Metabolism Of Malignant Cells Research Articles

Microproteins/micropeptides dysregulation contributes to cancer progression and development: A mechanistic review.

Microproteins, known as micropeptides, are small protein molecules encoded by short open reading frames. These recently identified molecules have been proven to be an essential part of the human proteome that participates in multiple processes, such as DNA repair, mitochondrial respiration, and regulating different signaling pathways. A growing body of studies has evidenced that microproteins exhibit dysregulated expression levels in various malignancies and contribute to tumor progression. It has been reported that microproteins interact with many proteins, such as enzymes (e.g., adenosine triphosphate synthase) and signal transducers (e.g., c-Jun), and regulate malignant cell metabolism, proliferation, and metastasis. Moreover, microproteins have been found to play a significant role in multidrug resistance in vitro and in vivo by their activity in DNA repair pathways. Considering that, this review intended to summarize the roles of microproteins in different aspects of tumorigenesis with diagnostic and therapeutic perspectives.

Cancer cell metabolism and antitumour immunity.

Accumulating evidence suggests that metabolic rewiring in malignant cells supports tumour progression not only by providing cancer cells with increased proliferative potential and an improved ability to adapt to adverse microenvironmental conditions but also by favouring the evasion of natural and therapy-driven antitumour immune responses. Here, we review cancer cell-intrinsic and cancercell-extrinsic mechanisms through which alterations of metabolism in malignant cells interfere with innate and adaptive immune functions in support of accelerated disease progression. Further, we discuss the potential of targeting such alterations to enhance anticancer immunity for therapeutic purposes.

Targeting Glucose Metabolism in Cancer Immunotherapy: A Promising Avenue for Enhanced Treatment Strategies

Cancer immunotherapy has emerged as an innovative strategy for treating multiple types of cancer. Recent breakthroughs have emphasized the significance of altering glucose metabolism in malignant cells; specifically, the heightened dependency of cancer cells on glucose metabolism (“Warburg effect”), where cancer cells alter their metabolic pathway so that they utilize glucose as an energy source, even when oxygen is present. This adaptation allows for rapid tumor cell proliferation, enabling them to grow and divide at an accelerated pace. Current research concentrates on the development of cancer treatments aimed at inhibiting glucose metabolism in cancer cells. This approach aims to improve the effectiveness of existing immunotherapies by utilizing the metabolic pathways of malignant cells through direct and indirect inhibition of glycolysis. This review discusses the interplay between glucose metabolism and immune response, explores current efforts in manipulating glucose metabolism for therapeutic benefits, and provides insights into future directions for optimizing this strategy.

IRF4 Rewires Tumor Metabolism with Functional Implications on the Immune Microenvironment in Lymphoma

Reprogramming of cellular metabolism is a hallmark of cancer leading to improved cellular fitness and providing a selective advantage to malignant cells. Recently, we found that follicular lymphoma (FL) patients with poor prognosis display increased transcriptional and protein expression of IRF4 (IRF4 high), altered immune signaling and an immunosuppressive tumor microenvironment (TME). Furthermore, high expression of IRF4 rewired B cell metabolism towards glycolysis ( Mondello P et al, Blood 2022;140 Suppl 1:168-169). However, it's unknown whether this metabolic reprogramming may be responsible in part for the impairment of immune cells in the TME, which also rely on glucose for their proliferation and function. Using RNA-seq on 88 FL patients, we found a negative correlation between IRF4-regulated genes associated with glycolysis and tumor infiltration by major immune cell subsets in IRF4 high compared to IRF4 low tumors ( Fig 1A), suggesting that IRF4-driven tumor glycolysis may reinforce an immune suppressed phenotype. Especially, we observed that ALDOA and GPI genes were inversely correlated with expression of CD4 + T cells (r=0.34, p=0.003; and r=0.38, p=0.015, respectively) and T reg cells (r=0.3, p=0.05; and r=0.35, p=0.027, respectively) in IRF4 high but not in IRF4 low tumors. As radioactive glucose is the cornerstone of PET assessment, we investigated the correlation between baseline SUV values and IRF4 expression. Remarkably, we found significantly higher levels of FDG uptake in FL IRF4 high compared to IRF4 low cases (p= 0.003), supporting the glycolytic features of Iymphoma expressing IRF4. To define the metabolic activity and function of lymphoma cells with different IRF4 status, we performed seahorse assay in TMD8 and SUDHL2 cells (lymphoma cells with IRF4 high expression) transfected with siIRF4 or siCtr for 72 hours. In line with the IRF4-dependence on glycolysis, TMD8 transfected with siCtr had a low baseline glycolytic ECAR, while the same value doubled after silencing IRF4. Notably, after adding oligomycin, which stimulates glycolysis, TMD8 control cells showed a lower glycolytic capability compared with those deprived of IRF4, probably due to the already maximal addiction to glycolysis ( Fig 1B left). We then investigated the glycolytic demand of germinal center (GC) B cells differently expressing IRF4. Using irf4 fl/fl mice ( Klein et al, Nat Immunol 2006) crossed with Cγ 1cre animals ( Cγ 1cre; irf4 fl/fl) to induce conditional deletion of irf4 in GC B cells, we tested ex-vivo sorted GC B cells from Cγ 1cre; irf4 fl/fl, Cγ 1cre; irf4 fl/+ and WT mice immunized for 10 days. This time point is critical for studying IRF4 as it corresponds to the GC reaction peak when GC B cells express elevated IRF4 levels promoting terminal differentiation to plasma cells. As expected, GC B cells minimally relied on glycolysis and showed a low ability to respond to oligomycin in WT mice ( Weiser et al, Nat Immunol 2020). However, GC B cells with partial or complete loss of irf4 demonstrated a superior ability to response to increased glycolytic demand ( Fig 1B right), further supporting the conclusion that IRF4 influences B cell dependency on glycolysis. As the malignant cell metabolism can alter nutrient availability in the TME, we explored if lymphoma cells differently expressing IRF4 may change glucose levels in the media and in turn impact T cells activity. We found low glucose and high lactate levels in the media of cell lines with IRF4 high expression, while the opposite was observed for those that were IRF4 low. We then cocultured primary human CD4 + T cells with TMD8 cells transfected with either siIRF4 or siCtr for 3 days at three different concentrations of D-glucose: high (22.2 mM, Gluc hi), normal (5.5 mM, Gluc nor) or absent (0 mM, Gluc nil). In line with our prior findings, coculture of B cells in which IRF4 was knocked out showed a significant decrease in abundance of T reg cells and an increase in T FH cells compared to control in presence of Gluc hi or Gluc nor. In contrast, there was no difference in the overall amount of either T cell subtype irrespective of IRF4 status with Gluc nil, supporting the prerequisite of microenvironmental glucose as primary fuel for T cell survival and function. In conclusion, IRF4 rewires the metabolism in B cells with profound functional implications on the TME. Our data suggest that glucose metabolism represents a functional vulnerability that might be exploited therapeutically.

Warburg-Effect Driven Lactic Acidosis Is Associated with Early Death in Patients with Lymphoma

Introduction: Lactic acidosis is a medical emergency and in most cases results from an imbalance between tissue oxygenation and oxygen demand (Type A). However, Type B lactic acidosis occurs in the absence of a recognizable impairment in systemic oxygen delivery, and is most commonly associated with drug or toxin ingestion, sepsis, severe nutritional deficits such as thiamine, and less commonly, malignancy. The capacity of shifting glucose metabolism to anaerobic pathway despite the presence of adequate oxygen, enhancing the rate of glucose uptake, accelerated glycolysis and hence producing more lactate, is a well-defined phenomenon in malignant cell metabolism, and is called the Warburg Effect (WE) or aerobic glycolysis. A systemic rise in lactate and lactic acidosis has been reported in hematologic malignancies and less commonly in solid tumors, and is thought to occur as a consequence of both increased production of lactate- reflecting high malignancy burden, and impaired lactate turnover or elimination by liver or kidney. Few case reports or series have demonstrated a possible correlation between early death and elevated lactate in patients with hematologic malignancy (Table 1). We aimed to characterize the incidence and clinical impact of Warburg-driven lactic acidosis on outcome including early death. Methods: A retrospective study of patients admitted to the haemato-oncology department in a large tertiary medical center between the years 2013- 2022, that presented with elevated lactate levels during the first week of admission. Patients with hematologic diseases other than lymphoma were excluded. Of 40 patients reviewed, 21 patients were excluded because of underlying hematological malignancy other than lymphoma. Medical records were retrieved and reviewed for a recognizable cause for elevated lactate, or non (assigned as the WE group). Early-death was defined as death of any cause within 4 weeks of admission. Results: Nineteen patients had lymphoma and elevated lactate levels within the first week of admission. Of these, 12 patients fulfilled the criteria for WE and 7 patients had their high lactate attributed to other causes such as hemorrhagic shock (n=1), sepsis (n=5) and concomitant metformin use (n=1). Median age among patients with WE was 72.5 years (range 66-86) with 41.6% (n=5) males. All but one had aggressive lymphoma. Diffuse Large B cell Lymphoma was the most frequent type (n=9) followed by T cell Lymphoma (n=2) and follicular lymphoma (n=1). Most patients had advanced disease (Stage III-IV in 75%) and the median International Prognostic Index (IPI) of the patients was 4 (range 3-5). The median lactate level was 31.5 mg/dL (range 25-64; with upper limit of normal value-20 mg/dL). Two patients received thiamin therapy once WE was recognized. Early-death rate was 75% (9 patients) among patients with WE. The established cause of death in this population was: sepsis (n=3), DIC (n=1), uremia and multi organ failure (n=2) and respiratory failure due to lung infiltration by lymphoma (n=3). Both steroid and chemotherapy were initiated earlier in surviving patients as compared to those who died early (within 4 weeks of admission); in surviving patients, chemotherapy was initiated at a median of 3 days before first elevated lactate was detected (range (-)7-1), and steroids at median of 3 days before first elevated lactate was detected, while in those who died early, chemotherapy was initiated at median of 5 days after first elevated lactate was detected (range (-)2-18) and steroids at median 1 day after first elevated lactate (p=0.29 and 0.05 for steroids and chemo, respectively). (Table 2). Conclusion: Warburg driven lactic acidosis is a marker of ‘acute cancer’ in lymphoma patients and is associated with a very high mortality rate. Early recognition of this very high-risk clinical scenario is of utmost importance and should prompt immediate anti-lymphoma therapy that may improve outcome.

Poly(ADP-Ribose) Glycohydrolase (PARG) Removes Repressive Poly-ADP Ribose Marks from DNA Polymerase Theta (PolΘ) to Stimulate DNA Double-Strand Breaks Repair in Myeloid Malignancies

Myeloid malignant cells (AML, MPN, CML) expressing oncogenic tyrosine kinases [OTKs: FLT3(ITD), JAK2(V617F), BCR/ABL1, respectively] accumulate DNA damage but altered DNA repair mechanisms protect them from apoptosis. We reported before that formaldehyde generated by altered serine/one-carbon cycle metabolism in malignant cells harboring OTKs contributed to accumulation of toxic DNA-protein crosslinks (DPCs) which were converted to highly lethal DNA double-strand breaks (DSBs). To counteract the toxicity of DSBs, OTKs enhanced the expression of DNA polymerase theta (PolΘ, encoded by POLQ gene), a unique DNA helicase-DNA polymerase fusion protein that promotes error-prone repair of DSBs by a mechanism referred to as PolΘ-mediated DNA end-joining (TMEJ). PolΘ plays an essential role in the initiation and maintenance of hematological malignancies harboring OTKs. Although the cellular activities of PolΘ have been widely studied, little is known about how PolΘ is regulated at the molecular level. For example, whether post-translational modifications of PolΘ are important for its TMEJ activity and regulation is unknown. Poly(ADP-ribose) polymerase 1 (PARP1) dependent poly-(ADP)-ribosylation (PARylation) of proteins is one of the major post-translational modification events involved in the DNA damage response (DDR). PARP1 - the founding member of the poly(ADP-ribose) polymerase family -transfers adenosine diphosphate (ADP)-ribose from nicotinamide adenine dinucleotide (NAD +) to substrate proteins enabling their PARylation. PARylation is tightly controlled by the glycohydrolase activity of poly(ADP-ribose) glycohydrolase (PARG). Thus, interplay between PARP1 and PARG is thought to regulate the PARylation status of relevant DDR proteins and multiple studies demonstrated that both PARP1 and PARG contribute to DDR. Here, we investigated whether the interplay of PARP1 and PARG is important for the regulation of TMEJ and the specific activities of PolΘ in myeloid malignancies harboring OTKs. We found that upon DNA damage OTKs exerted temporal effect on PolΘ, PARP1 and PARG detection in chromatin fractions. While both PolΘ and PARG displayed continuous time-dependent accumulation during 120 min. after irradiation, PARP1 levels sharply increased at 20 min. and dissipated at 120 min. We find that PARP1 binds to and directly PARylates PolΘ in vitro and in cells. However, PARylated PolΘ is unable to perform TMEJ in vitro due to its inability to bind DNA despite its PARylation dependent recruitment to DNA. Hence, PARG is needed to reactivate PolΘ DNA binding and its TMEJ activity by removing repressive PAR marks on PolΘ. In support of this, cellular studies show that PARG is essential for TMEJ and support a two-step mechanism of a PARP1-PARG regulatory axis of PolΘ and TMEJ. In the first step, the rate of PARP1-PolΘ spatiotemporal recruitment to DNA damage foci supports a rapid step whereby PARP1 PARylates PolΘ and facilitates its recruitment to the vicinity of DNA damage in an inactive state. The rate of PARG spatiotemporal recruitment supports a second step whereby subsequent recruitment of PARG corresponds to dissipation of PARP1 and PAR at DNA damage sites, dePARylation of PolΘ, and activation of TMEJ. These studies elucidate an unprecedented mechanism of PARP1-PARG activation of TMEJ and reveal the molecular basis by which PARP1 and PARG inhibition suppresses TMEJ. In conclusion, using intracellular and biochemical approaches we show here that both PARP1 and PARG are required for TMEJ despite their counteracting enzymatic activities. We also pinpointed a unique dynamic spatiotemporal interplay between PolΘ, PARP1 and PARG to regulate TMEJ activity. During the initial stage, PARP1 activated by DNA damage PARylates PolΘ and facilitates its recruitment to the vicinity of DNA damage. PARG removal of PAR from PolΘ reactivated its interaction with DNA resulting in robust TMEJ activity in vitro and in cells. Thus, DNA/chromatin binding and TMEJ activity of PolΘ were restored upon PARG-mediated removal of PAR repressive marks on PolΘ. This process seems to play a critical role in protecting OTK-positive hematopoietic malignant cells from genotoxic effect of formaldehyde generated by altered serine/one-carbon cycle metabolism.

Integrated regulatory and metabolic networks of the tumor microenvironment for therapeutic target prioritization.

Translation of genomic discovery, such as single-cell sequencing data, to clinical decisions remains a longstanding bottleneck in the field. Meanwhile, computational systems biological models, such as cellular metabolism models and cell signaling pathways, have emerged as powerful approaches to provide efficient predictions in metabolites and gene expression levels, respectively. However, there has been limited research on the integration between these two models. This work develops a methodology for integrating computational models of probabilistic gene regulatory networks with a constraint-based metabolism model. By using probabilistic reasoning with Bayesian Networks, we aim to predict cell-specific changes under different interventions, which are embedded into the constraint-based models of metabolism. Applications to single-cell sequencing data of glioblastoma brain tumors generate predictions about the effects of pharmaceutical interventions on the regulatory network and downstream metabolisms in different cell types from the tumor microenvironment. The model presents possible insights into treatments that could potentially suppress anaerobic metabolism in malignant cells with minimal impact on other cell types' metabolism. The proposed integrated model can guide therapeutic target prioritization, the formulation of combination therapies, and future drug discovery. This model integration framework is also generalizable to other applications, such as different cell types, organisms, and diseases.

Metabolic Tumor Imaging with Rapidly Signal-Enhanced 1-13 C-Pyruvate-d3.

The metabolism of malignant cells differs significantly from that of healthy cells and thus, it is possible to perform metabolic imaging to reveal not only the exact location of a tumor, but also intratumoral areas of high metabolic activity. Herein, we demonstrate the feasibility of metabolic tumor imaging using signal-enhanced 1-13 C-pyruvate-d3 , which is rapidly enhanced via para-hydrogen, and thus, the signal is amplified by several orders of magnitudes in less than a minute. Using as a model, human melanoma xenografts injected with signal-enhanced 1-13 C-pyruvate-d3, we show that the conversion of pyruvate into lactate can be monitored along with its kinetics, which could pave the way for rapidly detecting and monitoring changes in tumor metabolism.

STAT3 as a mediator of oncogenic cellular metabolism: Pathogenic and therapeutic implications.

The oncogenic transcription factor signal transducer and activator of transcription 3 (STAT3) is activated constitutively in a wide array of human cancers. It is an appealing molecular target for novel therapy as it directly regulates expression of genes involved in cell proliferation, survival, angiogenesis, chemoresistance and immune responsiveness. In addition to these well-established oncogenic roles, STAT3 has also been found to mediate a wide array of functions in modulating cellular behavior. The transcriptional function of STAT3 is canonically regulated through tyrosine phosphorylation. However, STAT3 phosphorylated at a single serine residue can allow incorporation of this protein into the inner mitochondrial membrane to support oxidative phosphorylation (OXPHOS) and maximize the utility of glucose sources. Conflictingly, its canonical transcriptional activity suppresses OXPHOS and favors aerobic glycolysis to promote oncogenic behavior. Apart from mediating the energy metabolism and controversial effects on ATP production, STAT3 signaling modulates lipid metabolism of cancer cells. By mediating fatty acid synthesis and beta oxidation, STAT3 promotes employment of available resources and supports survival in the conditions of metabolic stress. Thus, the functions of STAT3 extend beyond regulation of oncogenic genes expression to pleiotropic effects on a spectrum of essential cellular processes. In this review, we dissect the current knowledge on activity and mechanisms of STAT3 involvement in transcriptional regulation, mitochondrial function, energy production and lipid metabolism of malignant cells, and its implications to cancer pathogenesis and therapy.

Iron Metabolism in the Tumor Microenvironment: Contributions of Innate Immune Cells.

Cells of the innate immune system are a major component of the tumor microenvironment. They play complex and multifaceted roles in the regulation of cancer initiation, growth, metastasis and responses to therapeutics. Innate immune cells like neutrophils and macrophages are recruited to cancerous tissues by chemotactic molecules released by cancer cells and cancer-associated stromal cells. Once they reach the tumor, they can be instructed by a network of proteins, nucleic acids and metabolites to exert protumoral or antitumoral functions. Altered iron metabolism is a feature of cancer. Epidemiological studies suggest that increased presence of iron and/or iron binding proteins is associated with increased risks of cancer development. It has been shown that iron metabolism is involved in shaping the immune landscapes in inflammatory/infectious diseases and cancer-associated inflammation. In this article, we will dissect the contribution of macrophages and neutrophils to dysregulated iron metabolism in malignant cells and its impact on cancer growth and metastasis. The mechanisms involved in regulating the actions of macrophages and neutrophils will also be discussed. Moreover, we will examine the effects of iron metabolism on the phenotypes of innate immune cells. Both iron chelating and overloading agents are being explored in cancer treatment. This review highlights alternative strategies for management of iron content in cancer cells by targeting the iron donation and modulation properties of macrophages and neutrophils in the tumor microenvironment.

The Potential Oncostatic Effects of Melatonin against Prostate Cancer.

Melatonin is an endogenous indolamine, synthesized and secreted from the pineal gland. The environmental light-dark cycle is the primary regulator of melatonin synthesis. Darkness during the subjective night induces noradrenaline secretion, which stimulates pinealocytes for melatonin production. Melatonin exhibits anticancer effects and different physiological functions through the membrane-bound G-protein-coupled MT1 and MT2 receptors. Impaired circadian activity, indoor or outdoor light pollution, shift work, night work, and jet lag suppress normal melatonin synthesis. Decreased melatonin concentration causes impaired anticancer effects that adversely affect the progression of different cancers, including prostate. Melatonin differentially regulates the cell cycle, cell survival, and metabolism in malignant cells in contrast to normal prostate epithelial cells. Melatonin promotes the nuclear exclusion of androgen receptors without suppressing the expression of this receptor. This indirect effect blocks the androgenic response in prostate cancer cells. It acts as a cytostatic and cytotoxic agent, prevents cell proliferation, and activates an apoptotic response. Melatonin also inhibits HIF-1α activity and the expression of vascular endothelial growth factors to suppress angiogenesis. This indolamine restricts alteration of metabolic activity, invasion, and metastasis. Melatonin has therapeutic importance. It decreases the side effects of anticancer treatment and mitigates adverse effects after prostatectomy and radiotherapy. Melatonin blocks the recurrence of prostate cancer as well as hormone-refractory effects during androgen deprivation therapy. The present review discusses the multifaceted effects of melatonin against prostate cancer.

The design and fabrication of nanoengineered platinum needles with laser welded carbon nanotubes (CNTs) for the electrochemical biosensing of cancer lymph nodes.

A new biosensor for detecting cancer involved sentinel lymph nodes has been developed via the electrochemical tracing of fatty acid oxidation as a distinct metabolism of malignant cells invading lymph nodes (LNs). The system included integrated platinum needle electrodes that were decorated by carbon nanotubes (as hydrophobic agents) through laser-assisted nanowelding. It was applied to record the dielectric spectroscopy data from LN contents via electrochemical impedance spectroscopy. The system was applied for dielectric spectroscopy of LN contentsviaelectrochemical impedance approach. The reduced lipid content of involved LNs, due to fat metabolism by invasive cancer cells, would decrease the charge transfer resistance (RCT) of the LNs with respect to their normal counterparts. Multi-walled carbon nanotubes (MWCNTs) with superhydrophobic properties were used to enhance the interaction of Pt needle electrodes with the lipidic contents of lymph nodes. This is the first time that a fatty acid metabolism-based sensing approach has been introduced to detect involved LNs. Moreover, a novel electrode decorating method was applied to enhance the interfacial contact of this lipid detection probe (LDP). In order to avoid doubt about the biocompatibility of ferrocyanide, [Fe(CN)6]4- and ferricyanide, [Fe(CN)6]3-, a biocompatible injectable metal ion-based material, ferric carboxymaltose, was selected and applied as the electrolyte for the first time. Rabbit LNs were tested using the LDP in the animal model phase. The system was then used in vitro on 122 dissected human LNs in the operating room. Calibration of the results showed an excellent match between the dielectric response of the LDP (known as charge transfer resistance (RCT)) and the final pathological diagnoses. The LDP may have a promising future after further clinical investigations for intra-operative distinction between normal and cancerous LNs.

Metabolic reprogramming in childhood acute lymphoblastic leukemia.

The first observations of altered metabolism in malignant cells were made nearly 100 years ago and therapeutic strategies targeting cell metabolism have been in clinical use for several decades. In this review, we summarize our current understanding of cell metabolism dysregulation in childhood acute lymphoblastic leukemia (cALL). Reprogramming of cellular bioenergetic processes can be expected in the three distinct stages of cALL: at diagnosis, during standard chemotherapy, and in cases of relapse. Upregulation of glycolysis, dependency on anaplerotic energy sources, and activation of the electron transport chain have all been observed in cALL. While the current treatment strategies are tackling some of these aberrations, cALL cells are likely to be able to rewire their metabolism in order to escape therapy, which may contribute to a refractory disease and relapse. Finally, here we focus on novel therapeutic approaches emerging from our evolving understanding of the alterations of different metabolic networks in lymphoblasts.

Cancer associated fibroblast FAK regulates malignant cell metabolism

Emerging evidence suggests that cancer cell metabolism can be regulated by cancer-associated fibroblasts (CAFs), but the mechanisms are poorly defined. Here we show that CAFs regulate malignant cell metabolism through pathways under the control of FAK. In breast and pancreatic cancer patients we find that low FAK expression, specifically in the stromal compartment, predicts reduced overall survival. In mice, depletion of FAK in a subpopulation of CAFs regulates paracrine signals that increase malignant cell glycolysis and tumour growth. Proteomic and phosphoproteomic analysis in our mouse model identifies metabolic alterations which are reflected at the transcriptomic level in patients with low stromal FAK. Mechanistically we demonstrate that FAK-depletion in CAFs increases chemokine production, which via CCR1/CCR2 on cancer cells, activate protein kinase A, leading to enhanced malignant cell glycolysis. Our data uncover mechanisms whereby stromal fibroblasts regulate cancer cell metabolism independent of genetic mutations in cancer cells.

Microbial enzymes for deprivation of amino acid metabolism in malignant cells: biological strategy for cancer treatment.

Amino acid deprivation therapy (AADT) is emerging as a promising strategy for the development of novel therapeutics against cancer. This biological therapy relies upon the differences in the metabolism of cancer and normal cells. The rapid growth of tumors results in decreased expression of certain enzymes leading to auxotrophy for some specific amino acids. These auxotrophic tumors are targeted by amino acid-depleting enzymes. The depletion of amino acid selectively inhibits tumor growth as the normal cells can synthesize amino acids by their usual machinery. The enzymes used in AADT are mostly obtained from microbes for their easy availability. Microbial L-asparaginase is already approved by FDA for the treatment of acute lymphoblastic leukemia. Arginine deiminase and methionase are under clinical trials and the therapeutic potential of lysine oxidase, glutaminase and phenylalanine ammonia lyase is also being explored. The present review provides an overview of microbial amino acid depriving enzymes. Various attributes of these enzymes like structure, mode of action, production, formulations, and targeted cancers are discussed. The challenges faced and the combat strategies to establish AADT in standard cancer armamentarium are also reviewed.Key Points • Amino acid deprivation therapy is a potential therapy for auxotrophic tumors. • Microbial enzymes are used due to their ease of manipulation and high productivity. • Enzyme properties are improved by PEGylation, encapsulation, and genetic engineering. • AADT can be employed as combinational therapy for better containment of cancer.

Activity and Trafficking of Copper-Transporting ATPases in Tumor Development and Defense against Platinum-Based Drugs.

Membrane trafficking pathways emanating from the Golgi regulate a wide range of cellular processes. One of these is the maintenance of copper (Cu) homeostasis operated by the Golgi-localized Cu-transporting ATPases ATP7A and ATP7B. At the Golgi, these proteins supply Cu to newly synthesized enzymes which use this metal as a cofactor to catalyze a number of vitally important biochemical reactions. However, in response to elevated Cu, the Golgi exports ATP7A/B to post-Golgi sites where they promote sequestration and efflux of excess Cu to limit its potential toxicity. Growing tumors actively consume Cu and employ ATP7A/B to regulate the availability of this metal for oncogenic enzymes such as LOX and LOX-like proteins, which confer higher invasiveness to malignant cells. Furthermore, ATP7A/B activity and trafficking allow tumor cells to detoxify platinum (Pt)-based drugs (like cisplatin), which are used for the chemotherapy of different solid tumors. Despite these noted activities of ATP7A/B that favor oncogenic processes, the mechanisms that regulate the expression and trafficking of Cu ATPases in malignant cells are far from being completely understood. This review summarizes current data on the role of ATP7A/B in the regulation of Cu and Pt metabolism in malignant cells and outlines questions and challenges that should be addressed to understand how ATP7A and ATP7B trafficking mechanisms might be targeted to counteract tumor development.

Amino Acid Degrading Enzymes and their Application in Cancer Therapy.

Amino acids are essential components in various biochemical pathways. The deprivation of certain amino acids is an antimetabolite strategy for the treatment of amino acid-dependent cancers which exploits the compromised metabolism of malignant cells. Several studies have focused on the development and preclinical and clinical evaluation of amino acid degrading enzymes, namely L-asparaginase, L-methionine γ-lyase, L-arginine deiminase, L-lysine α-oxidase. Further research into cancer cell metabolism may therefore define possible targets for controlling tumor growth. The purpose of this review was to summarize recent progress in the relationship between amino acids metabolism and cancer therapy, with a particular focus on Lasparagine, L-methionine, L-arginine and L-lysine degrading enzymes and their formulations, which have been successfully used in the treatment of several types of cancer. We carried out a structured search among literature regarding to amino acid degrading enzymes. The main aspects of search were in vitro and in vivo studies, clinical trials concerning application of these enzymes in oncology. Most published research are on the subject of L-asparaginase properties and it's use for cancer treatment. L-arginine deiminase has shown promising results in a phase II trial in advanced melanoma and hepatocellular carcinoma. Other enzymes, in particular Lmethionine γ-lyase and L-lysine α-oxidase, were effective in vitro and in vivo. The findings of this review revealed that therapy based on amino acid depletion may have the potential application for cancer treatment but further clinical investigations are required to provide the efficacy and safety of these agents.

Time-dependent cytotoxicity of dichloroacetate and metformin against Lewis lung carcinoma

To study time-dependent cytotoxicity of sodium dichloroacetate (DCA) and metformin (MTF) against metastatic tumor cells and action of these agents on tumor cell migration. In the study low metastatic LLC/R9variant of Lewis lung carcinoma was used. The number of living cells in the cytotoxic test was evaluated using sulforhodamine B after 1, 2and 3days of cell incubation in vitro. The parameters of the sensitivity of tumor cells to the action of DCA and MTF in vitro were calculated using nonlinear and linear regression of experimental data. The effect of DCA and MTF on cellular motility in vitro was evaluated using a Boyden chamber by calculation of the number of cells that migrated to the bottom side of the filter within 3days of incubation. The statistical analysis of the data was carried out with the use of descriptive methods, Student's t-criterion, nonlinear, and linear regression analysis. IC50of DCA was found to be equal to 50.8±7.6mM at the first day of incubation with LLC/R9cells and decreased by 1.9 (p < 0.05) and 2.1 (p < 0.05) times at the 2nd and 3rd days, respectively. Despite the almost identical ІC50at the 2nd and 3rd days, an increase in the incubation period of cells with DCA for up to 3days increased the C0parameter, which reflects the maximum concentration of the agent that does not exhibit cytotoxic effects, by 93% (p < 0.05) compared to this at the 2nd day (16.2 ± 1.4mM vs 8.4 ± 1.0mM, correspondently). Unlike DCA, the LLC/R9cell population was not homogeneous by the sensitivity to the action of MTF; at least at the 3rd day, an appearance of MTF-resistant subpopulation was observed, accounting for 35% of all cells. IC50of MTF was equal to 12.1 ± 0.6mM, and unlike DCA, this index progressively decreased at the 2nd and 3rd days by 1.4 (p < 0.05) and 9.3times (p < 0.05) respectively. Action of DCA at a concentration of 25mM alone and in combination with MTF at the concentrations of 0.1mM and 0.7mM resulted in an increase in cell migration by 65% (p < 0.05), 63% (p < 0.05) and 78.5% (p < 0.05), respectively. There was no significant effect of MTF on the tumor cell migration. The sensitivity of the metastatic Lewis lung carcinoma cells to the action of the modifiers of the energy metabolism increased significantly with an increase in the incubation period, apparently, primarily due to the shortage of nutrient substrates and, in particular, glucose, indicating the relevance of their combined use as well as with other agents, which promote the deficiency of glucose in the tumor microenvironment.

Disulfide-masked iron prochelators: Effects on cell death, proliferation, and hemoglobin production

The iron metabolism of malignant cells, which is altered to ensure higher acquisition and utilization, motivates the investigation of iron chelation strategies in cancer treatment. In a prochelation approach aimed at increasing intracellular specificity, disulfide reduction/activation switches are incorporated on iron-binding scaffolds resulting in intracellularly activated scavengers. Herein, this strategy is applied to several tridentate donor sets including thiosemicarbazones, aroylhydrazones and semicarbazones. The novel prochelator systems are antiproliferative in breast adenocarcinoma cell lines (MCF-7 and metastatic MDA-MB-231) and do not result in the intracellular generation of oxidative stress. Consistent with iron deprivation, the tested prochelators lead to cell-cycle arrest at the G1/S interface and induction of apoptosis. Notably, although hemoglobin-synthesizing blood cells have the highest iron need in the human body, no significant impact on hemoglobin production was observed in the MEL (murine erythroleukemia) model of differentiating erythroid cells. This study provides new information on the intracellular effects of disulfide-based prochelators and indicates aroylhydrazone (AH1-S)2 as a promising prototype of a new class of antiproliferative prochelator systems.

Abstract 3807: Histone deacetylase inhibitors repress hypoxia signaling through affecting Hsp90

Abstract Histone deacetylase inhibitors (HDACis) are among the most promising recently developed anti-cancer agents. Recent work suggests that HDACis suppress both malignant cell metabolism and progression by interfering with the hypoxia-inducible factor (HIF) signaling, which plays a key role in tumor progression in both actual and falsely perceived cellular hypoxia (pseudohypoxia). However, the precise biochemical mechanism of HDACis repression of HIFs function remains unclear. In this study, we demonstrated that the class-2 histone deacetylase inhibitor SAHA potently inhibits hypoxia signaling via interfering with heat shock protein 90 (Hsp90). HDACis increase in the acetylation of Hsp90, resulting in less HIF-α recognition and nuclear translocation. Accumulated cytoplasmic HIF-α is not transcriptionally active and degradaded through proteosomal pathway. Finally, we demonstrated that SAHA remarkably reduces hypoxia signaling and decreases tumor growth in vivo. These findings provide insight into new possible therapeutic strategies for using HDACis in tumors with known HIF pathway aberrations. Citation Format: Chunzhang Yang, Zhengping Zhuang. Histone deacetylase inhibitors repress hypoxia signaling through affecting Hsp90. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3807.