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Effect of glucose metabolic inhibitors and ibrutinib on CLL metabolism. a, c, e, f Survival fraction (relative to non-treated control (NT)) of CLL lymphocytes treated with metabolic inhibitors for 48 h. b n = 21, c n = 12, e n = 19, f n = 23. b Glucose uptake after 24 h compound treatment (n = 15). d Relative ROS median values of CLL cells after 24 h of ibrutinib and/or NAC treatment (n = 20). *p < 0.05, **p < 0.001. g Model for basal wildtype and del11q CLL cell metabolism. The proposed metabolic flux in CLL lymphocytes is indicated with brown (wild-type) and pink (del11q) arrows. Del11q CLL lymphocytes promote glutamine synthesis, while decreasing GDH reaction. Glutamate production might be maintained via amino-acid catabolism through transaminase reactions, while glycolysis could provide the necessary metabolites to feed the TCA, PPP, and one-carbon metabolism. Wild-type CLL lymphocytes have a significant flux of glucose-derived carbons through PPP, while their amino-acid metabolism tends to generate ammonia
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Citations
... ATM and TP53, in addition to their role in controlling genomic integrity, also appear to be central regulators of carbon metabolism, thus correlating with metabolic and molecular profile [13]. In fact, del(11q) CLL cells would appear to be particularly sensitive to inhibition of glycolysis [14]. Similarly, Kluckova K et al. [15] reported that CLL cells in proliferation centers of several samples from patients with deletion of chromosome 17p (del17p) manifested increased spontaneous aerobic glycolysis in the BCR-unstimulated state (suggestive of a BCR-independent metabolic phenotype). ...
Chronic Lymphocytic Leukemia (CLL) is an indolent malignancy characterized by the accumulation of quiescent mature B cells. However, these cells are transcriptionally and translationally active, implicating an active metabolism. The recent literature suggests that CLL cells have an oxidative-type phenotype. Given the role of cell metabolism, which is able to influence the outcome of treatments, in other neoplasms, we aimed to assess its prognostic role in CLL patients by determining the ex vivo bioenergetic metabolic profile of CLL cells, evaluating the correlation with the patient clinical/biological characteristics and the in vivo response to BTK inhibitor treatment. Clustering analysis of primary samples identified two groups, characterized by low (CLL low) or high (CLL high) bioenergetic metabolic rates. Compared to the CLL high, CLL with lower bioenergetic metabolic rates belonged to patients characterized by a statistically significant higher white blood cell count and by unfavorable molecular genetics. More importantly, patients in the CLL low cluster displayed a better and more durable response to the BTK inhibitor ibrutinib, thus defining a bioenergetic metabolic subgroup that can benefit the most from this therapy.
... Glycolysis helps to fuel mitochondrial oxidative phosphorylation (OXPHOS) and is more active in unmutated IGHV CLL cells (associated with a poor prognosis) [9,10]. Amino-acids like glutamine and arginine are centrally positioned in the CLL cell's carbon and nitrogen metabolic pathways [11,12]. Ions like Ca2+ and Fe2 + /3+ influence the CLL cell's signal transduction mechanisms [13,14]. ...
Chronic lymphocytic leukemia (CLL) is still an incurable disease, with many patients developing resistance to conventional and targeted therapies. To better understand the physiology of CLL and facilitate the development of innovative treatment options, we examined specific metabolic features in the tumor CLL B-lymphocytes. We observed metabolic reprogramming, characterized by a high level of mitochondrial oxidative phosphorylation activity, a low glycolytic rate, and the presence of C2- to C6-carnitine end-products revealing an unexpected, essential role for peroxisomal fatty acid beta-oxidation (pFAO). Accordingly, downmodulation of ACOX1 (a rate-limiting pFAO enzyme overexpressed in CLL cells) was enough to shift the CLL cells’ metabolism from lipids to a carbon- and amino-acid-based phenotype. Complete blockade of ACOX1 resulted in lipid droplet accumulation and caspase-dependent death in CLL cells, including those from individuals with poor cytogenetic and clinical prognostic factors. In a therapeutic translational approach, ACOX1 inhibition spared non-tumor blood cells from CLL patients but led to the death of circulating, BCR-stimulated CLL B-lymphocytes and CLL B-cells receiving pro-survival stromal signals. Furthermore, a combination of ACOX1 and BTK inhibitors had a synergistic killing effect. Overall, our results highlight a less-studied but essential metabolic pathway in CLL and pave the way towards the development of new, metabolism-based treatment options.
... Metabolic reprogramming plays a pivotal role in cancer development and patient survival. Compared to other B-cell malignancies, chronic lymphocytic leukemia (CLL) is not highly active metabolically (1); however, it develops metabolic modifications that underlie its progression and resistance to drugs (2)(3)(4). Some of these modifications affect oxidative phosphorylation (OXPHOS) and help cancer cells use alternatives to glucose substrates to produce adenosine triphosphate (ATP) (5). ...
... Continuous treatment with anti-BTK and anti-BCL-2 therapies is typically required in the management of CLL, and the success of these therapies is heavily dependent on the emergence of drug resistance over time. Metabolic reprogramming, which leads to effects such as a higher oxygen consumption rate (OCR) and altered glutamine and glucose metabolism in cells, contributes to the cells' resistance to venetoclax and ibrutinib (4,17). An examination of the transcriptomic landscape of ibrutinib-resistant mantle cell lymphoma cells revealed enhanced reliance on OXPHOS and glutaminolysis (18). ...
... In addition, compound 968 (a GLS-1 inhibitor) was cytotoxic for the del(11q) CLL cells, in which ROS levels were increased upon GLS-1 inhibition. Glutamine and glutamate uptake was not affected by compound 968, suggesting that CLL lymphocytes are able to maintain constant concentrations of these metabolites (4). Nevertheless, compound 968 has different pharmacological characteristics than CB-839 and is less effective in disrupting glutaminyls (41). ...
Introduction:
Chronic lymphocytic leukemia (CLL) cells are metabolically flexible and adapt to modern anticancer treatments. Bruton tyrosine kinase (BTK) and B-cell lymphoma-2 (BCL-2) inhibitors have been widely used to treat CLL, but CLL cells become resistant to these treatments over time. CB-839 is a small-molecule glutaminase-1 (GLS-1) inhibitor that impairs glutamine use, disrupts downstream energy metabolism, and impedes the elimination of reactive oxygen species.
Methods:
To investigate the in vitro effects of CB-839 on CLL cells, we tested CB-839 alone and in combination with ibrutinib, venetoclax, or AZD-5991 on the HG-3 and MEC-1 CLL cell lines and on primary CLL lymphocytes.
Results:
We found that CB-839 caused dose-dependent decreases in GLS-1 activity and glutathione synthesis. CB-839-treated cells also showed increased mitochondrial superoxide metabolism and impaired energy metabolism, which were reflected in decreases in the oxygen consumption rate and depletion of the adenosine triphosphate pool and led to the inhibition of cell proliferation. In the cell lines, CB-839 combined with venetoclax or AZD-5991, but not with ibrutinib, demonstrated synergism with an increased apoptosis rate and cell proliferation inhibition. In the primary lymphocytes, no significant effects of CB-839 alone or in combination with venetoclax, ibrutinib, or AZD-5991 were observed.
Discussion:
Our findings suggest that CB-839 has limited efficacy in CLL treatment and shows limited synergy in combination with widely used CLL drugs.
... Another team explored metabolic plasticity of CLL cells using a panel of metabolic inhibitors. These Metabolic inhibitors, including glycolysis inhibitor (2-deoxyglucose) and oxidative phosphorylation (OxPhos) inhibitor (oligomycin), also resulted in cytotoxicity of CLL cells in-vitro [29]. These observations opened the door for the development of novel immune-therapeutics specifically targeting metabolic rewiring of malignant B cells. ...
Targeting toll-like receptors (TLRs), via TLR agonists, has been implicated in the regulation of immunometabolism. B-chronic lymphocytic leukemia (B-CLL) represents a suitable model for B-cell derived malignancies with shifted metabolic adaptations. Several signaling pathways have been found to be critical in metabolic reprogramming of CLL, including mechanistic target of rapamycin- hypoxia inducible factor-1α (mTOR- HIF-1α) pathway, the main metabolic regulator of glycolysis. Here, we investigated the effect of TLR7/8 agonist (Resiquimod) on the expression of mTOR and HIF-1α in patients with CLL. B cells were purified using Rosettesep Human B cell Enrichment Cocktail (Stem cell Technologies, Vancouver, BC, Canada#15,024) from peripheral venous blood of CLL patients (n = 20) and healthy individuals (n = 15). Isolated B cells were then cultured in both presence and absence of Resiquimod. Gene expression of mTOR and HIF-1α were assessed using qRT-PCR. Resiquimod significantly decreased mTOR and HIF-1α gene expression in both CLL (p < 0.001and p < 0.001, respectively) and Normal B cells (p = 0.004 and p = 0.001, respectively). Resiquimod may reprogram immunometabolism of malignant B-CLL cells via down-regulation of key glycolytic metabolic actors, mTOR and HIF-1α genes. Accordingly, Resiquimod may be an adjuvant as a therapeutic tool for CLL, which needs to be studied further.
Supplementary Information
The online version contains supplementary material available at 10.1007/s12288-023-01649-y.
... Overexpression of oxoglutarate dehydrogenase and isocitrate dehydrogenase was commonly found in CLL cells [125]. In line with this, CLL cells are sensitive to the pharmacological inhibition of oxidative phosphorylation by OXPHOS inhibitors (PK11195, oligomycin A, and metformin) (Figure 2) [126]. ...
... Given this evidence, 11q-deleted CLL cells are more sensitive to glycolysis inhibition. Currently, there are clinical trials targeting the mitochondrial OXPHOS via metformin (NCT01750567) alone or in combination with GLUT4 (glucose transporter) via ritonavir (NCT02948283) [126]. ...
... Del11q CLL patients exhibit higher glutamine synthesis and metabolism than their negative counterparts. Del11q CLL samples are susceptible to glutaminase inhibitors indicating the pivotal role of glutamine metabolism in Del11q CLL [126]. Glutamine is taken up by the cells via glutamine transporters such as SLC1A5, SLC38A1, and SLC38A2 [131]. ...
Simple Summary
Despite being a slow-proliferating disease, chronic lymphocytic leukemia (CLL) is an incurable and frequently reoccurring adult leukemia. Although large-scale genome-wide next-generation sequencing studies have provided insights into CLL’s transcriptome and mutational landscape, the molecular mechanisms underlying disease progression remain incompletely understood. Over time, the treatment landscape in CLL has shifted from chemoimmunotherapies (CIT) to targeted therapies, but resistance mechanisms have emerged, leading to progression such as Richter’s transformation (RT). As a result, there remains an unmet clinical need to identify new therapeutic strategies. In our review article, we aim to evaluate the past and current state of CLL treatment in both frontline and relapsed/refractory settings and also explore mitochondrial reprogramming, metabolic alterations, and RNA splicing as potential novel therapeutic targets.
Abstract
Over the past decade, the treatment landscape of CLL has vastly changed from the conventional FC (fludarabine and cyclophosphamide) and FCR (FC with rituximab) chemotherapies to targeted therapies, including inhibitors of Bruton tyrosine kinase (BTK) and phosphatidylinositol 3-kinase (PI3K) as well as inhibitors of BCL2. These treatment options dramatically improved clinical outcomes; however, not all patients respond well to these therapies, especially high-risk patients. Clinical trials of immune checkpoint inhibitors (PD-1, CTLA4) and chimeric antigen receptor T (CAR T) or NK (CAR NK) cell treatment have shown some efficacy; still, long-term outcomes and safety issues have yet to be determined. CLL remains an incurable disease. Thus, there are unmet needs to discover new molecular pathways with targeted or combination therapies to cure the disease. Large-scale genome-wide whole-exome and whole-genome sequencing studies have discovered genetic alterations associated with disease progression, refined the prognostic markers in CLL, identified mutations underlying drug resistance, and pointed out critical targets to treat the disease. More recently, transcriptome and proteome landscape characterization further stratified the disease and revealed novel therapeutic targets in CLL. In this review, we briefly summarize the past and present available single or combination therapies, focusing on potential emerging therapies to address the unmet clinical needs in CLL.
... Amino acids: amino acids play a crucial role in the progression of B-cell malignancies. In peculiar, addiction of tumor cells to glutamine is observed in many tumors including MM [392] and CLL with 11q deletion [393], and in the Burkitt lymphoma cell line P493 [394]. The glutaminase inhibitor CB-839 (telaglenastat) has been tested in phase I clinical trial in MM and WM, but the results are not yet available (NCT02071888). ...
Resistance to death is one of the hallmarks of human B cell malignancies and often contributes to the lack of a lasting response to today’s commonly used treatments. Drug discovery approaches designed to activate the death machinery have generated a large number of inhibitors of anti-apoptotic proteins from the B-cell lymphoma/leukemia 2 family and the B-cell receptor (BCR) signaling pathway. Orally administered small-molecule inhibitors of Bcl-2 protein and BCR partners (e.g., Bruton’s tyrosine kinase and phosphatidylinositol-3 kinase) have already been included (as monotherapies or combination therapies) in the standard of care for selected B cell malignancies. Agonistic monoclonal antibodies and their derivatives (antibody–drug conjugates, antibody–radioisotope conjugates, bispecific T cell engagers, and chimeric antigen receptor-modified T cells) targeting tumor-associated antigens (TAAs, such as CD19, CD20, CD22, and CD38) are indicated for treatment (as monotherapies or combination therapies) of patients with B cell tumors. However, given that some patients are either refractory to current therapies or relapse after treatment, novel therapeutic strategies are needed. Here, we review current strategies for managing B cell malignancies, with a focus on the ongoing clinical development of more effective, selective drugs targeting these molecules, as well as other TAAs and signaling proteins. The observed impact of metabolic reprogramming on B cell pathophysiology highlights the promise of targeting metabolic checkpoints in the treatment of these disorders.
... Previous research indicated the consumption of Gln was increased in CLL [65]. Elevated ammonia uptake and Gln synthetase expression are found in del11q CLL lymphocytes, favoring de novo Gln synthesis [66]. The expression of glutamate dehydrogenase (GDH) is decreased in del11q CLL cells, which benefits transaminase reactions using α-ketoglutarate for glutamate synthesis and reduces oxidative deamination of glutamate. ...
... MYC may upregulate the glucose metabolism in CLL cells leading to disease progression. The human tumor suppressor gene ATM, located on the long arm of chromosome 11, relates to DNA damage, cell cycle progression, p53 dysfunction and metabolism [66,140]. In CLL, the ATM mutation is a poor independent prognostic biomarker for time to first treatment (TTFT) [141]. ...
... In CLL, the ATM mutation is a poor independent prognostic biomarker for time to first treatment (TTFT) [141]. The underexpression of ATM is associated with the deletion of chromosome 11 (del11q), while del11q CLL lymphocytes reprogram glutamine metabolism and inhibit glucose metabolism [66]. Besides, increased expression of insulin receptors is found in del11q CLL [142]. ...
Metabolic reprogramming, fundamentally pivotal in carcinogenesis and progression of cancer, is considered as a promising therapeutic target against tumors. In chronic lymphocytic leukemia (CLL) cells, metabolic abnormalities mediate alternations in proliferation and survival compared with normal B cells. However, the role of metabolic reprogramming is still under investigation in CLL. In this review, the critical metabolic processes of CLL were summarized, particularly glycolysis, lipid metabolism and oxidative phosphorylation. The effects of T cells and stromal cells in the microenvironment on metabolism of CLL were also elucidated. Besides, the metabolic alternation is regulated by some oncogenes and tumor suppressor regulators, especially TP53, MYC and ATM. Thus, the agents targeting metabolic enzymes or signal pathways may impede the progression of CLL. Both the inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR) statins and the lipoprotein lipase inhibitor orlistat induce the apoptosis of CLL cells. In addition, a series of oxidative phosphorylation inhibitors play important roles in decreasing the proliferation of CLL cells. We epitomized recent advancements in metabolic reprogramming in CLL and discussed their clinical potentiality for innovative therapy options. Metabolic reprogramming plays a vital role in the initiation and progression of CLL. Therapeutic approaches targeting metabolism have their advantages in improving the survival of CLL patients. This review may shed novel light on the metabolism of CLL, leading to the development of targeted agents based on the reshaping metabolism of CLL cells.
... Resting B-cells and memory B-cells are more glycolytic than activated B-cells (77). However, in CLL, the mutated B-cells rely on OxPhos over glycolysis and have reduced glucose uptake unlike normal HSCs (78,79). CLL lymphocytes have an increase in ROS plus mitochondrial respiration and also have an active antioxidant activity via glutathione metabolism, protecting CLL cells from chemotherapy ( Figure 3) (80, 81). ...
Therapeutic targeting of leukemic stem cells is widely studied to control leukemia. An emerging approach gaining popularity is altering metabolism as a potential therapeutic opportunity. Studies have been carried out on hematopoietic and leukemic stem cells to identify vulnerable pathways without impacting the non-transformed, healthy counterparts. While many metabolic studies have been conducted using stem cells, most have been carried out in vitro or on a larger population of progenitor cells due to challenges imposed by the low frequency of stem cells found in vivo. This creates artifacts in the studies carried out, making it difficult to interpret and correlate the findings to stem cells directly. This review discusses the metabolic difference seen between hematopoietic stem cells and leukemic stem cells across different leukemic models. Moreover, we also shed light on the advancements of metabolic techniques and current limitations and areas for additional research of the field to study stem cell metabolism.
... While the impact of BCR-associated kinase inhibitors on cellular bioenergetics has not been well studied, emerging evidence suggests that metabolic state may impact therapeutic outcomes, and that cellular bioenergetic responses and BTK activity are interdependent. Ibrutinib was shown to induce a metabolic stress response in primary neoplastic lymphocytes, and in vitro drug sensitivity was modulated following perturbations in cellular bioenergetics [39,40]. In another study, CLL cells obtained from patients treated with ibrutinib exhibited a decreased OCR, on par with that of normal lymphocytes [41]. ...
Aberrant B-cell receptor (BCR) signaling is a key driver in lymphoid malignancies. Bruton tyrosine kinase (BTK) inhibitors that disrupt BCR signaling have received regulatory approvals in therapy of mantle cell lymphoma (MCL). However, responses are incomplete and patients who experience BTK inhibitor therapy failure have dire outcomes. CG-806 (luxeptinib) is a dual BTK/SYK inhibitor in clinical development in hematologic malignancies. Here we investigated the pre-clinical activity of CG-806 in MCL. In vitro treatment with CG-806 thwarted survival of MCL cell lines and patient-derived MCL cells in a dose-dependent manner. CG-806 blocked BTK and SYK activation and abrogated BCR signaling. Contrary to ibrutinib, CG-806 downmodulated the anti-apoptotic proteins Mcl-1 and Bcl-xL, abrogated survival of ibrutinib-resistant MCL cell lines, and partially reversed the pro-survival effects of stromal microenvironment-mimicking conditions in primary MCL cells. Dual BTK/SYK inhibition led to mitochondrial membrane depolarization accompanied by mitophagy and metabolic reprogramming toward glycolysis. In vivo studies of CG-806 demonstrated improved survival in one of the two tested aggressive MCL PDX models. While suppression of the anti-apoptotic Bcl-2 family proteins and NFκB signaling correlated with in vivo drug sensitivity, OxPhos and MYC transcriptional programs were upregulated in the resistant model following treatment with CG-806. BAX and NFKBIA were implicated in susceptibility to CG-806 in a whole-genome CRISPR-Cas9 library screen (in a diffuse large B-cell lymphoma cell line). A high-throughput in vitro functional drug screen demonstrated synergy between CG-806 and Bcl-2 inhibitors. In sum, dual BTK/SYK inhibitor CG-806 disrupts BCR signaling and induces metabolic reprogramming and apoptosis in MCL. The Bcl-2 network is a key mediator of sensitivity to CG-806 and combined targeting of Bcl-2 demonstrates synergy with CG-806 warranting continued exploration in lymphoid malignancies.
... 20 It has been reported that BTK has a role in negatively regulating ROS levels, and ibrutinib can enhance ROS production in CLL. [21][22] Then we used the DCFH-DA probe to evaluate the effect of 13l on ROS level in Z138 cells. As shown in Figure 4B, the green uorescence intensity gradually increased as the concentrations of 13l increased. ...
Development of Bruton’s tyrosine kinase (BTK) inhibitors is of great value and significance in the treatment of B-cell malignancies and autoimmune diseases. Herein, a novel class of pyrazolopyrimidine-based BTK inhibitors were designed and evaluated in mantle cell lymphoma (MCL) cell lines. We demonstrated that target compounds had made great progress in improvement of antiproliferative activity compared to lead compound. Compounds 13c , 13g , 13h , 13l , 13n and 13o demonstrated effectively antiproliferative activity in MCL cells lines with single-digit micromolar potency. Furthermore, compound 13l specifically disturbed mitochondrial membrane potential and increased reactive oxygen species level in Z138 cells in a dose-dependent manner. 13l induced cell apoptosis through the caspase 3- mediated apoptotic pathway in Z138 cells. Overall, this study provides valuable lead compounds for developing antitumor agents.
