, or multiple myeloma. Arm B included pts with acute myeloid/lymphocytic leukemia, chronic myelomonocytic/lymphocytic leukemia, or hairy cell leukemia. Pts must have received at least 2 prior lines of therapy for the current histology. In each arm, pts received AZD4573 IV on an 8-week cycle using an intra-patient ramp up starting at 6 mg and increasing in 3 mg increments to a target dose of 18 mg (cohort 1) or 12 mg (cohort 2). The initial dosing regimen was 2 days on/12 days off but was changed to once weekly (QW) in cohort 1. Pts in Cohort 3 received AZD4573 9 mg QW on a 4-week cycle including intra-patient ramp up (3 mg, 6 mg, 9 mg). The primary objectives were to assess safety and to determine the biologically effective dose/maximum tolerated dose (MTD). Adverse events (AE) were assessed using CTCAE v4.03. Secondary/exploratory endpoints included PK, PD, and antitumor efficacy. Results: Overall, 44 pts were treated and evaluated for safety and efficacy: 22 each in Arms A and B. Median age was 69.0 years (range, 26.0 - 84.0), 65.9% were male, and median number of prior lines of therapy was 3 (range, 1-11). At data cutoff (Sept 30, 2021), all pts had discontinued treatment, mainly due to disease progression (47.7%) or AEs (22.7%). All pts had treatment-emergent adverse events (TEAEs) which were serious in 88.6%. The most common TEAEs were diarrhea (59.1%), pyrexia (52.3%) and nausea (50.0%). The most common TEAE that led to discontinuation was pneumonia (4.5%). Treatment-related AEs (TRAEs) occurred in 93.2% of pts and were serious in 59.1%, but none led to death. The most common TRAEs were diarrhea (50%), nausea (47.7%) and tumor lysis syndrome (TLS, 40.9%: laboratory TLS, 38.6%; clinical TLS, 6.8%) (Table 1). Elevated liver enzymes were transient and resolved spontaneously. These increases were assumed to be mainly due to down-modulation of hepatic transporter proteins and reduced enzyme clearance rather than direct hepatocellular cytotoxicity (based on clinical, in-vitro and in-silico assessments). Regarding MTD, the 18 mg dose was not tolerated in either arm (clinical TLS and acute kidney injury in Arm A; hypotension and liver injury in Arm B). AZD4573 12 mg was tolerated in Arm A, but not tolerated in Arm B due to clinical TLS (n=2). The recommended Phase 2 dose (RP2D) was 12 mg QW in Arm A and 9 mg QW in Arm B. AZD4573 showed dose-proportional increases in AUC and Cmax, with moderate PK variability (~30-60% CV), and a t1/2 of ~5 hrs. Furthermore, AZD4573 led to dose-dependent reduction in pSer2 (>75%) and MCL-1 (>70%) in peripheral blood as evidence of target engagement and mechanism of action. Median duration of exposure was 10.1 wks in Arm A (range, 1.0-130.1) and 5.2 wks in Arm B (range, 1.0-35.7). In Arm A, there was 1 complete and 1 partial response in 17 DLBCL pts. Median overall survival was not reached in Arm A and was 8.8 months in Arm B, with 7 (31.8%) deaths in each. Conclusion: AZD4573 had manageable safety and a PK profile suitable for QW dosing in a broad range of hematologic malignancies including lymphoma and leukemia. The reduction in pSer2 and MCL-1 supports the RP2D of 12 mg for lymphoma and 9 mg for leukemia. A combination study of AZD4573 and acalabrutinib (NCT04630756) is ongoing to further examine the DLBCL response signal seen in this study. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

A bibliometric and scientific knowledge-map study of the chimeric antigen receptor (CAR) natural killer (NK) cell-related research from 2010 to 2022.

ObjectivesAs emerging adoptive immunotherapy after CAR-T cell therapy, CAR-NK cell therapy has been developing rapidly in recent years. Presently, the research on CAR-NK cells has become a hotspot in the field of tumor immunotherapy.MethodsIn this descriptive study, CtieSpace and VOSviewer were used to perform the bibliometric and scientific knowledge-map analysis of articles and reviews related to CAR-NK cells.Results5371 authors from 715 institutions in 65 countries published 1028 papers about CAR-NK cells in 346 journals. The number of publications related to CAR-NK cells was increasing overall, especially from 2018 to 2021. The United States was in a leading position. The most active institution was Univ Texas, MD Anderson Cancer Center (USA). The journal with the most publications was Frontiers in immunology, and the most co-cited journal was Blood. The researcher with the most published papers was Winfried S. Wels, while the most co-cited researcher was Shannon L Maude. The research of CAR-NK cells in hematological malignancies and solid tumors (especially the selection of targets and the evaluation of efficacy and safety) was a research hotspot in this field. The emerging topics mainly included three aspects. First, further improve the proliferation and persistence of NK cells in vivo. Secondly, optimizing and improving the CAR structure for NK cells to improve the anti-tumor ability of CAR-NK cells. Thirdly, the related research of CRISPR/Cas9 gene-editing technology in constructing engineered immune cells.ConclusionIn this study, a bibliometric and scientific knowledge-map study provided a unique and objective perspective for the CAR-NK cell field. This information would provide a helpful reference for researchers interested in this field.

Functional Outcomes for Children, Adolescents, and Young Adults with Osteonecrosis following Hip Core Decompression.

Patients with pediatric leukemia and sickle cell disease are at risk for developing osteonecrosis (ON), a disease that can result in pain, loss of function, and disability. Hip core decompression surgery is an option aimed to prevent femoral head collapse and avoid future arthroplasty. Describe functional outcomes and gait quality among a young population with hip ON before and after hip core decompression. Study included participants with hip ON secondary to treatment for hematologic malignancy or sickle cell disease, between 8 and 29 years old, requiring hip core decompression surgery. At one-year follow-up, 13 participants (9 male, median age 17 years) completed the Functional Mobility Assessment (FMA), range of motion, and GAITRite® testing. Participants demonstrated improved mobility and endurance on the FMA at 1-year post-operatively compared to pre-operatively, with higher scores for time on the Timed Up and Go (mean FMA score = 2.92 [SD = 1.32] vs. 2.07 [SD = 1.70]), time on the Timed Up and Down Stairs (3.69 [0.85] vs. 2.92 [1.66]), and 9-Minute Walk Test scores for distance walked (2.69 [0.63] vs. 2.23 [0.93]) and heart rate (4.54 [0.66] vs. 3.31 [1.38]). GAITRite® analysis also showed improvements in many gait parameters at one-year follow-up. Cancer treatment complications other than ON could have contributed to results, not all eligible participants agreed to participate, and follow-up was only one year. Young patients with hip ON demonstrated improvements in functional mobility, endurance, and gait quality one year following hip core decompression.

Place de l’allogreffe de cellules souches hématopoïétiques après traitement par CAR T-cell anti-CD19 : recommandations de la SFGM-TC

The role of allogeneic hematopoietic cell transplantation (allo-HCT) after CAR T- treatment cells in hematologic malignancies is currently controversial. Prolonged remissions after several years of follow-up suggest that there is a curative effect of CAR T-cells therapy, whereas allo-HCT was previously considered the only curative treatment in relapse situation. The aim of this harmonization workshop is to detail the existing data in the literature on the feasibility of allo-HCT after CAR T-cells and to propose to consider allograft in selected patients with B-acute lymphoblastic leukemia (B-ALL) and diffuse large B-cell lymphoma (DLBCL). In B-ALL, various intrinsic factors (inherent to the patient, to the disease, to the type of CAR T-cells) and especially various post CAR T-cells criteria (early expansion kinetics, residual disease at D28, early loss of B-cell aplasia) should lead to consider performing allo-HCT before the occurrence of a relapse. In DLBCL, although there are risk factors for relapse at diagnosis and prior to CAR T-cells therapy, response assessed by PET-CT at three months is critical and allo-HCT cannot currently be recommended in cases of complete or partial remission. In any case, if the age is appropriate for allogeneic transplantation, HLA typing should be performed before CAR T-cells treatment in order not to delay the allo-HCT project if needed.

PB2012: A PHASE 2 MULTIARM STUDY OF MAGROLIMAB COMBINATIONS IN PATIENTS WITH RELAPSED/REFRACTORY MULTIPLE MYELOMA

Background: Magrolimab is a first-in-class humanized monoclonal antibody that blocks the immune checkpoint CD47, a “do not eat me” signal, overexpressed on tumor cells. Binding of magrolimab to CD47 leads to phagocytosis of cancer cells in human solid tumors and hematologic malignancies. CD47 is highly expressed in multiple myeloma (MM), and overexpression contributes to MM pathogenesis. Blocking of CD47 induces immediate activation of macrophages and elimination of myeloma cells. Hence, a blockade of CD47 with magrolimab may have therapeutic benefit in patients with MM. Additionally, in vitro studies have shown synergy between magrolimab and daratumumab in MM cell line phagocytosis, and complimentary mechanisms suggest potential enhanced efficacy in other combinations. Aims: To evaluate the safety, tolerability, and efficacy of magrolimab in combination with commonly used myeloma therapies in patients with relapsed/refractory MM (RRMM). Methods: This phase 2 open-label multiarm study includes patients aged ≥18 years currently requiring treatment for RRMM; all participants must have received ≥3 previous lines of therapy for MM, including an immunomodulatory drug and a proteasome inhibitor. This study includes safety run-in and corresponding dose-expansion phases with the following combinations: magrolimab + daratumumab, magrolimab + pomalidomide + dexamethasone, magrolimab + carfilzomib + dexamethasone, and magrolimab + bortezomib + dexamethasone (Figure). Magrolimab + bortezomib + dexamethasone may be initiated based on the preliminary safety and efficacy of magrolimab + carfilzomib + dexamethasone, and, if initiated, only requires 1 prior line of therapy. The primary objectives of the safety run-in phase are to evaluate the safety and tolerability of magrolimab and determine the recommended phase 2 dose (RP2D), with incidence of dose-limiting toxicities (DLTs), adverse events, and laboratory abnormalities as the primary endpoint. The dose-expansion phase will evaluate efficacy, with objective response rate as the primary endpoint. Secondary endpoints include duration of response, progression-free survival, and overall survival. Minimal residual disease negativity, biomarker changes from baseline, immune cell signaling alterations, and the mutational profile of myeloma cells will also be explored. Magrolimab will be administered intravenously as an initial priming dose to mitigate on-target anemia, followed by a maintenance dose on days 8, 15, 22, and 29 during cycle 1 (35 days); days 1, 8, 15, and 22 during cycle 2 (28 days); and days 1 and 15 from cycle 3 (28 days) onward (maximum of 8 cycles in patients who previously received bortezomib). Daratumumab, pomalidomide, dexamethasone, carfilzomib, and bortezomib will be administered according to manufacturer’s prescribing information. Dose de-escalation may occur based on DLTs per protocol in the safety run-in cohorts; magrolimab doses will not be reduced in the dose-expansion cohorts. Patients will continue treatment until unacceptable toxicity, progressive disease, or patient/investigator choice to discontinue. Planned enrollment is approximately 153 patients, with up to 27 in each safety run-in cohort and 72 in the dose-expansion cohorts following completion of the safety run-ins. Results: Trial in progress. Image:Summary/Conclusion: Patient recruitment is ongoing. Clinical trial information: NCT04892446.

Abstract 555: Allogeneic Natural Killer cells engineered to express HER2 CAR, Interleukin 15 and TGF beta dominant negative receptor effectively control HER2+ tumors

Abstract Despite the success of HER2 targeted therapies in HER2+ breast and gastric cancer, additional therapies are needed to address treatment-resistant metastatic disease. Adoptive immune cell therapy is a promising therapeutic modality given the remarkable clinical responses seen with autologous chimeric antigen receptor (CAR) T cells in hematological malignancies. However, success of cell therapy in solid tumors has been more limited. Three major impediments to the success of adoptive cell therapies in solid tumors are the heterogeneity of antigen expression, the immunosuppressive tumor microenvironment (TME), and the inherent challenges of manufacturing autologous cells and consequent variability of these cell products. Engineered, off-the-shelf, allogeneic Natural Killer (NK) cells provide a solution to these challenges. We describe here CAT-179, a novel engineered CAR-NK cell therapeutic for HER2+ solid tumors. CAT-179 cells express three transgenes: a HER2-directed CAR to effectively eliminate tumor cells, a Transforming Growth Factor (TGF) β dominant negative receptor (DNR) for resistance to TGFβ -mediated immunosuppression in the TME, and Interleukin 15 (IL15) cytokine to enhance NK cell persistence and activity for durable response. High efficiency engineering of the large (~3.7Kb) cargo containing CAR, IL15, and DNR in CAT-179 is enabled by the non-viral TC Buster™ Transposon System. Transposon engineering of CAT-179 results in high and stable expression of CAR (45% CAR at day 7 post gene delivery) without the need for post-engineering selection. CAT-179 demonstrates both CAR-dependent and innate NK receptor-dependent tumor cell killing in vitro, reducing the likelihood of tumor escape through antigen loss. CAT-179 effectively kills in vitro both high HER2-expressing SKOV3 cells as well as lower HER2-expressing HT-29 cells. CAT-179 also demonstrates resistance to TGFβ mediated immunosuppression, as evidenced by 75% reduction in TGFβ -induced phosphorylation of SMAD2 as well as prevention of TGFβ induced downregulation of NK cell activating receptors and restoration of NK cell cytotoxic activity. These data suggest CAT-179 cells will be protected from TGFβ -mediated immune suppression in the TME. Finally, the addition of IL15 in CAT-179 significantly enhances persistence for at least fourteen days in vitro without the need for exogenous cytokines. Moreover, CAT-179 administration to NSG mice showed expansion and persistence of the transferred cell product. CAT-179 addresses key hurdles to allogeneic cell therapy for solid tumors and is a promising new therapeutic approach for HER2 expressing breast, gastric and other tumors. Citation Format: Celeste Richardson, Finola Moore, Andres Alvarez, Alexia Barandiaran, Luke Barron, Eugene Choi, Tucker Ezell, Charlotte Franco, Bashar Hamza, Jennifer Johnson, Annie Khamhoung, Taeyoon Kyung, Marilyn Marques, Dominic Picarella, Jared Sewell, Alex Storer, Meghan Walsh, Vipin Suri. Allogeneic Natural Killer cells engineered to express HER2 CAR, Interleukin 15 and TGF beta dominant negative receptor effectively control HER2+ tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 555.

Abstract 4049: Preclinical pharmacology profile of GTX-0196: A novel, potent and highly selective dihydroorotate dehydrogenase (DHODH) inhibitor for the treatment of hematologic malignancies

Abstract Dihydroorotate dehydrogenase (DHODH) is a key enzyme in the de novo pyrimidine synthesis [Löffler et al., 1997]. Initial clinical studies with the DHODH inhibitor brequinar in solid tumors showed limited efficacy due to dose limiting adverse effects [Peters et al, 2018]. Recently, there is renewed interest in the use of more potent and selective DHODH inhibitors for the treatment of myeloid and lymphoid malignancies [Sykes et al, 2016]. Here we report the identification of the novel potent and highly selective DHODH inhibitor GTX-0196, with excellent physicochemical properties. GTX-0196 exhibited a biochemical IC50 of 3.7 nM in a DHODH biochemical assay and an antiproliferative IC50 of 1.1 nM in the MOLM-13 cell line, with brequinar showing IC50 values of 6.9 and 50 nM, respectively. The activity in the MOLM-13 proliferation assay was uridine-dependent, indicating absence of general toxicity of the compound and confirming DHODH-mediated efficacy. Profiling of GTX-0196 in the BioPrint panel (Eurofins/CEREP, at 10 µM), showed <50% inhibition for all 133 target proteins tested (GPCR’s, NR´s, kinases and other enzymes), illustrating the high selectivity for DHODH (> 2000-fold selectivity versus other targets tested). Furthermore, no inhibition was observed in a human CYP panel (1A2, 2A6, 2B6, 2C8, 2C9, 2C19, 2D6, 2E1 and 3A4, at 30 µM) and in a panel of transporters (at 10 µM). High metabolic stability in human liver microsomes and hepatocytes was observed with t½ > 6h and 10h, respectively. Compared to an earlier lead compound, GTX-0196 demonstrated enhanced potency and metabolic stability (particularly in preclinical species). Potent and efficacious inhibition of proliferation through DHODH was confirmed in a large number of solid tumor and hematologic malignancy cell lines, illustrating the broad potential use of this chemical series. Furthermore, tumor growth inhibition was observed in the MOLM-13 xenograft mouse model. Due to its favorable properties, GTX-0196 has progressed towards IND-enabling studies. In parallel, we are exploring mono and combination therapy opportunities in hematological malignancies as well as combination therapy opportunities in solid tumors to investigate the full therapeutic potential in oncology for this optimized DHODH inhibitor. Löffler et al. Mol. Cell. Biochem. 1997Peters GJ et al, Nucleosides, Nucleotides and Nucleic Acids 2018Sykes DB et al. Cell 2016 Citation Format: Allard Kaptein, Sonia Lain, Diede Brunen, Edward van Wezel, Tjeerd Barf. Preclinical pharmacology profile of GTX-0196: A novel, potent and highly selective dihydroorotate dehydrogenase (DHODH) inhibitor for the treatment of hematologic malignancies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4049.

Chimeric Antigen Receptor T-cells (CARs) in Cancer Treatment.

Cancer is one of the leading causes of death worldwide. Chemotherapy, radiation therapy, and stem cell transplantation were the main cancer treatment approaches for several years but due to their limited effectiveness, there was a constant search for new therapeutic approaches. Cancer immunotherapy that utilizes and enhances the normal capacity of the patient's immune system was used to fight against cancer. Genetically engineered T-cells that express Chimeric Antigen Receptors (CARs) showed remarkable anti-tumor activity against hematologic malignancies and are now being investigated in a variety of solid tumors. The use of this therapy in the last few years has been successful, achieving great success in improving the quality of life and prolonging the survival time of patients with a reduction in remission rates. However, many challenges still need to be resolved in order for this technology to gain widespread adoption. This review summarizes various experimental approaches towards the use of CAR T-- cells in hematologic malignancies and solid tumors. Finally, we address the challenges posed by CAR T-cells and discuss strategies for improving the performance of these T-cells in fighting cancers.

Update in TIGIT Immune-Checkpoint Role in Cancer.

The in-depth characterization of cross-talk between tumor cells and T cells in solid and hematological malignancies will have to be considered to develop new therapeutical strategies concerning the reactivation and maintenance of patient-specific antitumor responses within the patient tumor microenvironment. Activation of immune cells depends on a delicate balance between activating and inhibitory signals mediated by different receptors. T cell immunoreceptor with immunoglobulin and ITIM domain (TIGIT) is an inhibitory receptor expressed by regulatory T cells (Tregs), activated T cells, and natural killer (NK) cells. TIGIT pathway regulates T cell-mediated tumor recognition in vivo and in vitro and represents an exciting target for checkpoint blockade immunotherapy. TIGIT blockade as monotherapy or in combination with other inhibitor receptors or drugs is emerging in clinical trials in patients with cancer. The purpose of this review is to update the role of TIGIT in cancer progression, looking at TIGIT pathways that are often upregulated in immune cells and at possible therapeutic strategies to avoid tumor aggressiveness, drug resistance, and treatment side effects. However, in the first part, we overviewed the role of immune checkpoints in immunoediting, the TIGIT structure and ligands, and summarized the key immune cells that express TIGIT.

Targeting of Mevalonate-Isoprenoid Pathway in Acute Myeloid Leukemia Cells by Bisphosphonate Drugs

Metabolic reprogramming represents a hallmark of tumorigenesis to sustain survival in harsh conditions, rapid growth and metastasis in order to resist to cancer therapies. These metabolic alterations involve glucose metabolism, known as the Warburg effect, increased glutaminolysis and enhanced amino acid and lipid metabolism, especially the cholesterol biosynthesis pathway known as the mevalonate pathway and these are upregulated in several cancer types, including acute myeloid leukemia (AML). In particular, it was demonstrated that the mevalonate pathway has a pivotal role in cellular transformation. Therefore, targeting this biochemical process with drugs such as statins represents a promising therapeutic strategy to be combined with other anticancer treatments. In the last decade, several studies have revealed that amino-bisphosphonates (BP), primarily used for bone fragility disorders, also exhibit potential anti-cancer activity in leukemic cells, as well as in patients with symptomatic multiple myeloma. Indeed, these compounds inhibit the farnesyl pyrophosphate synthase, a key enzyme in the mevalonate pathway, reducing isoprenoid formation of farnesyl pyrophosphate and geranylgeranyl pyrophosphate. This, in turn, inhibits the prenylation of small Guanosine Triphosphate-binding proteins, such as Ras, Rho, Rac, Rab, which are essential for regulating cell survival membrane ruffling and trafficking, interfering with cancer key signaling events involved in clonal expansion and maturation block of progenitor cells in myeloid hematological malignancies. Thus, in this review, we discuss the recent advancements about bisphosphonates’ effects, especially zoledronate, analyzing the biochemical mechanisms and anti-tumor effects on AML model systems. Future studies will be oriented to investigate the clinical relevance and significance of BP treatment in AML, representing an attractive therapeutic strategy that could be integrated into chemotherapy.

Covid-19 in pediatric hematology-oncology and stem cell transplant patients –The spectrum of illness, complications and comparison of first two waves

IntroductionIndian subcontinent witnessed first wave of COVID-19 around March 2020 and second wave in April 2021. The mutant delta variant was ≈2.5 times more transmissible and led to the severe second wave. We compared the impact of two waves on pediatric hematology and oncology patients at our tertiary care centre that was at heart of managing COVID-19. MethodsChildren between 0 and 18 years, who were treated for a haematological illness, malignancy or stem cell transplant with confirmed COVID-19 infection or who developed multisystem inflammatory syndrome in children were included. ResultsA total of 48 (22-first, 26-second wave) children were evaluated. Despite better understanding of disease and standardised management algorithms, we found a trend towards younger age, increased requirements of oxygen, severe pneumonia and other post-covid complications in admitted patients during the second wave. We observed early RTPCR negativity in second wave. Invasive aspergillosis, disseminated candidiasis, reactivation of tuberculosis, HLH and MISC were the main complications. No child died of COVID-19. ConclusionThe second wave hit pediatric hematology and oncology patients harder than the first wave.COVID-19 infection in these patients may lead to significant morbidity and complications that interfere with treatment of their primary illnesses. They need close monitoring for development of life threatening infections. Early recognition and prompt therapy can optimise outcomes.

Optimized flow cytometric detection of transient receptor potential vanilloid-1 (TRPV1) in human hematological malignancies

The ectopic overexpression of transient receptor potential vanilloid-1 (TRPV1) has been detected in numerous solid cancers, including breast, prostate, pancreatic, and tongue epithelium cancer. However, the expression of TRPV1 in hematological malignancies remains unknown. Here we show through in silico analysis that elevated TRPV1 mRNA expression occurs in a range of hematological malignancies and presents an optimized flow cytometry method to rapidly assess TRPV1 protein expression for both cell lines and primary patient samples. Three anti-TRPV1 antibodies were evaluated for intracellular TRPV1 detection using flow cytometry resulting in an optimized protocol for the evaluation of TRPV1 in hematological malignant cell lines and patients’ peripheral blood mononuclear cells (PBMC). Overexpression of TRPV1 was observed in THP-1 (acute monocytic leukemia) and U266B1 (multiple myeloma, MM), but not U937 (histiocytic lymphoma) compared to healthy PBMC. TRPV1 was also detected in all 49 patients including B-cell non-Hodgkin’s lymphoma (B-NHL), MM, and others and 20 healthy controls. TRPV1 expression was increased in 8% of patients (MM = 2, B-NHL = 2). In conclusion, we provide an optimized flow cytometry method for routine expression analysis of clinical samples and show that TRPV1 is increased in a subset of patients with hematological malignancies.

Immunosuppression-Induced Pulmonary Alveolar Proteinosis Following Bilateral Lung Transplantation

<h3>Introduction</h3> Pulmonary alveolar proteinosis (PAP) has been described secondary to immunosuppression-induced macrophage dysfunction following lung transplantation. The incidence of infection is known to be increased in the setting of PAP. We present a case of immunosuppression-associated PAP in a lung transplant recipient with mycobacterium avium complex (MAC) infection, responsive to reduced immunosuppression, serial whole lung lavage, and MAC treatment. <h3>Case Report</h3> A 69-year-old male underwent bilateral lung transplantation for hypersensitivity pneumonitis (HP). He was never treated with mTOR inhibitor therapy. One year post-transplant, he developed intermittent cough and exertional dyspnea. CT scan showed new bilateral peribronchovascular groundglass opacities and interlobular septal thickening, initially concerning for recurrent HP. BAL fluid grew MAC on serial AFB cultures; infection cleared with treatment. Transbronchial biopsy showed A1B0 at the time of MAC infection, which cleared following treatment for acute cellular rejection. He developed increased groundglass opacities with superimposed thickened intra- and interlobular septae in polygonal shapes, "crazy paving" suggestive of pulmonary alveolar proteinosis. Exertional supplemental O2 requirement increased to 10 LPM. Milky-white opaque BAL fluid was noted on bronchoscopy. Transbronchial biopsies showed focal alveolar filling by granular eosinophilic material, consistent with PAP. GM-CSF Ab testing was negative. He had no history of hematologic malignancy, myelodysplastic syndrome, or hematopoietic cell transplantation. Exposure history was negative for dust. <h3>Summary</h3> Whole lung lavage showed cloudy BAL fluid that progressively cleared with continued lavage. Following ACR treatment, immunosuppression was weaned. Groundglass opacities resolved and allograft function improved. He weaned to 4 LPM O2 with exertion with reduction in respiratory symptoms.

A Bibliometric and Knowledge-Map Analysis of CAR-T Cells From 2009 to 2021.

ObjectivesA bibliometric and knowledge-map analysis is used to explore hotspots’ evolution and development trends in the CAR-T cell field. By looking for research hotspots and new topics, we can provide new clues and ideas for researchers in this field.MethodsThe articles and reviews regarding CAR-T cells were retrieved and obtained from the Web of Science Core Collection (WOSCC) on October 28th, 2021. CtieSpace [version 5.8.R3 (64-bit)] and VOSviewer (version 1.6.17) were used to conduct the bibliometric and knowledge-map analysis.Results660 authors from 488 institutions in 104 countries/regions published 6,867 papers in 1,212 academic journals. The United States was absolutely in the leading position in this research field. The institution that contributed the most publications was the University of Pennsylvania. Carl H June published the most articles, while Shannon L Maude had the most co-citations. However, there was little cooperation between countries. After 2012, cooperation among various institutions was also small. The journals that published the most CAR-T cell-related papers were Frontiers in immunology and Cancers. Nevertheless, Blood and The New England Journal of Medicine were the most commonly co-cited journals. The most influential research hotspots were the research of CAR-T cells in hematological malignancies, the related research of cytokine release syndrome (CRS), CD19, and the anti-tumor activity and efficacy of CAR-T cells. The latest hotspots and topics included the study of CAR-T cells in solid tumors, universal CAR-T cells, CAR-NK cells, CD22, and anakinra (the IL-1 receptor antagonist). The research of CAR-T cells in solid tumors was a rapidly developing hot field. Emerging topics in this field mainly included the study of CAR-T cells in glioblastoma (related targets: IL13Rα2, EGFRvIII, and HER2), neuroblastoma (related target: GD2), sarcoma (related target: HER2), and pancreatic cancer (related target: mesothelin), especially glioblastoma.ConclusionAs an anti-tumor therapy with great potential and clinical application prospects, CAR-T cell therapy is still in a stage of rapid development. The related field of CAR-T cells will remain a research hotspot in the future.

Real-World Use of Isavuconazole as Primary Therapy for Invasive Fungal Infections in High-Risk Patients with Hematologic Malignancy or Stem Cell Transplant.

(1) Introduction: Invasive fungal infections (IFIs) are a major cause of morbidity and mortality among immunocompromised patients with hematologic malignancies (HM) and stem cell transplants (SCT). Isavuconazole was approved by FDA as a primary therapy for Invasive Aspergillosis (IA) and Mucormycosis. The aim of this study is to look at the real-world use of Isavuconazole in patients with HM and evaluate their clinical outcomes and safety. (2) Methods: We conducted a retrospective study of HM patients at MD Anderson Cancer Center who had definite, probable or possible mold infections between 1 April 2016 and 31 January 2020 and were treated with Isavuconazole for a period of at least 7 days. Clinical and radiological findings were assessed at baseline and at 6 and 12 weeks of follow up. (3) Results: We included 200 HM patients with IFIs that were classified as definite (11), probable (63) and possible (126). Aspergillus spp was the most commonly isolated pathogen. The majority of patients (59%) received prophylaxis with anti-mold therapy and Isavuconazole was used as a primary therapy in 43% of patients, and as salvage therapy in 58%. The switch to Isavuconazole was driven by the failure of the primary therapy in 66% of the cases and by adverse effects in 29%. Isavuconazole was used as monotherapy in 30% of the cases and in combination in 70%. Adverse events possibly related to Isavuconazole were reported in eight patients (4%) leading to drug discontinuation. Moreover, a favorable response with Isavuconazole was observed in 40% at 6 weeks and in 60% at 12 weeks. There was no significant difference between isavuconazole monotherapy and combination therapy (p = 0.16 at 6 weeks and p = 0.06 at 12 weeks). Finally, there was no significant difference in outcome when Isavuconazole was used after failure of other anti-mold prophylaxis or treatment versus when used de novo as an anti-mold therapy (p = 0.68 at 6 weeks and p = 0.25 at 12 weeks). (4) Conclusions: Whether used as first-line therapy or after the failure of other azole and non-azole prophylaxis or therapies, isavuconazole seems to have a promising clinical response and a good safety profile as an antifungal therapy in high-risk cancer patients with hematologic malignancies. Moreover, combination therapy did not improve the outcome compared to Isavuconazole therapy.

Evaluation of central line salvage for mucosal barrier injury laboratory-confirmed bloodstream infection (MBI-LCBI) management practices in patients with hematologic malignancies

Patients with underlying hematologic malignancy (HM) and/or allogeneic hematopoietic stem cell transplant (HCT) recipients are at risk for mucosal barrier injury laboratory-confirmed bloodstream infection (MBI-LCBI) secondary to bacterial translocation. There is sparse data comparing MBI-LCBI management practices, in particular central venous catheter (CVC) salvage versus removal. We created a 22-item poll of Infectious Disease specialists at major US cancer centers on management controversies. Response rate was 44% (31/70). CVC salvage was a common practice among 87.5%. This was followed by a single center retrospective study (2017–2019) comparing outcomes related to CVC practices. We identified 115 patients, 52% (60/115) admitted for chemotherapy and 33% (38/115) for allogeneic HCT. The majority of patients (78%, 90/115) had their CVC removed. There was no difference in 72 h defervescence, microbiological clearance, in-hospital mortality, and 90-day recurrent infection between CVC salvage versus removal. CVC salvage is a safe approach in certain clinical scenarios.

Cytotoxicity of Medicinal Mushrooms Oudemansiella canarii and Ganoderma lucidum (Agaricomycetes) Against Hematologic Malignant Cells via Activation of Apoptosis-Related Markers.

Cancer is the second leading cause of death worldwide, and despite of the of the availability of the advanced chemical treatments, development of effective and safe alternatives derived from natural resources are still of high interest. Mushroom is one of the important resources of pharmacologically active cytotoxic compounds. In this paper, we report the cytotoxicity of ethanolic extracts of Oudemansiella canarii (Jungh.) Höhn. and Ganoderma lucidum (W. Curt.: Fr.) P. Karst. against nine hematologic malignant cells and describe their molecular mechanisms. Cell lines were exposed to varying concentrations of mushroom extracts for 48 h and the cell proliferation and apoptosis parameters were determined. Western blot analysis was performed to determine the extract-induced changes in the level of apoptosis-related proteins in cancer cell lines and patient-derived mononuclear cells. Results revealed that O. canarii and G. lucidum extracts exhibited cytotoxicity with IC50 values of 26.8-66.0 ppm and 48.1-78.4 ppm, respectively, in all the cancer cell lines used. Mushroom extracts inhibited cell proliferation by 57.3-72.5% (O. canarii) and 44.2-67.4% (G. lucidum), which correlates to the activation of apoptosis as indicated by increased annexin V positivity, cells in sub G0/G1 phase and production of reactive oxygen species, and decreased mitochondrial membrane potential. Western blot analysis showed increase in the level of apoptotic markers (cleaved PARP1, cleaved caspase 3 and phosphorylation of histone 2AX) and activation of the stress-activated protein kinase (SAPK/JNK) signaling pathway. The extract-activated apoptosis was also observed in mononuclear cells isolated from the peripheral blood of leukemia and lymphoma patients. In conclusion, activation of pro-apoptotic markers is one of the major mechanisms of the cytotoxicity of O. canarii and G. lucidum extracts against hematologic malignant cells.

2022 – CHARACTERIZING HETEROGENEITY IN HEALTHY AND MALIGNANT HEMATOPOIETIC HIERARCHIES USING SINGLE CELL PROTEOMICS

In recent years, single-cell studies demonstrated a high degree of molecular and functional heterogeneity within complex cellular systems. Single-cell transcriptome profiling has provided the basis for our current knowledge about stem cell heterogeneity. Although single-cell transcriptomics has given tremendous insight into heterogeneity, transcript levels are not always a good proxy for protein abundance. In this study, we demonstrate for the first time that single-cell mass spectrometry (scMS)-based proteomics enables the exploration of the global protein landscape of human hematopoietic stem & progenitor cells (HSPCs) in healthy aging and hematologic malignancies. By combining fluorescence-activated cell sorting (FACS) with our current experimental and computational scMS workflow, we are able to capture heterogeneity within these complex cellular hierarchies. Our analyses encompass scMS measurement of over 6000 single-cells, quantifying ∼1000 proteins per cell, from young and elderly individuals as well as patients with myelodysplastic syndromes. Our approach is able to recapitulate the differentiation trajectories of early human hematopoiesis. Besides, our scMS data highlights that the current FACS marker combinations appear to fall short of precise delineation of early human progenitors. We further show that Endomucin, a recently published marker of human HSCs, marks the stem cells at the top of the hierarchy. Finally, by integrating scMS and CITE-seq data, we aim to provide a better resolution of disease states based on the combined analysis of proteome and transcriptome. Overall, the results presented here support the power of global single-cell proteomics as a tool to characterize normal and perturbed developmental hierarchies. In recent years, single-cell studies demonstrated a high degree of molecular and functional heterogeneity within complex cellular systems. Single-cell transcriptome profiling has provided the basis for our current knowledge about stem cell heterogeneity. Although single-cell transcriptomics has given tremendous insight into heterogeneity, transcript levels are not always a good proxy for protein abundance. In this study, we demonstrate for the first time that single-cell mass spectrometry (scMS)-based proteomics enables the exploration of the global protein landscape of human hematopoietic stem & progenitor cells (HSPCs) in healthy aging and hematologic malignancies. By combining fluorescence-activated cell sorting (FACS) with our current experimental and computational scMS workflow, we are able to capture heterogeneity within these complex cellular hierarchies. Our analyses encompass scMS measurement of over 6000 single-cells, quantifying ∼1000 proteins per cell, from young and elderly individuals as well as patients with myelodysplastic syndromes. Our approach is able to recapitulate the differentiation trajectories of early human hematopoiesis. Besides, our scMS data highlights that the current FACS marker combinations appear to fall short of precise delineation of early human progenitors. We further show that Endomucin, a recently published marker of human HSCs, marks the stem cells at the top of the hierarchy. Finally, by integrating scMS and CITE-seq data, we aim to provide a better resolution of disease states based on the combined analysis of proteome and transcriptome. Overall, the results presented here support the power of global single-cell proteomics as a tool to characterize normal and perturbed developmental hierarchies.

Role of Endothelial Cells in Hematological Malignancies

Endothelial cells (EC) are key elements of vascularized tissues that form a single-cell layer that connects the vessels to the surrounding tissues. EC participate in the regulation of blood hemostasis, leukocyte homing, acute inflammation, wound healing, and antigen presentation. EC subpopulations are characterized by diverse structures, functions, and molecular profiles. In bone marrow, EC are part of the hematopoietic stem cell vascular and endosteal niche, where they play well-defined roles in hematopoietic stem cell functioning and maintenance, and reside surrounding sinusoids and blood vessels. In the past years, the clinical and pathophysiological roles of EC have been explored due to their contribution to neoangiogenesis and alterations in the vascular endothelium functions in hematological diseases. The present review discusses the EC contribution to pathogenesis of hematological malignancies and their potential use as therapeutic target in these diseases. Keywords: Endothelial cells, hematological malignancies, bone marrow niche, pathogenesis, angiogenesis.

The anticancer mechanism of human antimicrobial peptide LL-37

Human antimicrobial peptides LL-37 have a variety of medicinal uses. It has been portrayed that this peptide has robust tumoricidal action in a range of malignancies, particularly ovarian cancer, lung cancer, breast cancer, prostate cancer, pancreatic cancer, malignant melanoma, and squamous cell carcinoma of the skin. It exhibits substantial anticancer action against a range of cancers, including colon cancer, gastric cancer, hematologic malignancy, and oral squamous cell carcinoma (OSCC), in comparison. In this review, we explored in depth the anticancer mechanism of action of LL-37 in numerous sorts of cancer. We have shown how LL-37 impedes colon cancer by eliciting caspase-independent apoptosis. LL-37, in addition, has been noticed to boost tumor-suppressive bone morphogenetic protein signaling in gastric cancer cells via restricting the proteasome, which has been previously reported. In this research, we investigated how DNA methylation interferes with the activity of the human CAMP (Cathelicidin antimicrobial peptide gene) promoter and, as a result, acts as a tumor inhibitor in mouth squamous cell carcinoma. Additionally, how LL-37 inhibits cancer cell development in hematologic malignancy has been explored through caspase-independent but Ca2+/calpain- and AIF-dependent processes.