RNAi screening reveals a synthetic chemical–genetic interaction between ATP synthase and PFK1 in cancer cells

To meet cellular bioenergetic and biosynthetic demands, cancer cells remodel their metabolism to increase glycolytic flux, a phenomenon known as the Warburg effect and believed to contribute to cancer malignancy. Among glycolytic enzymes, phosphofructokinase-1 (PFK1) has been shown to act as a rate-limiting enzyme and to facilitate the Warburg effect in cancer cells. In this study, however, we found that decreased PFK1 activity did not affect cell survival or proliferation in cancer cells. This raised a question regarding the importance of PFK1 in malignancy. To gain insights into the role of PFK1 in cancer metabolism and the possibility of adopting it as a novel anticancer therapeutic target, we screened for genes that caused lethality when they were knocked down in the presence of tryptolinamide (TLAM), a PFK1 inhibitor. The screen revealed a synthetic chemical–genetic interaction between genes encoding subunits of ATP synthase (complex V) and TLAM. Indeed, after TLAM treatment, the sensitivity of HeLa cells to oligomycin A (OMA), an ATP synthase inhibitor, was 13,000 times higher than that of untreated cells. Furthermore, this sensitivity potentiation by TLAM treatment was recapitulated by genetic mutations of PFK1. By contrast, TLAM did not potentiate the sensitivity of normal fibroblast cell lines to OMA, possibly due to their reduced energy demands compared to cancer cells. We also showed that the PFK1-mediated glycolytic pathway can act as an energy reservoir. Selective potentiation of the efficacy of ATP synthase inhibitors by PFK1 inhibition may serve as a foundation for novel anticancer therapeutic strategies.     

Ectopic endometriotic stromal cells-derived lactate induces M2 macrophage polarization via Mettl3/Trib1/ERK/STAT3 signalling pathway in endometriosis

Endometriosis is a gynaecological condition characterized by the growth of endometrium-like tissues within and outside of the pelvic cavity. Recent studies have demonstrated that aberrant infiltration of M2 macrophages is mainly responsible for the establishment of endometriotic lesions. A growing body of evidence shows that glycolysis and lactate accumulation have great impact on the regulation of immunomicroenvironment. However, the communication signal between glycolysis and macrophages is poorly defined in endometriosis. Hereby, we investigate the correlation between glycolysis and M2 macrophage infiltration in endometriosis. Next, we confirm that lactate is pivotal factor that drives macrophage M2-polarization to promote endometriotic stromal cells invasion in vitro and in vivo. In addition, we also identify that the activation of Mettl3 and its target gene Trib1 promote M2 macrophage polarization. Moreover, we also demonstrate that Trib1 induce M2 macrophage polarization via the activation of ERK/STAT3 signalling pathway. Finally, by injecting 2-DG into endometriosis mice model, we show that the restrain of glycolysis significantly reduces the progression of endometriosis, which provides evidence for lactate as a potential therapeutic strategy for the prevention and treatment of endometriosis.     

Ubiquitin-specific peptidase 24 accelerates aerobic glycolysis and tumor progression in gastric carcinoma through stabilizing PLK1 to activate NOTCH1

Ubiquitin-specific peptidase 24 (USP24), a member of the deubiquitinase family, plays an important role in tumor regulation. However, the role of USP24 in gastric cancer (GC) is unknown. The aim of our study was to explore the role of USP24 in GC to seek new therapeutic targets for GC. TCGA analysis showed that USP24 was upregulated in GC patients, and its high expression levels were associated with poor prognosis. It was found that overexpressed USP24 promoted GC cell proliferation and abnormal glycolysis. Further analysis and study showed that USP24 could promote the stability and increase the expression of oncogene PLK1. Knocking down USP24 can reduce the stability of PLK1 to reduce Notch 1 activity, thus inhibiting GC glycolysis, proliferation, and other processes and alleviating tumor progression. Knocking down USP24 can inhibit GC progression. In conclusion, USP24 was upregulated in GC and promoted the growth and glycolysis of GC cells, the mechanism of which was related to the deubiquitination of PLK1 and the increase of its stability. Silencing USP24 inhibited the GC process. This study suggests that the USP24/PLK1/Notch 1 axis may be a novel therapeutic target for gastric cancer.     

表没食子儿茶素没食子酸酯通过PI3K/Akt通路抑制胰腺癌细胞糖酵解的实验研究

目的：探讨表没食子儿茶素没食子酸酯（EGCG）对胰腺癌细胞的增殖及糖代谢的影响。方法：用CCK8试剂盒检测不同浓度EGCG对胰腺癌PANC-1细胞增殖的影响，用葡萄糖乳酸试剂盒检测EGCG对糖酵解的影响，Western blot法检测糖酵解酶及PI3K/Akt通路蛋白的表达。结果：CCK8结果显示，EGCG可以抑制PANC-1细胞的增殖，其抑制作用具有剂量依赖性和时间依赖性（P <0.05）。葡萄糖乳酸检测实验表明，EGCG可以抑制PANC-1细胞的葡萄糖消耗和乳酸生成（P <0.05）。Western blot结果表明，EGCG可以抑制糖酵解酶己糖激酶-2(HK-2)、磷酸果糖激酶（PFK）和乳酸脱氢酶（LDH）及磷酸化PI3K和Akt的表达，且EGCG浓度越高，抑制作用越明显（P <0.05）。结论：EGCG抑制人胰腺癌细胞系PANC-1的增殖和糖酵解，其抑制作用可能与抑制PANC-1细胞中PI3K/Akt通路有关。     

HKDC1 upregulation promotes glycolysis and disease progression,and confers chemoresistance onto gastric cancer

There is increasing evidence that hexokinase is involved in cell proliferation and migration. However, the function of the hexokinase domain containing protein-1 (HKDC1) in gastric cancer (GC) remains unclear. Immunohistochemistry analysis and big data mining were used to evaluate the correlation between HKDC1 expression and clinical features in GC. In addition, the biological function and molecular mechanism of HKDC1 in GC were studied by in vitro and in vivo assays. Our study indicated that HKDC1 expression was upregulated in GC tissues compared with adjacent nontumor tissues. High expression of HKDC1 was associated with worse prognosis. Functional experiments demonstrated that HKDC1 upregulation promoted glycolysis, cell proliferation, and tumorigenesis. In addition, HKDC1 could enhance GC invasion and metastasis by inducing epithelial–mesenchymal transition (EMT). Abrogation of HKDC1 could effectively attenuate its oncogenic and metastatic function. Moreover, HKDC1 promoted GC proliferation and migration in vivo. HKDC1 overexpression conferred chemoresistance to cisplatin, oxaliplatin, and 5-fluorouracil (5-Fu) onto GC cells. Furthermore, nuclear factor kappa-B (NF-κB) inhibitor PS-341 could attenuate tumorigenesis, metastasis, and drug resistance ability induced by HKDC1 overexpression in GC cells. Our results highlight a critical role of HKDC1 in promoting glycolysis, tumorigenesis, and EMT of GC cells via activating the NF-κB pathway. In addition, HKDC1-mediated drug resistance was associated with DNA damage repair, which further activated NF-κB signaling. HKDC1 upregulation may be used as a potential indicator for choosing an effective chemotherapy regimen for GC patients undergoing chemotherapy.     

ALKBH5-induced circular RNA NRIP1 promotes glycolysis in thyroid cancer cells by targeting PKM2

Although circular RNAs (circRNAs) are involved in cell proliferation, differentiation, apoptosis, and invasion, the underlying regulatory mechanisms of circRNAs in thyroid cancer have not been fully elucidated. This article aimed to study the role of circRNA regulated by N6-methyladenosine modification in papillary thyroid cancer (PTC). Quantitative real-time PCR, western blotting, and immunohistochemistry were used to investigate the expressions of circRNA nuclear receptor-interacting protein 1 (circNRIP1) in PTC tissues and adjacent noncancerous thyroid tissues. In vitro and in vivo assays were carried out to assess the effects of circNRIP1 on PTC glycolysis and growth. The N6-methyladenosine mechanisms of circNRIP1 were evaluated by methylated RNA immunoprecipitation sequencing, luciferase reporter gene, and RNA stability assays. Results showed that circNRIP1 levels were significantly upregulated in PTC tissues. Furthermore, elevated circNRIP1 levels in PTC patients were correlated with high tumor lymph node metastasis stage and larger tumor sizes. Functionally, circNRIP1 significantly promoted glycolysis, PTC cell proliferation in vitro, and tumorigenesis in vivo. Mechanistically, circNRIP1 acted as a sponge for microRNA (miR)-541-5p and miR-3064-5p and jointly upregulated pyruvate kinase M2 (PKM2) expression. Knockdown of m⁶A demethylase α-ketoglutarate-dependent dioxygenase alkB homolog 5 (ALKBH5) significantly enhanced circNRIP1 m⁶A modification and upregulated its expression. These results show that ALKBH5 knockdown upregulates circNRIP1, thus promoting glycolysis in PTC cells. Therefore, circNRIP1 can be a prognostic biomarker and therapeutic target for PTC by acting as a sponge for oncogenic miR-541-5p and miR-3064-5p to upregulate PKM2 expression.     

Juglone promotes antitumor activity against prostate cancer via suppressing glycolysis and oxidative phosphorylation

The treatments currently used for prostate cancer (PC) do not meet clinical needs, and thus, new therapies with greater effectiveness are urgently required. Metabolic reprogramming of tumor cells is emerging as an exciting field for cancer therapy. Although the Warburg effect is a common feature of glucose metabolism in many cancers, PC cells have a unique metabolic phenotype. Non-neoplastic prostate cells show reduced oxidative phosphorylation (OXPHOS) because large, accumulated zinc inhibits citrate oxidation. During transformation, there are low levels of zinc in PC cells, and the tricarboxylic acid (TCA) cycle is reactivated. However, metastatic PC exhibits the Warburg effect. Due to metabolic differences in prostate tissue, targeting metabolic alterations in PC cells is an attractive therapeutic strategy. In this study, we investigated the effect of juglone on energy metabolism in PC cells. We found that juglone inhibited cell proliferation and induced apoptosis. Mechanistically, we demonstrated that juglone suppressed OXPHOS and glycolysis due to its inhibition of hexokinase (HK), phosphofructokinase (PFK), and pyruvate kinase (PK) activity. Furthermore, downregulation of PFK and PK, but not HK contributed to the inhibition of these enzyme activities. The current study indicates that further development of juglone for PC treatment would be beneficial.