Deregulated calcium signaling in blood cancer: Underlying mechanisms and therapeutic potential

Intracellular calcium signaling regulates diverse physiological and pathological processes. In solid tumors, changes to calcium channels and effectors via mutations or changes in expression affect all cancer hallmarks. Such changes often disrupt transport of calcium ions (Ca$^{2+}$) in the endoplasmic reticulum (ER) or mitochondria, impacting apoptosis. Evidence rapidly accumulates that this is similar in blood cancer. Principles of intracellular Ca$^{2+}$ signaling are outlined in the introduction. We describe different Ca$^{2+}$-toolkit components and summarize the unique relationship between extracellular Ca$^{2+}$ in the endosteal niche and hematopoietic stem cells. The foundational data on Ca$^{2+}$ homeostasis in red blood cells is discussed, with the demonstration of changes in red blood cell disorders. This leads to the role of Ca$^{2+}$ in neoplastic erythropoiesis. Then we expand onto the neoplastic impact of deregulated plasma membrane Ca$^{2+}$ channels, ER Ca$^{2+}$ channels, Ca$^{2+}$ pumps and exchangers, as well as Ca$^{2+}$ sensor and effector proteins across all types of hematologic neoplasms. This includes an overview of genetic variants in the Ca$^{2+}$-toolkit encoding genes in lymphoid and myeloid cancers as recorded in publically available cancer databases. The data we compiled demonstrate that multiple Ca$^{2+}$ homeostatic mechanisms and Ca$^{2+}$ responsive pathways are altered in hematologic cancers. Some of these alterations may have genetic basis but this requires further investigation. Most changes in the Ca$^{2+}$-toolkit do not appear to define/associate with specific disease entities but may influence disease grade, prognosis, treatment response, and certain complications. Further elucidation of the underlying mechanisms may lead to novel treatments, with the aim to tailor drugs to different patterns of deregulation. To our knowledge this is the first review of its type in the published literature. We hope that the evidence we compiled increases awareness of the calcium signaling deregulation in hematologic neoplasms and triggers more clinical studies to help advance this field

N-Methyl-D-aspartate receptors in hematopoietic cells: what have we learned?

The N-methyl-D-aspartate receptor (NMDAR) provides a pathway for glutamate-mediated inter-cellular communication, best known for its role in the brain but with multiple examples of functionality in non-neuronal cells. Data previously published by others and us provided ex vivo evidence that NMDARs regulate platelet and red blood cell (RBC) production. Here, we summarize what is known about these hematopoietic roles of the NMDAR. Types of NMDAR subunits expressed in megakaryocytes (platelet precursors) and erythroid cells are more commonly found in the developing rather than adult brain, suggesting trophic functions. Nevertheless, similar to their neuronal counterparts, hematopoietic NMDARs function as ion channels, and are permeable to calcium ions (Ca2+). Inhibitors that block open NMDAR (memantine and MK-801) interfere with megakaryocytic maturation and proplatelet formation in primary culture. The effect on proplatelet formation appears to involve Ca2+ influx-dependent regulation of the cytoskeletal remodeling. In contrast to normal megakaryocytes, NMDAR effects in leukemic Meg-01 cells are diverted away from differentiation to increase proliferation. NMDAR hypofunction triggers differentiation of Meg-01 cells with the bias toward erythropoiesis. The underlying mechanism involves changes in the intracellular Ca2+ homeostasis, cell stress pathways, and hematopoietic transcription factors that upon NMDAR inhibition shift from the predominance of megakaryocytic toward erythroid regulators. This ability of NMDAR to balance both megakaryocytic and erythroid cell fates suggests receptor involvement at the level of a bipotential megakaryocyte-erythroid progenitor. In human erythroid precursors and circulating RBCs, NMDAR regulates intracellular Ca2+ homeostasis. NMDAR activity supports survival of early proerythroblasts, and in mature RBCs NMDARs impact cellular hydration state, hemoglobin oxygen affinity, and nitric oxide synthase activity. Overexcitation of NMDAR in mature RBCs leads to Ca2+ overload, K+ loss, RBC dehydration, and oxidative stress, which may contribute to the pathogenesis of sickle cell disease. In summary, there is growing evidence that glutamate-NMDAR signaling regulates megakaryocytic and erythroid cells at different stages of maturation, with some intriguing differences emerging in NMDAR expression and function between normal and diseased cells. NMDAR signaling may provide new therapeutic opportunities in hematological disease, but in vivo applicability needs to be confirmed

N-methyl-D-aspartate receptor regulates the circadian clock in megakaryocytic cells and impacts cell proliferation through BMAL1

Peripheral circadian clocks control cell proliferation and survival, but little is known about their role and regulation in megakaryocytic cells. N-methyl-D-aspartate receptor (NMDAR) regulates the central clock in the brain. The purpose of this study was to determine whether NMDAR regulates the megakaryocytic cell clock and whether the megakaryocytic clock regulates cell proliferation and cell death. We found that both the Meg-01 megakaryocytic cell line and native murine megakaryocytes expressed circadian clock genes. Megakaryocyte-directed deletion of Grin1 in mice caused significant disruption of the circadian rhythm pathway at the transcriptional level and increased expression of BMAL1 at the protein level. Similarly, both pharmacological (MK-801) and genetic (GRIN–/–) inhibition of NMDAR in Meg-01 cells in vitro resulted in widespread changes in clock gene expression including increased expression of BMAL1, the core clock transcription factor. BMAL1 overexpression reduced Meg-01 cell proliferation and altered the time-dependent expression of the cell cycle regulators MYC and WEE1, whereas BMAL1 knockdown led to increased cell death in Meg-01-GRIN1–/– cells. Our results demonstrate that NMDAR regulates the circadian clock in megakaryocytic cells and that the circadian clock component BMAL1 contributes to the control of Meg-01 cell proliferation and survival

Ionotropic glutamate receptors in platelets: opposing effects and a unifying hypothesis

Ionotropic glutamate receptors include α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPAR), kainate receptors (KAR), and N-methyl-D-aspartate receptors (NMDAR). All function as cation channels; AMPAR and KAR are more permeable to sodium and NMDAR to calcium ions. Compared to the brain, receptor assemblies in platelets are unusual, suggesting distinctive functionalities.There is convincing evidence that AMPAR and KAR amplify platelet function and thrombus formation in vitro and in vivo. Transgenic mice lacking GluA1 and GluK2 (AMPAR and KAR subunits, respectively) have longer bleeding times and prolonged time to thrombosis in an arterial model. In humans, rs465566 KAR gene polymorphism associates with altered in vitro platelet responses suggesting enhanced aspirin effect. The NMDAR contribution to platelet function is less well defined. NMDA at low concentrations (≤10 μM) inhibits platelet aggregation and high concentrations (≥100 μM) have no effect. However, open NMDAR channel blockers interfere with platelet activation and aggregation induced by other agonists in vitro; anti-GluN1 antibodies interfere with thrombus formation under high shear rates ex vivo; and rats vaccinated with GluN1 develop iron deficiency anemia suggestive of mild chronic bleeding. In this review, we summarize data on glutamate receptors in platelets and propose a unifying model that reconciles some of the opposing effects observed

Insights into the role of JAK2-I724T variant in myeloproliferative neoplasms from a unique cohort of New Zealand patients

This study aimed to compile bioinformatic and experimental information for JAK2 missense variants previously reported in myeloproliferative neoplasms (MPN) and determine if germline JAK2-I724T, recently found to be common in New Zealand Polynesians, associates with MPN. For all JAK2 variants found in the literature, gnomAD_exome allele frequencies were extracted and REVEL scores were calculated using the dbNSFP database. We investigated the prevalence of JAK2-I724T in a cohort of 111 New Zealand MPN patients using a TaqMan assay, examined its allelic co-occurrence with JAK2-V617F using Oxford Nanopore sequencing, and modelled the impact of I724T on JAK2 using I-Mutant and ChimeraX software. Several non-V617F JAK2 variants previously reported in MPN had REVEL scores greater than 0.5, suggesting pathogenicity. JAK2-I724T (REVEL score 0.753) was more common in New Zealand Polynesian MPN patients (n = 2/27; 7.4%) than in other New Zealand patients (n = 0/84; 0%) but less common than expected for healthy Polynesians (n = 56/377; 14.9%). Patients carrying I724T (n = 2), one with polycythaemia vera and one with essential thrombocythaemia, had high-risk MPN. Both patients with JAK2-I724T were also positive for JAK2-V617F, found on the same allele as I724T, as well as separately. In silico modelling did not identify noticeable structural changes that would give JAK2-I724T a gain-of-function. Several non-canonical JAK2 variants with high REVEL scores have been reported in MPN, highlighting the need to further understand their relationship with disease. The JAK2-I724T variant does not drive MPN, but additional investigations are required to exclude any potential modulatory effect on the MPN phenotype.</p