Human papillomavirus-16 E6 activates the pentose phosphate pathway to promote cervical cancer cell proliferation by inhibiting G6PD lactylation.

High-risk human papillomaviruses (HPVs) are the causative agents of cervical cancer. Here, we report that HPV16 E6E7 promotes cervical cancer cell proliferation by activating the pentose phosphate pathway (PPP). We found that HPV16 E6 activates the PPP primarily by increasing glucose-6-phosphate dehydrogenase (G6PD) enzyme activity. Mechanistically, HPV16 E6 promoted G6PD dimer formation by inhibiting its lactylation. Importantly, we suggest that G6PD K45 was lactylated during G6PD-mediated antioxidant stress. In primary human keratinocytes and an HPV-negative cervical cancer C33A cells line ectopically expressing HPV16 E6, the transduction of G6PD K45A (unable to be lactylated) increased GSH and NADPH levels and, correspondingly, decreasing ROS levels. Conversely, the re-expression of G6PD K45T (mimicking constitutive lactylation) in HPV16-positive SiHa cells line inhibited cell proliferation. In vivo, the inhibition of G6PD enzyme activity with 6-aminonicotinamide (6-An) or the re-expression of G6PD K45T inhibited tumor proliferation. In conclusion, we have revealed a novel mechanism of HPV oncoprotein-mediated malignant transformation. These findings might provide effective strategies for treating cervical and HPV-associated cancers.

ACOX1 deficiency-induced lipid metabolic disorder facilitates chronic interstitial fibrosis development in renal allografts.

Chronic interstitial fibrosis presents a significant challenge to the long-term survival of transplanted kidneys. Our research has shown that reduced expression of acyl-coenzyme A oxidase 1 (ACOX1), which is the rate-limiting enzyme in the peroxisomal fatty acid ß-oxidation pathway, contributes to the development of fibrosis in renal allografts. ACOX1 deficiency leads to lipid accumulation and excessive oxidation of polyunsaturated fatty acids (PUFAs), which mediate epithelial-mesenchymal transition (EMT) and extracellular matrix (ECM) reorganization respectively, thus causing fibrosis in renal allografts. Furthermore, activation of Toll-like receptor 4 (TLR4)-nuclear factor kappa-B (NF-κB) signaling induced ACOX1 downregulation in a DNA methyltransferase 1 (DNMT1)-dependent manner. Overconsumption of PUFA resulted in endoplasmic reticulum (ER) stress, which played a vital role in facilitating ECM reorganization. Supplementation with PUFAs contributed to delayed fibrosis in a rat model of renal transplantation. The study provides a novel therapeutic approach that can delay chronic interstitial fibrosis in renal allografts by targeting the disorder of lipid metabolism.

Lactylation stabilizes DCBLD1 activating the pentose phosphate pathway to promote cervical cancer progression.

BACKGROUND: Discoidin, CUB, and LCCL domain-containing type I (DCBLD1) is identified as an oncogene involved in multiple regulation of tumor progression, but specific mechanisms remain unclear in cervical cancer. Lactate-mediated lactylation modulates protein function. Whether DCBLD1 can be modified by lactylation and the function of DCBLD1 lactylation are unknown. Therefore, this study aims to investigate the lactylation of DCBLD1 and identify its specific lactylation sites. Herein, we elucidated the mechanism by which lactylation modification stabilizes the DCBLD1 protein. Furthermore, we investigated DCBLD1 overexpression activating pentose phosphate pathway (PPP) to promote the progression of cervical cancer. METHODS: DCBLD1 expression was examined in human cervical cancer cells and adjacent non-tumorous tissues using quantitative reverse transcription-polymerase chain reaction, western blotting, and immunohistochemistry. In vitro and in vivo studies were conducted to investigate the impact of DCBLD1 on the progression of cervical cancer. Untargeted liquid chromatography-tandem mass spectrometry (LC-MS/MS) metabolomics studies were used to characterize DCBLD1-induced metabolite alterations. Western blot, immunofuorescence and transmission electron microscopy were performed to detect DCBLD1 degradation of G6PD by activating autophagy. Chromatin immunoprecipitation, dual luciferase reporter assay for detecting the mechanism by which lactate increases DCBLD1 transcription. LC-MS/MS was employed to verify specific modification sites within the DCBLD1 protein. RESULTS: We found that lactate increased DCBLD1 expression, activating the PPP to facilitate the proliferation and metastasis of cervical cancer cells. DCBLD1 primarily stimulated PPP by upregulating glucose-6-phosphate dehydrogenase (G6PD) expression and enzyme activity. The mechanism involved the increased enrichment of HIF-1α in the DCBLD1 promoter region, enhancing the DCBLD1 mRNA expression. Additionally, lactate-induced DCBLD1 lactylation stabilized DCBLD1 expression. We identified DCBLD1 as a lactylation substrate, with a predominant lactylation site at K172. DCBLD1 overexpression inhibited G6PD autophagic degradation, activating PPP to promote cervical cancer progression. In vivo, 6-An mediated inhibition of G6PD enzyme activity, inhibiting tumor proliferation. CONCLUSIONS: Our findings revealed a novel post-translational modification type of DCBDL1, emphasizing the significance of lactylation-driven DCBDL1-mediated PPP in promoting the progression of cervical cancer.

Human papillomavirus type 16 E6 promotes cervical cancer proliferation by upregulating transketolase enzymatic activity through the activation of protein kinase B.

Over 99% of precancerous cervical lesions are associated with human papillomavirus (HPV) infection, with HPV types 16 and 18 (especially type 16) found in over 70% of cervical cancer cases globally. E6, a critical HPV gene, triggers malignant proliferation by degrading p53; however, this mechanism alone cannot fully explain the oncogenic effects of HPV16 E6. Therefore, we aimed to investigate new targets of HPV oncogenic mechanisms. Our results revealed significant changes in nonoxidative pentose phosphate pathway (PPP) metabolites in HPV16-positive cells. However, the role of nonoxidative PPP in HPV-associated cell transformation and tumor development remained unexplored. In this study, we investigated the impact and mechanisms of HPV16 E6 on cervical cancer proliferation using the HPV-negative cervical cancer cell line (C33A). HPV16 E6 was found to promote cervical cancer cell proliferation both in vitro and in vivo, activating the nonoxidative PPP. Transketolase (TKT), a key enzyme in the nonoxidative PPP, is highly expressed in cervical cancer tissues and associated with poor prognosis. HPV16 E6 promotes cervical cancer cell proliferation by upregulating TKT activity through the activation of AKT. In addition, oxythiamine (OT), a TKT inhibitor, hindered tumor growth, with enhanced effects when combined with cisplatin (DDP). In conclusion, HPV16 E6 promotes cervical cancer proliferation by upregulating TKT activity through the activation of AKT. OT demonstrates the potential to inhibit HPV16-positive cervical cancer growth, and when combined with DDP, could further enhance the tumor-suppressive effect of DDP.

Peroxisome proliferator-activated receptors: A key link between lipid metabolism and cancer progression.

Lipids represent the essential components of membranes, serve as fuels for high-energy processes, and play crucial roles in signaling and cellular function. One of the key hallmarks of cancer is the reprogramming of metabolic pathways, especially abnormal lipid metabolism. Alterations in lipid uptake, lipid desaturation, de novo lipogenesis, lipid droplets, and fatty acid oxidation in cancer cells all contribute to cell survival in a changing microenvironment by regulating feedforward oncogenic signals, key oncogenic functions, oxidative and other stresses, immune responses, or intercellular communication. Peroxisome proliferator-activated receptors (PPARs) are transcription factors activated by fatty acids and act as core lipid sensors involved in the regulation of lipid homeostasis and cell fate. In addition to regulating whole-body energy homeostasis in physiological states, PPARs play a key role in lipid metabolism in cancer, which is receiving increasing research attention, especially the fundamental molecular mechanisms and cancer therapies targeting PPARs. In this review, we discuss how cancer cells alter metabolic patterns and regulate lipid metabolism to promote their own survival and progression through PPARs. Finally, we discuss potential therapeutic strategies for targeting PPARs in cancer based on recent studies from the last five years.

Targeted changes in blood lipids improves fibrosis in renal allografts.

BACKGROUND: Chronic interstitial fibrosis is the primary barrier against the long-term survival of transplanted kidneys. Extending the lifespan of allografts is vital for ensuring the long-term health of patients undergoing kidney transplants. However, few targets and their clinical applications have been identified. Moreover, whether dyslipidemia facilitates fibrosis in renal allograft remains unclear. METHODS: Blood samples were collected from patients who underwent kidney transplantation. Correlation analyses were conducted between the Banff score and body mass index, and serum levels of triacylglycerol, total cholesterol, low-density lipoprotein cholesterol, and high-density lipoprotein cholesterol. A rat model of renal transplantation was treated with the lipid-lowering drug, fenofibrate, and kidney fibrosis levels were determined by histochemical staining. Targeted metabolomic detection was conducted in blood samples from patients who underwent kidney transplantation and were divided into fibrotic and non-fibrotic groups. Rats undergoing renal transplantation were fed either an n-3 or n-6 polyunsaturated fatty acid (PUFA)-enriched diet. Immunohistochemical and Masson's trichrome staining were used to determine the degree of fibrosis. RESULTS: Hyperlipidemia was associated with fibrosis development. Treatment with fenofibrate contributed to improve fibrosis in a rat model of renal transplantation. Moreover, n-3 PUFAs from fibrotic group showed significant downregulation compared to patients without fibrotic renal allografts, and n-3 PUFAs-enriched diet contributed to delayed fibrosis in a rat model of renal transplantation. CONCLUSIONS: This study suggests that hyperlipidemia facilitates fibrosis of renal allografts. Importantly, a new therapeutic approach was provided that may delay chronic interstitial fibrosis in transplanted kidneys by augmenting the n-3 PUFA content in the diet.

The connection between tricarboxylic acid cycle enzyme mutations and pseudohypoxic signaling in pheochromocytoma and paraganglioma.

Pheochromocytomas and paragangliomas (PPGLs) are rare neuroendocrine tumors originating from chromaffin cells, holding significant clinical importance due to their capacity for excessive catecholamine secretion and associated cardiovascular complications. Roughly 80% of cases are associated with genetic mutations. Based on the functionality of these mutated genes, PPGLs can be categorized into distinct molecular clusters: the pseudohypoxia signaling cluster (Cluster-1), the kinase signaling cluster (Cluster-2), and the WNT signaling cluster (Cluster-3). A pivotal factor in the pathogenesis of PPGLs is hypoxia-inducible factor-2α (HIF2α), which becomes upregulated even under normoxic conditions, activating downstream transcriptional processes associated with pseudohypoxia. This adaptation provides tumor cells with a growth advantage and enhances their ability to thrive in adverse microenvironments. Moreover, pseudohypoxia disrupts immune cell communication, leading to the development of an immunosuppressive tumor microenvironment. Within Cluster-1a, metabolic perturbations are particularly pronounced. Mutations in enzymes associated with the tricarboxylic acid (TCA) cycle, such as succinate dehydrogenase (SDHx), fumarate hydratase (FH), isocitrate dehydrogenase (IDH), and malate dehydrogenase type 2 (MDH2), result in the accumulation of critical oncogenic metabolic intermediates. Notable among these intermediates are succinate, fumarate, and 2-hydroxyglutarate (2-HG), which promote activation of the HIFs signaling pathway through various mechanisms, thus inducing pseudohypoxia and facilitating tumorigenesis. SDHx mutations are prevalent in PPGLs, disrupting mitochondrial function and causing succinate accumulation, which competitively inhibits α-ketoglutarate-dependent dioxygenases. Consequently, this leads to global hypermethylation, epigenetic changes, and activation of HIFs. In FH-deficient cells, fumarate accumulation leads to protein succination, impacting cell function. FH mutations also trigger metabolic reprogramming towards glycolysis and lactate synthesis. IDH1/2 mutations generate D-2HG, inhibiting α-ketoglutarate-dependent dioxygenases and stabilizing HIFs. Similarly, MDH2 mutations are associated with HIF stability and pseudohypoxic response. Understanding the intricate relationship between metabolic enzyme mutations in the TCA cycle and pseudohypoxic signaling is crucial for unraveling the pathogenesis of PPGLs and developing targeted therapies. This knowledge enhances our comprehension of the pivotal role of cellular metabolism in PPGLs and holds implications for potential therapeutic advancements.

The role of liver kinase B1 in tumor progression through regulation of lipid metabolism

The somatic mutation of liver kinase B1 (LKB1) has been implicated in various tumors, which is reflected in the survival, proliferation, and metastasis of tumor cells. However, the regulation of LKB1 in lipid metabolism, a process that is involved in tumor progression is not completely clear. We conclude that LKB1 deficiency results in abnormal expression and activation of multiple molecules related to lipid metabolism which locate downstream of AMP-activated protein kinase (AMPK) or salt-induced kinase (SIK). Abnormal lipid metabolism induced by LKB1 deficiency contributes to the proliferation and metastasis of tumor cells through energy regulation. (AU)

Recent findings in the regulation of G6PD and its role in diseases.

Glucose-6-phosphate dehydrogenase (G6PD) is the only rate-limiting enzyme in the pentose phosphate pathway (PPP). Rapidly proliferating cells require metabolites from PPP to synthesize ribonucleotides and maintain intracellular redox homeostasis. G6PD expression can be abnormally elevated in a variety of cancers. In addition, G6PD may act as a regulator of viral replication and vascular smooth muscle function. Therefore, G6PD-mediated activation of PPP may promote tumor and non-neoplastic disease progression. Recently, studies have identified post-translational modifications (PTMs) as an important mechanism for regulating G6PD function. Here, we provide a comprehensive review of various PTMs (e.g., phosphorylation, acetylation, glycosylation, ubiquitination, and glutarylation), which are identified in the regulation of G6PD structure, expression and enzymatic activity. In addition, we review signaling pathways that regulate G6PD and evaluate the role of oncogenic signals that lead to the reprogramming of PPP in tumor and non-neoplastic diseases as well as summarize the inhibitors that target G6PD.

The role of transketolase in human cancer progression and therapy.

Transketolase (TKT) is an enzyme that is ubiquitously expressed in all living organisms and has been identified as an important regulator of cancer. Recent studies have shown that the TKT family includes the TKT gene and two TKT-like (TKTL) genes; TKTL1 and TKTL2. TKT and TKTL1 have been reported to be involved in the regulation of multiple cancer-related events, such as cancer cell proliferation, metastasis, invasion, epithelial-mesenchymal transition, chemoradiotherapy resistance, and patient survival and prognosis. Therefore, TKT may be an ideal target for cancer treatment. More importantly, the levels of TKTL1 were detected using EDIM technology for the early detection of some malignancies, and TKTL1 was more sensitive and specific than traditional tumor markers. Detecting TKTL1 levels before and after surgery could be used to evaluate the surgery's effect. While targeted TKT suppresses cancer in multiple ways, in some cases, it has detrimental effects on the organism. In this review, we discuss the role of TKT in different tumors and the detailed mechanisms while evaluating its value and limitations in clinical applications. Therefore, this review provides a basis for the clinical application of targeted therapy for TKT in the future, and a strategy for subsequent cancer-related research.

The role of liver kinase B1 in tumor progression through regulation of lipid metabolism.

The somatic mutation of liver kinase B1 (LKB1) has been implicated in various tumors, which is reflected in the survival, proliferation, and metastasis of tumor cells. However, the regulation of LKB1 in lipid metabolism, a process that is involved in tumor progression is not completely clear. We conclude that LKB1 deficiency results in abnormal expression and activation of multiple molecules related to lipid metabolism which locate downstream of AMP-activated protein kinase (AMPK) or salt-induced kinase (SIK). Abnormal lipid metabolism induced by LKB1 deficiency contributes to the proliferation and metastasis of tumor cells through energy regulation.

LKB1 deficiency-induced metabolic reprogramming in tumorigenesis and non-neoplastic diseases.

BACKGROUND: Live kinase B1 (LKB1) is a tumor suppressor that is mutated in Peutz-Jeghers syndrome (PJS) and a variety of cancers. Lkb1 encodes serine-threonine kinase (STK) 11 that activates AMP-activated protein kinase (AMPK) and its 13 superfamily members, regulating multiple biological processes, such as cell polarity, cell cycle arrest, embryo development, apoptosis, and bioenergetics metabolism. Increasing evidence has highlighted that deficiency of LKB1 in cancer cells induces extensive metabolic alterations that promote tumorigenesis and development. LKB1 also participates in the maintenance of phenotypes and functions of normal cells through metabolic regulation. SCOPE OF REVIEW: Given the important role of LKB1 in metabolic regulation, we provide an overview of the association of metabolic alterations in glycolysis, aerobic oxidation, the pentose phosphate pathway (PPP), gluconeogenesis, glutamine, lipid, and serine induced by aberrant LKB1 signals in tumor progression, non-neoplastic diseases, and functions of immune cells. MAJOR CONCLUSIONS: In this review, we summarize layers of evidence demonstrating that disordered metabolisms in glucose, glutamine, lipid, and serine caused by LKB1 deficiency promote carcinogenesis and non-neoplastic diseases. The metabolic reprogramming resulting from the loss of LKB1 confers cancer cells with growth or survival advantages. Nevertheless, it also causes a metabolic frangibility for LKB1-deficient cancer cells. The metabolic regulation of LKB1 also plays a vital role in maintaining cellular phenotype in the progression of non-neoplastic diseases. In addition, lipid metabolic regulation of LKB1 plays an important role in controlling the function, activity, proliferation, and differentiation of several types of immune cells. We conclude that in-depth knowledge of metabolic pathways regulated by LKB1 is conducive to identifying therapeutic targets and developing drug combinations to treat cancers and metabolic diseases and achieve immunoregulation.

Fenofibrate Improved Interstitial Fibrosis of Renal Allograft through Inhibited Epithelial-Mesenchymal Transition Induced by Oxidative Stress.

The best treatment for end-stage renal disease is renal transplantation. However, it is often difficult to maintain a renal allograft healthy for a long time following transplantation. Interstitial fibrosis and tubular atrophy (IF/TA) are significant histopathologic characteristics of a compromised renal allograft. There is no effective therapy to improve renal allograft function once IF/TA sets in. Although there are many underlying factors that can induce IF/TA, the pathogenesis of IF/TA has not been fully elucidated. It has been found that epithelial-mesenchymal transition (EMT) significantly contributes to the development of IF/TA. Oxidative stress is one of the main causes that induce EMT in renal allografts. In this study, we have used H2O2 to induce oxidative stress in renal tubular epithelial cells (NRK-52e) of rats. We also pretreated NRK-52e cells with an antioxidant (N-acetyl L-cysteine (NAC)) 1 h prior to the treatment with H2O2. Furthermore, we used fenofibrate (a peroxisome proliferator-activated receptor α agonist) to treat NRK-52e cells and a renal transplant rat model. Our results reveal that oxidative stress induces EMT in NRK-52e cells, and pretreatment with NAC can suppress EMT in these cells. Moreover, fenofibrate suppresses fibrosis by ameliorating oxidative stress-induced EMT in a rat model. Thus, fenofibrate may effectively prevent the development of fibrosis in renal allograft and improve the outcome.

PARP-1 inhibitors sensitize HNSCC cells to APR-246 by inactivation of thioredoxin reductase 1 (TrxR1) and promotion of ROS accumulation.

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer worldwide. Mutations of TP53 may reach 70% - 85% in HNSCC patients without human papillomavirus (HPV) infection. Recurrence rate remains particularly high for HNSCC patients with mutations in the TP53 gene although patients are responsive to surgery, irradiation, and chemotherapy early in the treatment. p53-Reactivation and Induction of Massive Apoptosis-1 (PRIMA-1) and its methylated analogue PRIMA-1Met (also known as APR-246) are quinuclidine compounds that rescue the DNA-binding activity of mutant p53 (mut-p53) and restore the potential of wild-type p53. In the current report, we demonstrated that inhibition of poly (ADP-ribose) polymerase-1 (PARP-1) with 6(5H)-phenanthridinone (PHEN) and N-(6-Oxo-5,6-dihydrophenanthridin-2-yl)-(N, N-dimethylamino) acetamide hydrochloride (PJ34) sensitizes UMSCC1, UMSCC14, and UMSCC17A, three HNSCC cell lines to the treatment of APR-246. PHEN enhances APR-246-induced apoptosis, but not programmed necrosis or autophagic cell death in HNSCC cells. The PARP-1 inhibition-induced sensitization of HNSCC cells to APR-246 is independent of TP53 mutation. Instead, PARP-1 inhibition promotes APR-246-facilitated inactivation of thioredoxin reductase 1 (TrxR1), leading to ROS accumulation and DNA damage. Overexpression of TrxR1 or application of antioxidant N-acetyl-L-cysteine (NAC) depletes the ROS increase, reduces DNA damage, and decreases cell death triggered by APR-246/PHEN in HNSCC cells. Thus, we have characterized a new function of PARP-1 inhibitor in HNSCC cells by inactivation of TrxR1 and elevation of ROS and provide a novel therapeutic strategy for HNSCC by the combination of PARP-1 inhibitors and APR-246.

Piperlongumine and p53-reactivator APR-246 selectively induce cell death in HNSCC by targeting GSTP1.

TP53 mutations frequently occur in head and neck squamous cell carcinoma (HNSCC) patients without human papillomavirus infection. The recurrence rate for these patients is distinctly high. It has been actively explored to identify agents that target TP53 mutations and restore wild-type (WT) TP53 activities in HNSCC. PRIMA-1 (p53-reactivation and induction of massive apoptosis-1) and its methylated analogue PRIMA-1Met (also called APR-246) were found to be able to reestablish the DNA-binding activity of p53 mutants and reinstate the functions of WT p53. Herein we report that piperlongumine (PL), an alkaloid isolated from Piper longum L., synergizes with APR-246 to selectively induce apoptosis and autophagic cell death in HNSCC cells, whereas primary and immortalized mouse embryonic fibroblasts and spontaneously immortalized non-tumorigenic human skin keratinocytes (HaCat) are spared from the damage by the co-treatment. Interestingly, PL-sensitized HNSCC cells to APR-246 are TP53 mutation-independent. Instead, we demonstrated that glutathione S-transferase pi 1 (GSTP1), a GST family member that catalyzes the conjugation of GSH with electrophilic compounds to fulfill its detoxification function, is highly expressed in HNSCC tissues. Administration of PL and APR-246 significantly suppresses GSTP1 activity, resulting in the accumulation of ROS, depletion of GSH, elevation of GSSG, and DNA damage. Ectopic expression of GSTP1 or pre-treatment with antioxidant N-acetyl-L-cysteine (NAC) abrogates the ROS elevation and decreases DNA damage, apoptosis, and autophagic cell death prompted by PL/APR-246. In addition, administration of PL and APR-246 impedes UMSCC10A xenograft tumor growth in SCID mice. Taken together, our data suggest that HNSCC cells are selectively sensitive to the combination of PL and APR-246 due to a remarkably synergistic effect of the co-treatment in the induction of ROS by suppression of GSTP1.