RESUMO
The primary impediment to the success of immunotherapy lies in the immune evasion orchestrated by tumors, contributing to the suboptimal overall response rates observed. Despite this recognition, the intricacies of the underlying mechanisms remain incompletely understood. Through preliminary detection of clinical patient tissues, we have found that ALDH1A1 was a key gene for the prognosis of cancer patients and tumor glycolysis. In vitro experiments and tumor formation in nude mice suggested that targeting ALDH1A1 could inhibit tumor growth. Through further analysis of xenograft tumor models in immune-normal mice and flow cytometry, we found that deficiency in ALDH1A1 could promote immune system suppression of tumors in vivo. Specifically, RNA-seq analysis, combined with qPCR and western blot, identified the transcription factor ZBTB7B as downstream of ALDH1A1. The binding sites of the transcription factor ZBTB7B on the LDHA promoter region, which is responsible for regulating the rate-limiting enzyme gene LDHA in glycolysis, were determined using luciferase reporter gene detection and Chip-qPCR, respectively. In addition, the increased SUMOylation of ZBTB7B stabilized its transcriptional activity. Further in vivo and in vitro experiments confirmed that the combination of targeting ALDH1A1 and ZBTB7B with immune checkpoint inhibitors could synergistically inhibit tumors in vivo. Finally, after conducting additional verification of patient tissue and clinical data, we have confirmed the potential translational value of targeting ALDH1A1 and ZBTB7B for tumor immunotherapy. These results emphasize the potential translational significance of targeting ALDH1A1 and ZBTB7B in the realm of tumor immunotherapy. The convergence of ALDH1A1 inhibition and immune checkpoint blockade, particularly with PD-L1/PD-1 mAb, presents a compelling avenue for curtailing tumor immune escape.
Assuntos
Família Aldeído Desidrogenase 1 , Glicólise , Camundongos Nus , Retinal Desidrogenase , Evasão Tumoral , Humanos , Animais , Família Aldeído Desidrogenase 1/metabolismo , Família Aldeído Desidrogenase 1/genética , Retinal Desidrogenase/metabolismo , Retinal Desidrogenase/genética , Camundongos , Linhagem Celular Tumoral , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias/imunologia , Neoplasias/genética , Neoplasias/patologia , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Regiões Promotoras Genéticas/genética , L-Lactato Desidrogenase/metabolismo , L-Lactato Desidrogenase/genética , Feminino , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Homeobox C4 (HOXC4) is a member of homeobox family and acts as a transcription factor in regulating morphological development. The current study aimed to determine its role in pancreatic cancer (PC). Bioinformatics analysis was employed to assess the expression and clinical significance of HOXC4 in PC, while the expression of HOXC4 was further confirmed in PC tissues through quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry (IHC). The impact of HOXC4 on PC cell proliferation was evaluated using various assays including Cell Counting Kit-8, colony formation, apoptosis detection, cell cycle analysis, and subcutaneous tumorigenesis. Extracellular acidification rate, glucose uptake, and lactate production measurements were detected to examine the impact of HOXC4 on glycolysis. The relationship between HOXC4 and lactate dehydrogenase A (LDHA) was investigated using CHIP assay, luciferase reporter assay, and western blot. Notably, there was a substantial increase in HOXC4 expression in PC, and patients with elevated HOXC4 levels exhibited shorter survival durations. HOXC4 knockdown resulted in significantly reduced proliferation and colony formation in PC cells, accompanied by increased apoptosis and G1 phase arrest. The overexpression of HOXC4 resulted in contrasting effects. In vivo, the proliferation of PC cells was diminished upon the knockdown of HOXC4. HOXC4 exhibited an increase in LDHA expression by binding to its promoter. The suppressive effects of HOXC4 knockdown on PC cells were counteracted upon the restoration of LDHA. In conclusion, HOXC4 promoted the proliferation of PC cells by increasing LDHA-mediated glycolysis. HOXC4 can act as a target for PC therapy.
Assuntos
Proliferação de Células , Glicólise , Proteínas de Homeodomínio , Neoplasias Pancreáticas , Humanos , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Proliferação de Células/genética , Glicólise/genética , Linhagem Celular Tumoral , Animais , Camundongos , Regulação Neoplásica da Expressão Gênica , L-Lactato Desidrogenase/metabolismo , L-Lactato Desidrogenase/genética , Apoptose/genética , Masculino , Feminino , Camundongos NusRESUMO
Background: Mitochondrial creatine kinase (MtCK) plays a pivotal role in cellular energy metabolism, exhibiting enhanced expression in various tumors, including colorectal cancer (CRC). Creatine kinase mitochondrial 2 (CKMT2) is a subtype of MtCK; however, its clinical significance, biological functions, and underlying molecular mechanisms in CRC remain elusive. Methods: We employed immunohistochemical staining to discern the expression of CKMT2 in CRC and adjacent nontumor tissues of patients. The correlation between CKMT2 levels and clinical pathological factors was assessed. Additionally, we evaluated the association between CKMT2 and the prognosis of CRC patients using Kaplan-Meier survival curves and Cox regression analysis. Meanwhile, quantitative reverse transcription polymerase chain reaction (qRT-PCR) was used to detect the expression levels of CKMT2 in different CRC cell lines. Finally, we explored the biological functions and potential molecular mechanisms of CKMT2 in CRC cells through various techniques, including qRT-PCR, cell culture, cell transfection, western blot, Transwell chamber assays, flow cytometry, and co-immunoprecipitation. Results: We found that CKMT2 was significantly overexpressed in CRC tissues compared with adjacent nontumor tissues. The expression of CKMT2 is correlated with pathological types, tumor size, distant metastasis, and survival in CRC patients. Importantly, CKMT2 emerged as an independent prognostic factor through Cox regression analysis. Experimental downregulation of CKMT2 expression in CRC cell lines inhibited the migration and promoted apoptosis of these cells. Furthermore, we identified a novel role for CKMT2 in promoting aerobic glycolysis in CRC cells through interaction with lactate dehydrogenase B (LDHB). Conclusion: In this study, we found the elevated expression of CKMT2 in CRC, and it was a robust prognostic indicator in CRC patients. CKMT2 regulates glucose metabolism via amplifying the Warburg effect through interaction with LDHB, which promotes the growth and progression of CRC. These insights unveil a novel regulatory mechanism by which CKMT2 influences CRC and provide promising targets for future CRC therapeutic interventions.
Assuntos
Neoplasias Colorretais , Efeito Warburg em Oncologia , Humanos , Neoplasias Colorretais/patologia , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/mortalidade , Masculino , Feminino , Linhagem Celular Tumoral , Prognóstico , Creatina Quinase Mitocondrial/metabolismo , Creatina Quinase Mitocondrial/genética , Progressão da Doença , L-Lactato Desidrogenase/metabolismo , L-Lactato Desidrogenase/genética , Pessoa de Meia-Idade , Proliferação de Células , Apoptose , Regulação Neoplásica da Expressão GênicaRESUMO
The Warburg effect is a hallmark of cancer that refers to the preference of cancer cells to metabolize glucose anaerobically rather than aerobically1,2. This results in substantial accumulation of lacate, the end product of anaerobic glycolysis, in cancer cells3. However, how cancer metabolism affects chemotherapy response and DNA repair in general remains incompletely understood. Here we report that lactate-driven lactylation of NBS1 promotes homologous recombination (HR)-mediated DNA repair. Lactylation of NBS1 at lysine 388 (K388) is essential for MRE11-RAD50-NBS1 (MRN) complex formation and the accumulation of HR repair proteins at the sites of DNA double-strand breaks. Furthermore, we identify TIP60 as the NBS1 lysine lactyltransferase and the 'writer' of NBS1 K388 lactylation, and HDAC3 as the NBS1 de-lactylase. High levels of NBS1 K388 lactylation predict poor patient outcome of neoadjuvant chemotherapy, and lactate reduction using either genetic depletion of lactate dehydrogenase A (LDHA) or stiripentol, a lactate dehydrogenase A inhibitor used clinically for anti-epileptic treatment, inhibited NBS1 K388 lactylation, decreased DNA repair efficacy and overcame resistance to chemotherapy. In summary, our work identifies NBS1 lactylation as a critical mechanism for genome stability that contributes to chemotherapy resistance and identifies inhibition of lactate production as a promising therapeutic cancer strategy.
Assuntos
Proteínas de Ciclo Celular , Resistencia a Medicamentos Antineoplásicos , Ácido Láctico , Proteínas Nucleares , Reparo de DNA por Recombinação , Animais , Feminino , Humanos , Masculino , Camundongos , Hidrolases Anidrido Ácido/metabolismo , Anaerobiose , Proteínas de Ciclo Celular/química , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Quebras de DNA de Cadeia Dupla , Proteínas de Ligação a DNA/metabolismo , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Instabilidade Genômica , Ácido Láctico/metabolismo , Lisina/química , Lisina/metabolismo , Lisina Acetiltransferase 5/metabolismo , Lisina Acetiltransferase 5/genética , Proteína Homóloga a MRE11/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Neoplasias/genética , Proteínas Nucleares/química , Proteínas Nucleares/metabolismo , Organoides , Glicólise , Terapia Neoadjuvante , L-Lactato Desidrogenase/antagonistas & inibidores , L-Lactato Desidrogenase/deficiência , L-Lactato Desidrogenase/genética , L-Lactato Desidrogenase/metabolismo , Anticonvulsivantes/farmacologiaRESUMO
Breast cancer is the most prevalent malignancy and the most significant contributor to mortality in female oncology patients. Potassium Two Pore Domain Channel Subfamily K Member 1 (KCNK1) is differentially expressed in a variety of tumors, but the mechanism of its function in breast cancer is unknown. In this study, we found for the first time that KCNK1 was significantly up-regulated in human breast cancer and was correlated with poor prognosis in breast cancer patients. KCNK1 promoted breast cancer proliferation, invasion, and metastasis in vitro and vivo. Further studies unexpectedly revealed that KCNK1 increased the glycolysis and lactate production in breast cancer cells by binding to and activating lactate dehydrogenase A (LDHA), which promoted histones lysine lactylation to induce the expression of a series of downstream genes and LDHA itself. Notably, increased expression of LDHA served as a vicious positive feedback to reduce tumor cell stiffness and adhesion, which eventually resulted in the proliferation, invasion, and metastasis of breast cancer. In conclusion, our results suggest that KCNK1 may serve as a potential breast cancer biomarker, and deeper insight into the cancer-promoting mechanism of KCNK1 may uncover a novel therapeutic target for breast cancer treatment.
Assuntos
Neoplasias da Mama , Proliferação de Células , Histonas , Animais , Feminino , Humanos , Camundongos , Neoplasias da Mama/patologia , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica , Glicólise/genética , Histonas/metabolismo , L-Lactato Desidrogenase/metabolismo , L-Lactato Desidrogenase/genética , Lactato Desidrogenase 5/metabolismo , Lactato Desidrogenase 5/genética , Camundongos Endogâmicos BALB C , Camundongos Nus , Invasividade Neoplásica , Metástase Neoplásica , Canais de Potássio de Domínios Poros em Tandem/metabolismo , Canais de Potássio de Domínios Poros em Tandem/genética , Prognóstico , Regulação para Cima/genéticaRESUMO
Plasmodium knowlesi is the only Plasmodium that causes zoonotic disease among the Plasmodium that cause infection in humans. It is fatal due to its short asexual growth cycle within 24 h. Lactate dehydrogenase (LDH), an enzyme that catalyzes the final step of glycolysis, is a biomarker for diagnosing infection by Plasmodium spp. parasite. Therefore, this study aimed to efficiently produce the soluble form of P. knowlesi LDH (PkLDH) using a bacterial expression system for studying malaria caused by P. knowlesi. Recombinant pET-21a(+)-PkLDH plasmid was constructed by inserting the PkLDH gene into a pET-21a(+) expression vector. Subsequently, the recombinant plasmid was inserted into the protein-expressing Escherichia coli Rosetta(DE3) strain, and the optimal conditions for overexpression of the PkLDH protein were established using this strain. We obtained a yield of 52.0 mg/L PkLDH from the Rosetta(DE3) strain and confirmed an activity of 483.9 U/mg through experiments. This methodology for high-efficiency PkLDH production can be utilized for the development of diagnostic methods and drug candidates for distinguishing malaria caused by P. knowlesi.
Assuntos
Clonagem Molecular , L-Lactato Desidrogenase , Malária , Plasmodium knowlesi , Plasmodium knowlesi/genética , Plasmodium knowlesi/enzimologia , L-Lactato Desidrogenase/genética , L-Lactato Desidrogenase/metabolismo , Clonagem Molecular/métodos , Malária/parasitologia , Malária/diagnóstico , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Animais , Humanos , Expressão Gênica , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismoRESUMO
Liver fibrosis is a significant health concern globally due to its association with severe liver conditions like cirrhosis and liver cancer. Histone lactylation has been implicated in the progression of hepatic fibrosis, but its specific role in liver fibrosis, particularly regarding H3K18 lactylation, remained unclear. To investigate this, we established in vivo and in vitro models of liver fibrosis using carbon tetrachloride (CCl4) injection in rats and stimulation of hepatic stellate cells (HSCs) with TGF-ß1, respectively. We found that histone lactylation, particularly H3K18 lactylation, was upregulated in both CCl4-induced rats and TGF-ß1-activated HSCs, indicating its potential involvement in liver fibrosis. Further experiments revealed that lactate dehydrogenase A (LDHA) knockdown inhibited H3K18 lactylation and had a beneficial effect on liver fibrosis by suppressing HSC proliferation, migration, and extracellular matrix (ECM) deposition. This suggests that H3K18 lactylation promotes liver fibrosis progression. Chromatin immunoprecipitation (ChIP) and luciferase reporter assays demonstrated that H3K18 lactylation facilitated the transcription of SOX9, a transcription factor associated with fibrosis. Importantly, overexpression of SOX9 counteracted the effects of LDHA silencing on activated HSCs, indicating that SOX9 is downstream of H3K18 lactylation in promoting liver fibrosis. In summary, this study uncovers a novel mechanism by which H3K18 lactylation contributes to liver fibrosis by activating SOX9 transcription. This finding opens avenues for exploring new therapeutic strategies for hepatic fibrosis targeting histone lactylation pathways.
Assuntos
Progressão da Doença , Células Estreladas do Fígado , Histonas , Cirrose Hepática , Ratos Sprague-Dawley , Fatores de Transcrição SOX9 , Animais , Humanos , Masculino , Ratos , Tetracloreto de Carbono , Movimento Celular/genética , Proliferação de Células , Células Estreladas do Fígado/metabolismo , Células Estreladas do Fígado/patologia , Histonas/metabolismo , Histonas/genética , L-Lactato Desidrogenase/metabolismo , L-Lactato Desidrogenase/genética , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Cirrose Hepática/genética , Cirrose Hepática/induzido quimicamente , Fatores de Transcrição SOX9/metabolismo , Fatores de Transcrição SOX9/genética , Transcrição Gênica , Fator de Crescimento Transformador beta1/metabolismoRESUMO
AIMS: Despite the success of single-cell RNA sequencing in identifying cellular heterogeneity in ischemic stroke, clarifying the mechanisms underlying these associations of differently expressed genes remains challenging. Several studies that integrate gene expression and gene expression quantitative trait loci (eQTLs) with genome wide-association study (GWAS) data to determine their causal role have been proposed. METHODS: Here, we combined Mendelian randomization (MR) framework and single cell (sc) RNA sequencing to study how differently expressed genes (DEGs) mediating the effect of gene expression on ischemic stroke. The hub gene was further validated in the in vitro model. RESULTS: We identified 2339 DEGs in 10 cell clusters. Among these DEGs, 58 genes were associated with the risk of ischemic stroke. After external validation with eQTL dataset, lactate dehydrogenase B (LDHB) is identified to be positively associated with ischemic stroke. The expression of LDHB has also been validated in sc RNA-seq with dominant expression in microglia and astrocytes, and melatonin is able to reduce the LDHB expression and activity in vitro ischemic models. CONCLUSION: Our study identifies LDHB as a novel biomarker for ischemic stroke via combining the sc RNA-seq and MR analysis.
Assuntos
AVC Isquêmico , L-Lactato Desidrogenase , Melatonina , Análise da Randomização Mendeliana , Análise de Sequência de RNA , Animais , Humanos , Estudo de Associação Genômica Ampla/métodos , AVC Isquêmico/genética , AVC Isquêmico/metabolismo , Isoenzimas/genética , Isoenzimas/metabolismo , L-Lactato Desidrogenase/metabolismo , L-Lactato Desidrogenase/genética , Análise da Randomização Mendeliana/métodos , Locos de Características Quantitativas , Análise de Sequência de RNA/métodos , Análise de Célula Única/métodos , CamundongosRESUMO
BACKGROUND: Breast cancer (BC) is the most commonly diagnosed cancer in women. Treatment approaches that differ between estrogen-positive (ER+) and triple-negative BC cells (TNBCs) and may subsequently affect cancer biomarkers, such as H19 and telomerase, are an emanating delight in BC research. For instance, all-trans-Retinoic acid (ATRA) could represent a potent regulator of these oncogenes, regulating microRNAs, mostly let-7a microRNA (miR-let-7a), which targets the glycolysis pathway, mainly pyruvate kinase M2 (PKM2) and lactate dehydrogenase A (LDHA) enzymes. Here, we investigated the potential role of ATRA in H19, telomerase, miR-let-7a, and glycolytic enzymes modulation in ER + and TNBC cells. METHODS: MCF-7 and MDA-MB-231 cells were treated with 5 µM ATRA and/or 100 nM fulvestrant. Then, ATRA-treated or control MCF-7 cells were transfected with either H19 or hTERT siRNA. Afterward, ATRA-treated or untreated MDA-MB-231 cells were transfected with estrogen receptor alpha ER(α) or beta ER(ß) expression plasmids. RNA expression was evaluated by RTâqPCR, and proteins were assessed by Western blot. PKM2 activity was measured using an NADH/LDH coupled enzymatic assay, and telomerase activity was evaluated with a quantitative telomeric repeat amplification protocol assay. Student's t-test or one-way ANOVA was used to analyze data from replicates. RESULTS: Our results showed that MCF-7 cells were more responsive to ATRA than MDA-MB-231 cells. In MCF-7 cells, ATRA and/or fulvestrant decreased ER(α), H19, telomerase, PKM2, and LDHA, whereas ER(ß) and miR-let-7a increased. H19 or hTERT knockdown with or without ATRA treatment showed similar results to those obtained after ATRA treatment, and a potential interconnection between H19 and hTERT was found. However, in MDA-MB-231 cells, RNA expression of the aforementioned genes was modulated after ATRA and/or fulvestrant, with no significant effect on protein and activity levels. Overexpression of ER(α) or ER(ß) in MDA-MB-231 cells induced telomerase activity, PKM2 and LDHA expression, in which ATRA treatment combined with plasmid transfection decreased glycolytic enzyme expression. CONCLUSIONS: To the best of our knowledge, our study is the first to elucidate a new potential interaction between the estrogen receptor and glycolytic enzymes in ER + BC cells through miR-let-7a.
Assuntos
Neoplasias da Mama , Glicólise , MicroRNAs , RNA Longo não Codificante , Telomerase , Tretinoína , Humanos , Tretinoína/farmacologia , Glicólise/efeitos dos fármacos , Telomerase/metabolismo , Telomerase/genética , MicroRNAs/genética , MicroRNAs/metabolismo , Feminino , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Células MCF-7 , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Receptores de Estrogênio/metabolismo , L-Lactato Desidrogenase/metabolismo , L-Lactato Desidrogenase/genéticaRESUMO
BACKGROUND: Zymomonas mobilis is well known for its outstanding ability to produce ethanol with both high specific productivity and with high yield close to the theoretical maximum. The key enzyme in the ethanol production pathway is the pyruvate decarboxylase (PDC) which is converting pyruvate to acetaldehyde. Since it is widely considered that its gene pdc is essential, metabolic engineering strategies aiming to produce other compounds derived from pyruvate need to find ways to reduce PDC activity. RESULTS: Here, we present a new platform strain (sGB027) of Z. mobilis in which the native promoter of pdc was replaced with the IPTG-inducible PT7A1, allowing for a controllable expression of pdc. Expression of lactate dehydrogenase from E. coli in sGB027 allowed the production of D-lactate with, to the best of our knowledge, the highest reported specific productivity of any microbial lactate producer as well as with the highest reported lactate yield for Z. mobilis so far. Additionally, by expressing the L-alanine dehydrogenase of Geobacillus stearothermophilus in sGB027 we produced L-alanine, further demonstrating the potential of sGB027 as a base for the production of compounds other than ethanol. CONCLUSION: We demonstrated that our new platform strain can be an excellent starting point for the efficient production of various compounds derived from pyruvate with Z. mobilis and can thus enhance the establishment of this organism as a workhorse for biotechnological production processes.
Assuntos
Escherichia coli , Etanol , Ácido Láctico , Engenharia Metabólica , Piruvato Descarboxilase , Zymomonas , Zymomonas/metabolismo , Zymomonas/genética , Piruvato Descarboxilase/metabolismo , Piruvato Descarboxilase/genética , Engenharia Metabólica/métodos , Etanol/metabolismo , Ácido Láctico/metabolismo , Ácido Láctico/biossíntese , Escherichia coli/metabolismo , Escherichia coli/genética , L-Lactato Desidrogenase/metabolismo , L-Lactato Desidrogenase/genética , Alanina/metabolismo , Ácido Pirúvico/metabolismo , FermentaçãoRESUMO
Lactate dehydrogenase A (LDHA), a critical enzyme involved in glycolysis, is broadly involved multiple biological functions in human cancers. It is reported that LDHA can impact tumor immune surveillance and induce the transformation of tumor-associated macrophages, highlighting its unnoticed function of LDHA in immune system. However, in human cancers, the role of LDHA in prognosis and immunotherapy hasn't been investigated. In this study, we analyzed the expression pattern and prognostic value of LDHA in pan-cancer and explored its association between tumor microenvironment (TME), immune infiltration subtype, stemness scores, tumor mutation burden (TMB), and immunotherapy resistance. We found that LDHA expression is tumor heterogeneous and that its high expression is associated with poor prognosis in multiple human cancers. In addition, LDHA expression was positively correlated with the presence of mononuclear/macrophage cells, and also promoted the infiltration of a range of immune cells. Genomic alteration of LDHA was common in different types of cancer, while with prognostic value in pan-cancers. Pan-cancer analysis revealed that the significant correlations existed between LDHA expression and tumor microenvironment (including stromal cells and immune cells) as well as stemness scores (DNAss and RNAss) across cancer types. Drug sensitivity analysis also revealed that LDHA was able to predict response to chemotherapy and immunotherapy. Furthermore, it was confirmed that knockdown of LDHA reduced proliferation and migration ability of lung cancer cells. Taken together, LDHA could serve as a prognostic biomarker and a potential immunotherapy marker.
Assuntos
Resistencia a Medicamentos Antineoplásicos , Imunoterapia , Neoplasias , Microambiente Tumoral , Humanos , Microambiente Tumoral/imunologia , Prognóstico , Neoplasias/imunologia , Neoplasias/genética , Neoplasias/terapia , Resistencia a Medicamentos Antineoplásicos/genética , Biomarcadores Tumorais/metabolismo , Biomarcadores Tumorais/genética , Regulação Neoplásica da Expressão Gênica , L-Lactato Desidrogenase/metabolismo , L-Lactato Desidrogenase/genética , Linhagem Celular TumoralRESUMO
BACKGROUND: Head and neck squamous cell carcinoma (HNSCC) is one of the most common malignancies. Lactate dehydrogenase family genes (LDHs) play a critical role in tumor metabolism, but their functions in HNSCC have not been investigated thoroughly. Thus, we aimed to explore the value of LDHs in HNSCC. METHODS: The association between LDHs expression and mutations, methylation, copy number variations (CNVs), alternative splicing (AS) and competing endogenous RNA (ceRNA) was investigated in The Cancer Genome Atlas (TCGA). The expression level of LDHs in OSCC tissues and adjacent normal tissues was verified by qPCR. Algorithms, such as ssGSEA, ESTIMATE, xCell and TIDE were utilized to analyze the characteristics of immune infiltration. Pathway alternations were enriched by GO, GSEA and KEGG analysis. The Mantel test was employed to elucidate the correlation between metabolism and the tumor microenvironment (TME). Subsequently, MTT and colony formation assays were utilized to assess the impact of LDHB knockdown on cellular proliferation. Additionally, ATP and lactate assays were performed to examine metabolic alterations. Co-culture experiments further investigated the effect of LDHB knockdown on T cell differentiation. RESULTS: LDHs were completely analyzed in multiple databases, among which LDHB was differentially expressed in HNSCC and significantly associated with prognosis. Low LDHB expression had better clinicopathological characteristics. Downregulated LDHB expression was associated with enhanced immune cell infiltration and could influence tumor metabolism. Despite having worse cytotoxic T lymphocyte dysfunction, the LDHBlow group was predicted to respond more favorably to immune checkpoint inhibitors (ICIs) therapy. Moreover, the correlation between metabolism and TME was depicted. In vitro, LDHB knockdown resulted in inhibited cell proliferation, increased lactate levels and decreased ATP levels, while promoted the Th1 differentiation of T cells. CONCLUSIONS: Our study provided a comprehensive analysis of the LDHs and illustrated low LDHB expression could inhibit tumor cell proliferation and ATP production by influencing metabolism, with improved immune cell infiltration and better response to immunotherapy.
Assuntos
Neoplasias de Cabeça e Pescoço , Imunoterapia , L-Lactato Desidrogenase , Carcinoma de Células Escamosas de Cabeça e Pescoço , Humanos , Carcinoma de Células Escamosas de Cabeça e Pescoço/metabolismo , Carcinoma de Células Escamosas de Cabeça e Pescoço/genética , L-Lactato Desidrogenase/metabolismo , L-Lactato Desidrogenase/genética , Neoplasias de Cabeça e Pescoço/metabolismo , Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/patologia , Neoplasias de Cabeça e Pescoço/terapia , Microambiente Tumoral , Biomarcadores Tumorais/metabolismo , Biomarcadores Tumorais/genética , Linhagem Celular Tumoral , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Diferenciação Celular , IsoenzimasRESUMO
Lactate dehydrogenase B (LDHB) reversibly catalyzes the conversion of pyruvate to lactate or lactate to pyruvate and expressed in various malignancies. However, the role of LDHB in modulating immune responses against hepatocellular carcinoma (HCC) remains largely unknown. Here, we found that down-regulation of lactate dehydrogenase B (LDHB) was coupled with the promoter hypermethylation and knocking down the DNA methyltransferase 3A (DNMT 3A) restored LDHB expression levels in HCC cell lines. Bioinformatics analysis of the HCC cohort from The Cancer Genome Atlas revealed a significant positive correlation between LDHB expression and immune regulatory signaling pathways and immune cell infiltrations. Moreover, immune checkpoint inhibitors (ICIs) have shown considerable promise for HCC treatment and patients with higher LDHB expression responded better to ICIs. Finally, we found that overexpression of LDHB suppressed HCC growth in immunocompetent but not in immunodeficient mice, suggesting that the host immune system was involved in the LDHB-medicated tumor suppression. Our findings indicate that DNMT3A-mediated epigenetic silencing of LDHB may contribute to HCC progression through remodeling the tumor immune microenvironment, and LDHB may become a potential prognostic biomarker and therapeutic target for HCC immunotherapy.
Assuntos
Carcinoma Hepatocelular , DNA Metiltransferase 3A , Epigênese Genética , L-Lactato Desidrogenase , Neoplasias Hepáticas , Microambiente Tumoral , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Carcinoma Hepatocelular/imunologia , Carcinoma Hepatocelular/metabolismo , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/imunologia , Neoplasias Hepáticas/metabolismo , Microambiente Tumoral/imunologia , Humanos , Animais , Camundongos , L-Lactato Desidrogenase/metabolismo , L-Lactato Desidrogenase/genética , DNA Metiltransferase 3A/metabolismo , Regulação Neoplásica da Expressão Gênica , Metilação de DNA , Isoenzimas/genética , Isoenzimas/metabolismo , Linhagem Celular Tumoral , Inativação Gênica , PrognósticoRESUMO
Lactate dehydrogenase A (LDHA) is known to promote the growth and invasion of various types of tumors, affects tumor resistance, and is associated with tumor immune escape. But how LDHA reshapes the tumor microenvironment and promotes the progression of renal cell carcinoma (RCC) remains unclear. In this study, we found that LDHA was highly expressed in clear cell RCC (ccRCC), and this high expression was associated with macrophage infiltration, while macrophages were highly infiltrated in ccRCC, affecting patient prognosis via M2-type polarization. Our in vivo and in vitro experiments demonstrated that LDHA and M2-type macrophages could enhance the proliferation, invasion, and migration abilities of ccRCC cells. Mechanistically, high expression of LDHA in ccRCC cells upregulated the expression of EPHA2 in exosomes derived from renal cancer. Exosomal EPHA2 promoted M2-type polarization of macrophages by promoting activation of the PI3K/AKT/mTOR pathway in macrophages, thereby promoting the progression of ccRCC. All these findings suggest that EPHA2 may prove to be a potential therapeutic target for advanced RCC.
Assuntos
Carcinoma de Células Renais , Progressão da Doença , Exossomos , Neoplasias Renais , Macrófagos , Receptor EphA2 , Carcinoma de Células Renais/patologia , Carcinoma de Células Renais/metabolismo , Carcinoma de Células Renais/genética , Receptor EphA2/metabolismo , Receptor EphA2/genética , Humanos , Neoplasias Renais/patologia , Neoplasias Renais/metabolismo , Neoplasias Renais/genética , Exossomos/metabolismo , Animais , Macrófagos/metabolismo , Macrófagos/patologia , Camundongos , Linhagem Celular Tumoral , Proliferação de Células , L-Lactato Desidrogenase/metabolismo , L-Lactato Desidrogenase/genética , Movimento Celular , Regulação Neoplásica da Expressão Gênica , Masculino , Microambiente Tumoral , Prognóstico , Serina-Treonina Quinases TOR/metabolismo , Feminino , Transdução de Sinais , Camundongos Nus , Proteínas Proto-Oncogênicas c-akt/metabolismoRESUMO
Pikeperch (Sander lucioperca) is a freshwater species and an internationally highly demanded fish in aquaculture. Despite intensive research efforts on this species, fundamental knowledge of skeletal muscle biology and structural characteristics is missing. Therefore, we conducted a comprehensive analysis of skeletal muscle parameters in adult pikeperch from two different origins, wild-caught specimens from a lake and those reared in a recirculating aquaculture system. The analyses comprised the biochemical characteristics (nucleic acid, protein content), enzyme activities (creatine kinase, lactate dehydrogenase, NADP-dependent isocitrate dehydrogenase), muscle-specific gene and protein expression (related to myofibre formation, regeneration and permanent growth, muscle structure), and muscle fibre structure. The findings reveal distinct differences between the skeletal muscle of wild and farmed pikeperch. Specifically, nucleic acid content, enzyme activity, and protein expression varied significantly. The higher enzyme activity observed in wild pikeperch suggests greater metabolically activity in their muscles. Conversely, farmed pikeperch indicated a potential for pronounced muscle growth. As the data on pikeperch skeletal muscle characteristics is sparse, the purpose of our study is to gain fundamental insights into the characteristics of adult pikeperch muscle. The presented data serve as a foundation for further research on percids' muscle biology and have the potential to contribute to advancements and adaptations in aquaculture practices.
Assuntos
Aquicultura , Músculo Esquelético , Percas , Animais , Músculo Esquelético/metabolismo , Percas/genética , Percas/crescimento & desenvolvimento , Percas/metabolismo , Proteínas de Peixes/genética , Proteínas de Peixes/metabolismo , Animais Selvagens , L-Lactato Desidrogenase/metabolismo , L-Lactato Desidrogenase/genética , Proteínas Musculares/metabolismo , Proteínas Musculares/genética , Creatina Quinase/metabolismo , Creatina Quinase/genéticaRESUMO
Cancer cells exhibit a unique metabolic preference for the glycolytic pathway over oxidative phosphorylation for maintaining the tumor microenvironment. Lactate dehydrogenase A (LDHA) is a key enzyme that facilitates glycolysis by converting pyruvate to lactate and has been shown to be upregulated in multiple cancers due to the hypoxic tumor microenvironment. Diclofenac (DCF), a nonsteroidal anti-inflammatory drug, has been shown to exhibit anticancer effects by interfering with the glucose metabolism pathway. However, the specific targets of this drug remain unknown. Using in silico, biochemical, and biophysical studies, we show that DCF binds to LDHA adjacent to the substrate binding site and inhibits its activity in a dose-dependent and allosteric manner in HeLa cells. Thus, DCF inhibits the hypoxic microenvironment and induces apoptosis-mediated cell death. DCF failed to induce cytotoxicity in HeLa cells when LDHA was knocked down, confirming that DCF exerts its antimitotic effects via LDHA inhibition. DCF-induced LDHA inhibition alters pyruvate, lactate, NAD+, and ATP production in cells, and this could be a possible mechanism through which DCF inhibits glucose uptake in cancer cells. DCF-induced ATP deprivation leads to mitochondria-mediated oxidative stress, which results in DNA damage, lipid peroxidation, and apoptosis-mediated cell death. Reduction in intracellular ATP levels additionally activates the sensor kinase, adenosine monophosphate-activated protein kinase (AMPK), which further downregulates phosphorylated ribosomal S6 kinase (p-S6K), leading to apoptosis-mediated cell death. We find that in LDHA knocked down cells, intracellular ATP levels were depleted, resulting in the inhibition of p-S6K, suggesting the involvement of DCF-induced LDHA inhibition in the activation of the AMPK/S6K signaling pathway.
Assuntos
Proteínas Quinases Ativadas por AMP , Apoptose , Diclofenaco , Humanos , Células HeLa , Diclofenaco/farmacologia , Apoptose/efeitos dos fármacos , Proteínas Quinases Ativadas por AMP/metabolismo , Proteínas Quinases Ativadas por AMP/genética , L-Lactato Desidrogenase/metabolismo , L-Lactato Desidrogenase/antagonistas & inibidores , L-Lactato Desidrogenase/genética , Estresse Oxidativo/efeitos dos fármacos , Trifosfato de Adenosina/metabolismo , Anti-Inflamatórios não Esteroides/farmacologia , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismoRESUMO
Oligodendrocytes and astrocytes are metabolically coupled to neuronal compartments. Pyruvate and lactate can shuttle between glial cells and axons via monocarboxylate transporters. However, lactate can only be synthesized or used in metabolic reactions with the help of lactate dehydrogenase (LDH), a tetramer of LDHA and LDHB subunits in varying compositions. Here we show that mice with a cell type-specific disruption of both Ldha and Ldhb genes in oligodendrocytes lack a pathological phenotype that would be indicative of oligodendroglial dysfunctions or lack of axonal metabolic support. Indeed, when combining immunohistochemical, electron microscopical, and in situ hybridization analyses in adult mice, we found that the vast majority of mature oligodendrocytes lack detectable expression of LDH. Even in neurodegenerative disease models and in mice under metabolic stress LDH was not increased. In contrast, at early development and in the remyelinating brain, LDHA was readily detectable in immature oligodendrocytes. Interestingly, by immunoelectron microscopy LDHA was particularly enriched at gap junctions formed between adjacent astrocytes and at junctions between astrocytes and oligodendrocytes. Our data suggest that oligodendrocytes metabolize lactate during development and remyelination. In contrast, for metabolic support of axons mature oligodendrocytes may export their own glycolysis products as pyruvate rather than lactate. Lacking LDH, these oligodendrocytes can also "funnel" lactate through their "myelinic" channels between gap junction-coupled astrocytes and axons without metabolizing it. We suggest a working model, in which the unequal cellular distribution of LDH in white matter tracts facilitates a rapid and efficient transport of glycolysis products among glial and axonal compartments.
Assuntos
Axônios , Glicólise , L-Lactato Desidrogenase , Oligodendroglia , Animais , Oligodendroglia/metabolismo , Axônios/metabolismo , L-Lactato Desidrogenase/metabolismo , L-Lactato Desidrogenase/genética , Glicólise/fisiologia , Camundongos , Regulação para Baixo/fisiologia , Camundongos Endogâmicos C57BL , Lactato Desidrogenase 5/metabolismo , Astrócitos/metabolismo , Astrócitos/ultraestrutura , Camundongos Transgênicos , Isoenzimas/metabolismo , Isoenzimas/genética , Junções Comunicantes/metabolismo , Junções Comunicantes/ultraestrutura , Camundongos KnockoutRESUMO
Genotoxic therapy triggers reactive oxygen species (ROS) production and oxidative tissue injury. S-nitrosylation is a selective and reversible posttranslational modification of protein thiols by nitric oxide (NO), and 5,6,7,8-tetrahydrobiopterin (BH4) is an essential cofactor for NO synthesis. However, the mechanism by which BH4 affects protein S-nitrosylation and ROS generation has not been determined. Here, we showed that ionizing radiation disrupted the structural integrity of BH4 and downregulated GTP cyclohydrolase I (GCH1), which is the rate-limiting enzyme in BH4 biosynthesis, resulting in deficiency in overall protein S-nitrosylation. GCH1-mediated BH4 synthesis significantly reduced radiation-induced ROS production and fueled the global protein S-nitrosylation that was disrupted by radiation. Likewise, GCH1 overexpression or the administration of exogenous BH4 protected against radiation-induced oxidative injury in vitro and in vivo. Conditional pulmonary Gch1 knockout in mice (Gch1fl/fl; Sftpa1-Cre+/- mice) aggravated lung injury following irradiation, whereas Gch1 knock-in mice (Gch1lsl/lsl; Sftpa1-Cre+/- mice) exhibited attenuated radiation-induced pulmonary toxicity. Mechanistically, lactate dehydrogenase (LDHA) mediated ROS generation downstream of the BH4/NO axis, as determined by iodoacetyl tandem mass tag (iodoTMT)-based protein quantification. Notably, S-nitrosylation of LDHA at Cys163 and Cys293 was regulated by BH4 availability and could restrict ROS generation. The loss of S-nitrosylation in LDHA after irradiation increased radiosensitivity. Overall, the results of the present study showed that GCH1-mediated BH4 biosynthesis played a key role in the ROS cascade and radiosensitivity through LDHA S-nitrosylation, identifying novel therapeutic strategies for the treatment of radiation-induced lung injury.
Assuntos
Biopterinas , GTP Cicloidrolase , Lesão Pulmonar , Espécies Reativas de Oxigênio , Animais , Biopterinas/análogos & derivados , Biopterinas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Camundongos , Lesão Pulmonar/metabolismo , Lesão Pulmonar/etiologia , GTP Cicloidrolase/metabolismo , GTP Cicloidrolase/genética , Humanos , Tolerância a Radiação/genética , Lactato Desidrogenase 5/metabolismo , Camundongos Knockout , Óxido Nítrico/metabolismo , L-Lactato Desidrogenase/metabolismo , L-Lactato Desidrogenase/genética , Processamento de Proteína Pós-Traducional , Radiação IonizanteRESUMO
Lysine acetyltransferase 7 (KAT7), a histone acetyltransferase, has recently been identified as an oncoprotein and has been implicated in the development of various malignancies. However, its specific role in head and neck squamous carcinoma (HNSCC) has not been fully elucidated. Our study revealed that high expression of KAT7 in HNSCC patients is associated with poor survival prognosis and silencing KAT7 inhibits the Warburg effect, leading to reduced proliferation, invasion, and metastatic potential of HNSCC. Further investigation uncovered a link between the high expression of KAT7 in HNSCC and tumor-specific glycolytic metabolism. Notably, KAT7 positively regulates Lactate dehydrogenase A (LDHA), a key enzyme in metabolism, to promote lactate production and create a conducive environment for tumor proliferation and metastasis. Additionally, KAT7 enhances LDHA activity and upregulates LDHA protein expression by acetylating the lysine 118 site of LDHA. Treatment with WM3835, a KAT7 inhibitor, effectively suppressed the growth of subcutaneously implanted HNSCC cells in mice. In conclusion, our findings suggest that KAT7 exerts pro-cancer effects in HNSCC by acetylating LDHA and may serve as a potential therapeutic target. Inhibiting KAT7 or LDHA expression holds promise as a therapeutic strategy to suppress the growth and progression of HNSCC.
Assuntos
Proliferação de Células , Neoplasias de Cabeça e Pescoço , Histona Acetiltransferases , Carcinoma de Células Escamosas de Cabeça e Pescoço , Humanos , Animais , Neoplasias de Cabeça e Pescoço/patologia , Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/metabolismo , Carcinoma de Células Escamosas de Cabeça e Pescoço/patologia , Carcinoma de Células Escamosas de Cabeça e Pescoço/genética , Carcinoma de Células Escamosas de Cabeça e Pescoço/metabolismo , Acetilação , Linhagem Celular Tumoral , Histona Acetiltransferases/metabolismo , Histona Acetiltransferases/genética , Camundongos , L-Lactato Desidrogenase/metabolismo , L-Lactato Desidrogenase/genética , Lisina Acetiltransferases/metabolismo , Lisina Acetiltransferases/genética , Regulação Neoplásica da Expressão Gênica , Camundongos Nus , Efeito Warburg em Oncologia , Masculino , Feminino , Movimento Celular , Ensaios Antitumorais Modelo de Xenoenxerto , Invasividade Neoplásica , Isoenzimas/metabolismo , Isoenzimas/genéticaRESUMO
Abundant macrophage infiltration and altered tumor metabolism are two key hallmarks of glioblastoma. By screening a cluster of metabolic small-molecule compounds, we show that inhibiting glioblastoma cell glycolysis impairs macrophage migration and lactate dehydrogenase inhibitor stiripentol emerges as the top hit. Combined profiling and functional studies demonstrate that lactate dehydrogenase A (LDHA)-directed extracellular signal-regulated kinase (ERK) pathway activates yes-associated protein 1 (YAP1)/ signal transducer and activator of transcription 3 (STAT3) transcriptional co-activators in glioblastoma cells to upregulate C-C motif chemokine ligand 2 (CCL2) and CCL7, which recruit macrophages into the tumor microenvironment. Reciprocally, infiltrating macrophages produce LDHA-containing extracellular vesicles to promote glioblastoma cell glycolysis, proliferation, and survival. Genetic and pharmacological inhibition of LDHA-mediated tumor-macrophage symbiosis markedly suppresses tumor progression and macrophage infiltration in glioblastoma mouse models. Analysis of tumor and plasma samples of glioblastoma patients confirms that LDHA and its downstream signals are potential biomarkers correlating positively with macrophage density. Thus, LDHA-mediated tumor-macrophage symbiosis provides therapeutic targets for glioblastoma.