ABSTRACT
FMS-related tyrosine kinase 3 ligand (FLT3L), encoded by FLT3LG, is a hematopoietic factor essential for the development of natural killer (NK) cells, B cells, and dendritic cells (DCs) in mice. We describe three humans homozygous for a loss-of-function FLT3LG variant with a history of various recurrent infections, including severe cutaneous warts. The patients' bone marrow (BM) was hypoplastic, with low levels of hematopoietic progenitors, particularly myeloid and B cell precursors. Counts of B cells, monocytes, and DCs were low in the patients' blood, whereas the other blood subsets, including NK cells, were affected only moderately, if at all. The patients had normal counts of Langerhans cells (LCs) and dermal macrophages in the skin but lacked dermal DCs. Thus, FLT3L is required for B cell and DC development in mice and humans. However, unlike its murine counterpart, human FLT3L is required for the development of monocytes but not NK cells.
Subject(s)
Killer Cells, Natural , Membrane Proteins , Animals , Female , Humans , Male , Mice , B-Lymphocytes/metabolism , B-Lymphocytes/cytology , Bone Marrow/metabolism , Cell Lineage , Dendritic Cells/metabolism , Hematopoiesis , Hematopoietic Stem Cells/metabolism , Hematopoietic Stem Cells/cytology , Killer Cells, Natural/metabolism , Killer Cells, Natural/immunology , Langerhans Cells/metabolism , Membrane Proteins/metabolism , Membrane Proteins/genetics , Monocytes/metabolism , Skin/metabolism , Mice, Inbred C57BLABSTRACT
Unlike macrophage networks composed of long-lived tissue-resident cells within specific niches, conventional dendritic cells (cDCs) that generate a 3D network in lymph nodes (LNs) are short lived and continuously replaced by DC precursors (preDCs) from the bone marrow (BM). Here, we examined whether specific anatomical niches exist within which preDCs differentiate toward immature cDCs. In situ photoconversion and Prtn3-based fate-tracking revealed that the LN medullary cords are preferential entry sites for preDCs, serving as specific differentiation niches. Repopulation and fate-tracking approaches demonstrated that the cDC1 network unfolded from the medulla along the vascular tree toward the paracortex. During inflammation, collective maturation and migration of resident cDC1s to the paracortex created discontinuity in the medullary cDC1 network and temporarily impaired responsiveness. The decrease in local cDC1 density resulted in higher Flt3L availability in the medullary niche, which accelerated cDC1 development to restore the network. Thus, the spatiotemporal development of the cDC1 network is locally regulated in dedicated LN niches via sensing of cDC1 densities.
Subject(s)
Lymph Nodes , Macrophages , Cell Differentiation , Dendritic CellsABSTRACT
Poor maternal diet during pregnancy is a risk factor for severe lower respiratory infections (sLRIs) in the offspring, but the underlying mechanisms remain elusive. Here, we demonstrate that in mice a maternal low-fiber diet (LFD) led to enhanced LRI severity in infants because of delayed plasmacytoid dendritic cell (pDC) recruitment and perturbation of regulatory T cell expansion in the lungs. LFD altered the composition of the maternal milk microbiome and assembling infant gut microbiome. These microbial changes reduced the secretion of the DC growth factor Flt3L by neonatal intestinal epithelial cells and impaired downstream pDC hematopoiesis. Therapy with a propionate-producing bacteria isolated from the milk of high-fiber diet-fed mothers, or supplementation with propionate, conferred protection against sLRI by restoring gut Flt3L expression and pDC hematopoiesis. Our findings identify a microbiome-dependent Flt3L axis in the gut that promotes pDC hematopoiesis in early life and confers disease resistance against sLRIs.
Subject(s)
Microbiota , Respiratory Tract Infections , Animals , Female , Mice , Pregnancy , Dendritic Cells , Diet , PropionatesABSTRACT
Despite advances in single-cell multi-omics, a single stem or progenitor cell can only be tested once. We developed clonal multi-omics, in which daughters of a clone act as surrogates of the founder, thereby allowing multiple independent assays per clone. With SIS-seq, clonal siblings in parallel "sister" assays are examined either for gene expression by RNA sequencing (RNA-seq) or for fate in culture. We identified, and then validated using CRISPR, genes that controlled fate bias for different dendritic cell (DC) subtypes. This included Bcor as a suppressor of plasmacytoid DC (pDC) and conventional DC type 2 (cDC2) numbers during Flt3 ligand-mediated emergency DC development. We then developed SIS-skew to examine development of wild-type and Bcor-deficient siblings of the same clone in parallel. We found Bcor restricted clonal expansion, especially for cDC2s, and suppressed clonal fate potential, especially for pDCs. Therefore, SIS-seq and SIS-skew can reveal the molecular and cellular mechanisms governing clonal fate.
Subject(s)
Dendritic Cells/metabolism , Proto-Oncogene Proteins/genetics , Proto-Oncogene Proteins/metabolism , Repressor Proteins/genetics , Repressor Proteins/metabolism , Animals , Cell Differentiation/genetics , Cell Line , Cell Lineage/genetics , Female , Gene Expression/genetics , HEK293 Cells , Humans , Male , Membrane Proteins/genetics , Membrane Proteins/metabolism , Mice, Inbred C57BL , Stem Cells/metabolismABSTRACT
In tumors, a subset of CD8+ T cells expressing the transcription factor TCF-1 drives the response to immune checkpoint blockade. We examined the mechanisms that maintain these cells in an autochthonous model of lung adenocarcinoma. Longitudinal sampling and single-cell sequencing of tumor-antigen specific TCF-1+ CD8+ T cells revealed that while intratumoral TCF-1+ CD8+ T cells acquired dysfunctional features and decreased in number as tumors progressed, TCF-1+ CD8+ T cell frequency in the tumor draining LN (dLN) remained stable. Two discrete intratumoral TCF-1+ CD8+ T cell subsets developed over time-a proliferative SlamF6+ subset and a non-cycling SlamF6- subset. Blocking dLN egress decreased the frequency of intratumoral SlamF6+ TCF-1+ CD8+ T cells. Conventional type I dendritic cell (cDC1) in dLN decreased in number with tumor progression, and Flt3L+anti-CD40 treatment recovered SlamF6+ T cell frequencies and decreased tumor burden. Thus, cDC1s in tumor dLN maintain a reservoir of TCF-1+ CD8+ T cells and their decrease contributes to failed anti-tumor immunity.
Subject(s)
Adenocarcinoma of Lung/immunology , CD8-Positive T-Lymphocytes/immunology , Dendritic Cells/immunology , Lung Neoplasms/immunology , Lymph Nodes/immunology , T Cell Transcription Factor 1/immunology , Animals , Mice , T-Lymphocyte Subsets/immunologyABSTRACT
Mutant isocitrate dehydrogenase (IDH) 1 and 2 play a pathogenic role in cancers, including acute myeloid leukemia (AML), by producing oncometabolite 2-hydroxyglutarate (2-HG). We recently reported that tyrosine phosphorylation activates IDH1 R132H mutant in AML cells. Here, we show that mutant IDH2 (mIDH2) R140Q commonly has K413 acetylation, which negatively regulates mIDH2 activity in human AML cells by attenuating dimerization and blocking binding of substrate (α-ketoglutarate) and cofactor (NADPH). Mechanistically, K413 acetylation of mitochondrial mIDH2 is achieved through a series of hierarchical phosphorylation events mediated by tyrosine kinase FLT3, which phosphorylates mIDH2 to recruit upstream mitochondrial acetyltransferase ACAT1 and simultaneously activates ACAT1 and inhibits upstream mitochondrial deacetylase SIRT3 through tyrosine phosphorylation. Moreover, we found that the intrinsic enzyme activity of mIDH2 is much higher than mIDH1, thus the inhibitory K413 acetylation optimizes leukemogenic ability of mIDH2 in AML cells by both producing sufficient 2-HG for transformation and avoiding cytotoxic accumulation of intracellular 2-HG.
Subject(s)
Isocitrate Dehydrogenase/genetics , Leukemia, Myeloid, Acute/metabolism , Acetyl-CoA C-Acetyltransferase/metabolism , Acetylation , Animals , Antineoplastic Agents/pharmacology , Female , Humans , Isocitrate Dehydrogenase/metabolism , Ketoglutaric Acids/metabolism , Leukemia, Myeloid, Acute/genetics , Lysine/genetics , Lysine/metabolism , Male , Mice , Mice, Inbred NOD , Mutation/genetics , NADP/metabolism , Nuclear Proteins/metabolism , Phosphorylation , Polymorphism, Single Nucleotide/genetics , Primary Cell Culture , Protein Binding , Protein Processing, Post-Translational , Protein-Tyrosine Kinases/metabolismABSTRACT
Dendritic cells (DCs) are professional antigen-presenting cells that play a key role in shaping adaptive immunity. DCs have a unique ability to sample their environment, capture and process exogenous antigens into peptides that are then loaded onto major histocompatibility complex class I molecules for presentation to CD8+ T cells. This process, called cross-presentation, is essential for initiating and regulating CD8+ T cell responses against tumors and intracellular pathogens. In this review, we will discuss the role of DCs in cancer immunity, the molecular mechanisms underlying antigen cross-presentation by DCs, the immunosuppressive factors that limit the efficiency of this process in cancer, and approaches to overcome DC dysfunction and therapeutically promote antitumoral immunity.
Subject(s)
Cross-Priming , Neoplasms , Humans , CD8-Positive T-Lymphocytes , Antigen Presentation , Antigens , Neoplasms/therapy , Immunotherapy , Dendritic CellsABSTRACT
Arenaviral vaccine vectors encoding simian immunodeficiency virus (SIV) immunogens are capable of inducing efficacious humoral and cellular immune responses in nonhuman primates. Several studies have evaluated the use of immune modulators to further enhance vaccine-induced T-cell responses. The hematopoietic growth factor Flt3L drives the expansion of various bone marrow progenitor populations, and administration of Flt3L was shown to promote expansion of dendritic cell populations in spleen and blood, which are targets of arenaviral vectors. Therefore, we evaluated the potential of Flt3 signaling to enhance the immunogenicity of arenaviral vaccines encoding SIV immunogens (SIVSME543 Gag, Env, and Pol) in rhesus macaques, with a rhesus-specific engineered Flt3L-Fc fusion protein. In healthy animals, administration of Flt3L-Fc led to a 10- to 100-fold increase in type 1 dendritic cells 7 days after dosing, with no antidrug antibody (ADA) generation after repeated dosing. We observed that administration of Flt3L-Fc fusion protein 7 days before arenaviral vaccine increased the frequency and activation of innate immune cells and enhanced T-cell activation with no treatment-related adverse events. Flt3L-Fc administration induced early innate immune activation, leading to a significant enhancement in magnitude, breadth, and polyfunctionality of vaccine-induced T-cell responses. The Flt3L-Fc enhancement in vaccine immunogenicity was comparable to a combination with αCTLA-4 and supports the use of safe and effective variants of Flt3L to augment therapeutic vaccine-induced T-cell responses.IMPORTANCEInduction of a robust human immunodeficiency virus (HIV)-specific CD4+ and CD8+ T-cell response through therapeutic vaccination is considered essential for HIV cure. Arenaviral vaccine vectors encoding simian immunodeficiency virus (SIV) immunogens have demonstrated strong immunogenicity and efficacy in nonhuman primates. Here, we demonstrate that the immunogenicity of arenaviral vectors encoding SIV immunogens can be enhanced by administration of Flt3L-Fc fusion protein 7 days before vaccination. Flt3L-Fc-mediated increase in dendritic cells led to robust improvements in vaccine-induced T- and B-cell responses compared with vaccine alone, and Flt3L-Fc dosing was not associated with any treatment-related adverse events. Importantly, immune modulation by either Flt3L-Fc or αCTLA-4 led to comparable enhancement in vaccine response. These results indicate that the addition of Flt3L-Fc fusion protein before vaccine administration can significantly enhance vaccine immunogenicity. Thus, safe and effective Flt3L variants could be utilized as part of a combination therapy for HIV cure.
Subject(s)
Dendritic Cells , Macaca mulatta , SAIDS Vaccines , Simian Immunodeficiency Virus , Animals , Simian Immunodeficiency Virus/immunology , Dendritic Cells/immunology , SAIDS Vaccines/immunology , Simian Acquired Immunodeficiency Syndrome/immunology , Simian Acquired Immunodeficiency Syndrome/prevention & control , Membrane Proteins/immunology , Membrane Proteins/genetics , fms-Like Tyrosine Kinase 3/immunology , fms-Like Tyrosine Kinase 3/genetics , Antibodies, Viral/immunology , Antibodies, Viral/blood , Genetic Vectors , Immunogenicity, Vaccine , CD8-Positive T-Lymphocytes/immunologyABSTRACT
Small tandem DNA duplications in the range of 15 to 300 base-pairs play an important role in the aetiology of human disease and contribute to genome diversity. Here, we discuss different proposed mechanisms for their occurrence and argue that this type of structural variation mainly results from mutagenic repair of chromosomal breaks. This hypothesis is supported by both bioinformatical analysis of insertions occurring in the genome of different species and disease alleles, as well as by CRISPR/Cas9-based experimental data from different model systems. Recent work points to fill-in synthesis at double-stranded DNA breaks with complementary sequences, regulated by end-joining mechanisms, to account for small tandem duplications. We will review the prevalence of small tandem duplications in the population, and we will speculate on the potential sources of DNA damage that could give rise to this mutational signature. With the development of novel algorithms to analyse sequencing data, small tandem duplications are now more frequently detected in the human genome and identified as oncogenic gain-of-function mutations. Understanding their origin could lead to optimized treatment regimens to prevent therapy-induced activation of oncogenes and might expose novel vulnerabilities in cancer.
Subject(s)
Chromosome Breakage , DNA End-Joining Repair , Genome, Human , Microsatellite Repeats , Humans , CRISPR-Cas SystemsABSTRACT
MiR-126 and miR-155 are key microRNAs (miRNAs) that regulate, respectively, hematopoietic cell quiescence and proliferation. Herein we showed that in acute myeloid leukemia (AML), the biogenesis of these two miRNAs is interconnected through a network of regulatory loops driven by the FMS-like tyrosine kinase 3-internal tandem duplication (FLT3-ITD). In fact, FLT3-ITD induces the expression of miR-155 through a noncanonical mechanism of miRNA biogenesis that implicates cytoplasmic Drosha ribonuclease III (DROSHA). In turn, miR-155 down-regulates SH2-containing inositol phosphatase 1 (SHIP1), thereby increasing phosphor-protein kinase B (AKT) that in turn serine-phosphorylates, stabilizes, and activates Sprouty related EVH1 domain containing 1 (SPRED1). Activated SPRED1 inhibits the RAN/XPO5 complex and blocks the nucleus-to-cytoplasm transport of pre-miR-126, which cannot then complete the last steps of biogenesis. The net result is aberrantly low levels of mature miR-126 that allow quiescent leukemia blasts to be recruited into the cell cycle and proliferate. Thus, miR-126 down-regulation in proliferating AML blasts is downstream of FLT3-ITDdependent miR-155 expression that initiates a complex circuit of concatenated regulatory feedback (i.e., miR-126/SPRED1, miR-155/human dead-box protein 3 [DDX3X]) and feed-forward (i.e., miR-155/SHIP1/AKT/miR-126) regulatory loops that eventually converge into an output signal for leukemic growth.
Subject(s)
Leukemia, Myeloid, Acute , MicroRNAs , fms-Like Tyrosine Kinase 3 , DEAD-box RNA Helicases/metabolism , Down-Regulation , Humans , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/metabolism , MicroRNAs/metabolism , Mutation , fms-Like Tyrosine Kinase 3/genetics , fms-Like Tyrosine Kinase 3/metabolismABSTRACT
Corilagin (CLG) has antitumor activities in certain human malignant cancers. Herein, the effects and mechanisms of CLG on osteosarcoma (OS) were investigated. OS cell viability and proliferation were detected by MTT and colony formation assay. Cell cycle and apoptosis were examined using flow cytometry. The interaction between TRAF6 and FLT3 was investigated using a co-immunoprecipitation assay. Results demonstrated that CLG treatment inhibited OS cell viability and proliferation but promoted OS cell autophagy and apoptosis in a concentration-dependent manner. Mechanically, CLG inhibited TRAF6-mediated FLT3 ubiquitination degradation. TRAF6 overexpression abolished the effects of CLG on OS cell proliferation, autophagy, and apoptosis. Finally, CLG administration inhibited OS tumor growth in mice by inducing autophagy-dependent apoptosis. Taken together, CLG inhibited OS progression by facilitating mTOR/ULK1 pathway-mediated autophagy through inhibiting TRAF6-mediated FLT3 ubiquitination, which indicated that CLG was a promising candidate for the treatment of OS.
ABSTRACT
Data regarding the use of FMS-like tyrosine kinase 3 (FLT3) and isocitrate dehydrogenase 1/2 (IDH1/2) inhibitors in acute lymphoblastic leukaemia (ALL) are lacking. We identified 14 patients with FLT3- or IDH1/2-mutated ALL. Three early T-cell precursor-ALL patients received FLT3 or IDH2 inhibitors. Patient 1 maintains a complete remission (CR) with enasidenib after intolerance to chemotherapy. Patient 2 maintained a CR for 27 months after treatment with enasidenib for relapsed disease. Patient 3 was treated with venetoclax and gilteritinib at the time of relapse and maintained a CR with gilteritinib for 8 months. These cases suggest that FLT3 and IDH inhibitors could represent a viable therapeutic option for ALL patients with these mutations.
Subject(s)
Aminopyridines , Aniline Compounds , Leukemia, Myeloid, Acute , Precursor Cell Lymphoblastic Leukemia-Lymphoma , Pyrazines , Triazines , Humans , fms-Like Tyrosine Kinase 3/genetics , Neoplasm Recurrence, Local , Enzyme Inhibitors/therapeutic use , Mutation , Precursor Cell Lymphoblastic Leukemia-Lymphoma/drug therapy , Precursor Cell Lymphoblastic Leukemia-Lymphoma/genetics , Leukemia, Myeloid, Acute/geneticsABSTRACT
We have conducted a retrospective, single-centre analysis of 20 patients with relapsed or refractory FLT3-mutated acute myeloid leukaemia (FLT3m AML) who received a salvage quadruplet regimen consisting of gilteritinib, venetoclax, low-dose cytarabine and actinomycin D (G-ACTIVE). G-ACTIVE resulted in a 95% (19/20) overall response rate and 75% (15/20) complete remission and complete remission with an incomplete platelet recovery (CR + CRp) rate. Out of 13 transplant-eligible patients, 11 (86%) proceeded to an allogeneic stem cell transplantation. The median overall survival and relapse-free survival after G-ACTIVE were 32 and 12.9 months respectively. The Day 60 mortality rate was 15%.
Subject(s)
Aniline Compounds , Bridged Bicyclo Compounds, Heterocyclic , Cytarabine , Leukemia, Myeloid, Acute , Pyrazines , Sulfonamides , Humans , Dactinomycin , Retrospective Studies , Neoplasm Recurrence, Local , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/genetics , fms-Like Tyrosine Kinase 3/geneticsABSTRACT
Patients with FLT3-mutated acute myeloid leukaemia (AML) that relapse or are refractory (R/R) to intensive induction have poor outcomes. Gilteritinib has recently become standard-of-care for patients with R/R FLT3-mutated AML. We investigated whether adding venetoclax to gilteritinib (gilt-ven) improves outcomes as compared with gilteritinib monotherapy. We included patients treated with gilteritinib (n = 19) and gilt-ven (n = 17) for R/R AML after intensive chemotherapy. Gilteritinib and gilt-ven groups did not differ in terms of mCRc rates (53% and 65%, p = 0.51) and realization of allogeneic haematopoietic stem-cell transplantation (HSCT, 47% and 35%, p = 0.5). Overall survival (OS) was comparable between groups, although a trend towards better OS was seen with gilt-ven (12-month OS 58.8% [95% CI 39.5%-87.6%]) versus gilteritinib (42.1% [95% CI 24.9%-71.3%] for gilteritinib). Early salvage with gilt-ven versus any other gilteritinib-based approach was associated with the best outcome (p = 0.031). Combination therapy was associated with increased haematological toxicity. In summary, gilt-ven did not improve remissions or HSCT-realization rates in patients with R/R FLT3-mutated AML as compared with gilteritinib and was associated with increased haematological toxicity. Although OS did not differ, a trend towards better survival was suggested with gilt-ven and a survival benefit was shown for gilt-ven approach when sequenced early for salvage.
ABSTRACT
In vitro culture of bone marrow (BM) with Fms-like tyrosine kinase 3 ligand (Flt3L) is widely used to study development and function of type 1 conventional dendritic cells (cDC1). Hematopoietic stem cells (HSCs) and many progenitor populations that possess cDC1 potential in vivo do not express Flt3 and thus may not contribute to Flt3L-mediated cDC1 production in vitro. Here, we present a KitL/Flt3L protocol that recruits such HSCs and progenitors into the production of cDC1. Kit ligand (KitL) is used to expand HSCs and early progenitors lacking Flt3 expression into later stage where Flt3 is expressed. Following this initial KitL phase, a second Flt3L phase is used to support the final production of DCs. With this two-stage culture, we achieved approximately tenfold increased production of both cDC1 and cDC2 compared to Flt3L culture. cDC1 derived from this culture are similar to in vivo cDC1 in their dependence on IRF8, ability to produce IL-12, and induction of tumor regression in cDC1-deficient tumor-bearing mice. This KitL/Flt3L system for cDC1 production will be useful in further analysis of cDC1 that rely on in vitro generation from BM.
Subject(s)
Hematopoietic Stem Cells , Stem Cell Factor , Mice , Animals , Bone Marrow , Bone Marrow Cells , Dendritic CellsABSTRACT
The development of dendritic cells (DCs) depends on signaling via the FMS-like tyrosine kinase 3 (Flt3) receptor. How Flt3 signaling impacts terminally differentiated DC function is unknown. This is important given the increasing interest in exploiting Flt3 for vaccination and tumor immunotherapy. Here, we examined DCs in mice harboring constitutively activated Flt3 (Flt3-ITD). Flt3ITD/ITD mice possessed expanded splenic DC subsets including plasmacytoid DC, conventional DC (cDC)1, cDC2, double positive (DP) cDC1 (CD11c+ CD8+ CD11b- CD103+ CD86+), noncanonical (NC) cDC1 (CD11c+ CD8+ CD11b- CD103- CD86-) and single positive (SP) cDC1 (CD11c+ CD8+ CD11b- CD103- CD86+). Outcomes of constitutive Flt3 signaling differed depending on the cDC subset examined. In comparison with wild type (WT) DCs, all Flt3ITD/ITD cDCs displayed an altered surface phenotype with changes in costimulatory molecules, major histocompatibility complex class I (MHC I) and II (MHC II). Cytokine secretion patterns, antigen uptake, antigen proteolysis and antigen presenting function differed between WT and Flt3ITD/ITD subsets, particularly cDC2. In summary, Flt3 signaling impacts the function of terminally differentiated cDCs with important consequences for antigen presentation.
Subject(s)
Antigen Presentation , Dendritic Cells , Signal Transduction , fms-Like Tyrosine Kinase 3 , Animals , Dendritic Cells/immunology , Dendritic Cells/metabolism , fms-Like Tyrosine Kinase 3/metabolism , Mice , Mice, Inbred C57BL , Cytokines/metabolism , Cell DifferentiationABSTRACT
BACKGROUND: Patients with relapsed or refractory acute myeloid leukemia (R/R AML) and FLT3-internal tandem duplication (FLT3-ITD) respond infrequently to salvage chemotherapy. OBJECTIVE: To investigate the efficacy of sorafenib plus triplet therapy with venetoclax, azacitidine, and homoharringtonine (VAH) as a salvage therapy in this population. METHODS: This multicenter, single-arm, phase 2 study was conducted at 12 hospitals across China. Eligible patients had R/R AML with FLT3-ITD (aged 18-65 years) who were treated with VAH. The primary endpoint was composite complete remission (CRc) after two cycles. Secondary outcomes included the overall response rate (ORR), safety, and survival. RESULTS: Between July 9, 2020, and March 19, 2022, 58 patients were assessed for eligibility, 51 of whom were enrolled. The median patient age was 47 years (interquartile range [IQR] 31-57). CRc was 76.5% with ORR of 82.4%. At a median follow-up of 17.7 months (IQR, 8.7-24.7), the median duration of CRc was not reached (NR), overall survival was 18.1 months (95% confidence interval [CI], 11.8-NR) and event-free survival was 11.4 months (95% CI, 5.6-NR). Grade 3 or 4 adverse events occurring in ≥10% of patients included neutropenia in 47 (92.2%), thrombocytopenia in 41 (80.4%), anemia in 35 (68.6%), febrile neutropenia in 29 (56.9%), pneumonia in 13 (25.5%), and sepsis in 6 (11.8%) patients. Treatment-related death occurred in two (3.9%) patients. CONCLUSIONS: The sorafenib plus VAH regimen was well tolerated and highly active against R/R AML with FLT3-ITD. This regimen may be a suitable therapeutic option for this population, but larger population trials are needed to be explored. TRIAL REGISTRATION: Clinical Trials Registry: NCT04424147.
Subject(s)
Azacitidine , Bridged Bicyclo Compounds, Heterocyclic , Leukemia, Myeloid, Acute , Sulfonamides , Humans , Azacitidine/therapeutic use , fms-Like Tyrosine Kinase 3/genetics , fms-Like Tyrosine Kinase 3/therapeutic use , Homoharringtonine/therapeutic use , Leukemia, Myeloid, Acute/therapy , Pathologic Complete Response , Sorafenib/adverse effects , Adolescent , Young Adult , Adult , Middle Aged , AgedABSTRACT
The ability to detect low-level disease is key to our understanding of clonal heterogeneity in acute myeloid leukemia (AML) and residual disease that elude conventional assays and seed relapse. We developed a high-sensitivity next-generation sequencing (HS-NGS) clinical assay, able to reliably detect low levels (1 × 10-5) of FLT3-ITD, a frequent, therapeutically targetable and prognostically relevant mutation in AML. By applying this assay to 289 longitudinal samples from 62 patients at initial diagnosis and/or clinical follow-up (mean follow-up of 22 months), we reveal the frequent occurrence of FLT3-ITD subclones at diagnosis and demonstrate a significantly decreased relapse risk when FLT3-ITD is cleared after induction or thereafter. We perform pairwise sequencing of diagnosis and relapse samples from 23 patients to uncover more detailed patterns of FLT3-ITD clonal evolution at relapse than is detectable by less-sensitive assays. Finally, we show that rising ITD level during consecutive biopsies is a harbinger of impending relapse. Our findings corroborate the emerging clinical utility of high-sensitivity FLT3-ITD testing and expands our understanding of clonal dynamics in FLT3-ITD-positive AML.
ABSTRACT
FLT3L-Fc is a cytokine-Fc fusion agonizing receptor-type tyrosine-protein kinase FLT3 (fms-related tyrosine kinase 3; CD135). FLT3 is expressed on dendritic cells (DCs) as well as myeloid and lymphoid progenitors. Nonclinical pharmacokinetics, pharmacodynamics and safety of FLT3L-Fc were investigated in rats and cynomolgus monkeys. FLT3L-Fc induced robust pharmacodynamic responses, evidenced by marked expansion of peripheral blood cDC1s, cDC2s, and pDCs (up to 301-fold in rats and 378-fold in monkeys), peaking at 8-10 days after the first dose. FLT3L-Fc was well tolerated with no adverse findings at doses up to 10 mg/kg administered intravenously twice three weeks apart. In both species, major clinical pathology findings consisted of expansion of white blood cell (WBC) populations including lymphocytes, monocytes, neutrophils, basophils, and large unstained cells, which were pronounced after the first dose. The WBC findings were associated microscopically with histiocytic and mononuclear cell infiltrates in multiple organs. Tissue immunohistochemistry in monkeys showed that the leukocyte infiltrates consisted of hematopoietic progenitor cells and histiocytes with a reactive morphology and were associated with a slight stimulation of regional T and B cell populations. Additional FLT3L-Fc-associated changes included decreases in red blood cell (RBC) mass, increases in RBC distribution width, variable changes in reticulocytes, and transient alterations in platelet counts (rats only). The RBC and WBC findings were associated microscopically with increased hematopoietic cellularity of the bone marrow in both species and increased splenic megakaryocytic extramedullary hematopoiesis in rats. The totality of nonclinical safety data support the clinical development of FLT3L-Fc.
Subject(s)
Membrane Proteins , Neoplasms , Rats , Animals , Dendritic Cells , Hematopoietic Stem Cells , ImmunotherapyABSTRACT
The FLT3-ITD mutation represents the most frequent genetic alteration in newly diagnosed acute myeloid leukemia (AML) patient and is associated with poor prognosis. Mutation result in the retention of a constitutively active form of this receptor in the endoplasmic reticulum (ER) and the subsequent modification of its downstream effectors. Here, we assessed the impact of such retention on ER homeostasis and found that mutant cells present lower levels of ER stress due to the overexpression of ERO1α, one of the main proteins of the protein folding machinery at the ER. Overexpression of ERO1α resulted essential for ITD mutant cells survival and chemoresistance and also played a crucial role in shaping the type of glucose metabolism in AML cells, being the mitochondrial pathway the predominant one in those with a higher ER stress (non-mutated cells) and the glycolytic pathway the predominant one in those with lower ER stress (mutated cells). Our data indicate that FLT3 mutational status dictates the route for glucose metabolism in an ERO1α depending on manner and this provides a survival advantage to tumors carrying these ITD mutations.