Phosphofructokinase Muscle Research Articles

Evaluation of Serum Pyruvate Kinase M2 and Key Glycolytic Signatures and Its Correlation with Phosphatidylinositol-3-kinase/Akt/Mammalian Target of Rapamycin Pathway in Women with Breast Carcinoma

Abstract Background: A desired target for cancer therapy is a cancer-specific metabolism which is associated with innate or acquired therapeutic resistance, malignancies, and aggressive cancer growth. Nevertheless, these cells utilize manifold metabolic routes to prolong their mass explosion. Human hexokinase 2 (HK2), phosphofructokinase, muscle (PFKM), pyruvate kinase M2 (PKM2), and lactate dehydrogenase A (LDHA) are fundamental signature players of cancer glycolysis playing a role in cancer cell regulation. Reflecting their metabolic roles may be possible with a better grasp of the metabolism transition in the early onset and spread of cancer. This study was designed to assess the serum protein levels of HK2, PFKM, PKM2, and LDHA in women with breast carcinoma and its correlation with phosphatidylinositol-3-kinase (PI3K)/Akt and the mammalian target of rapamycin (mTOR) pathway. Methods: In the current research, case–control investigation was undertaken with breast cancer women (n = 30) and compatible healthy women (n = 30) visiting Justice K. S. Hegde Charitable Hospital, Deralakatte, Mangaluru, who were selected on the basis of inclusion and exclusion criteria. Baseline characteristics for the study population were marked down. The HK2, PFKM, PKM2, and LDHA levels within serum were estimated by means of sandwich enzyme-linked immunosorbent assay. Results: We observed a difference that was significant among the two groups with regard to age and postmenopausal status and also a difference in first-degree family history. The levels of PKM2 and key glycolytic enzymes in serum varied between the case and control groups. There was a positive correlation that was observed in breast carcinoma subjects with glycolytic proteins and signaling pathway (PI3K/Akt/mTOR). Conclusions: Nevertheless, assured processes that persuade the target enzymes remain vague and require further in-depth exploration. Overall, discovering anticancer therapeutics that target glycolytic enzymes involved in glucose metabolism remains a dilemma.

ISG15 blocks cardiac glycolysis and ensures sufficient mitochondrial energy production during Coxsackievirus B3 infection.

Virus infection triggers inflammation and, may impose nutrient shortage to the heart. Supported by type I interferon (IFN) signalling, cardiomyocytes counteract infection by various effector processes, with the IFN-stimulated gene of 15 kDa (ISG15) system being intensively regulated and protein modification with ISG15 protecting mice Coxsackievirus B3 (CVB3) infection. The underlying molecular aspects how the ISG15 system affects the functional properties of respective protein substrates in the heart are unknown. Based on the protective properties due to protein ISGylation, we set out a study investigating CVB3-infected mice in depth and found cardiac atrophy with lower cardiac output in ISG15-/- mice. By mass spectrometry, we identified the protein targets of the ISG15 conjugation machinery in heart tissue and explored how ISGylation affects their function. The cardiac ISGylome showed a strong enrichment of ISGylation substrates within glycolytic metabolic processes. Two control enzymes of the glycolytic pathway, hexokinase 2 (HK2) and phosphofructokinase muscle form (PFK1), were identified as bona fide ISGylation targets during infection. In an integrative approach complemented with enzymatic functional testing and structural modelling, we demonstrate that protein ISGylation obstructs the activity of HK2 and PFK1. Seahorse-based investigation of glycolysis in cardiomyocytes revealed that, by conjugating proteins, the ISG15 system prevents the infection-/IFN-induced up-regulation of glycolysis. We complemented our analysis with proteomics-based advanced computational modelling of cardiac energy metabolism. Our calculations revealed an ISG15-dependent preservation of the metabolic capacity in cardiac tissue during CVB3 infection. Functional profiling of mitochondrial respiration in cardiomyocytes and mouse heart tissue by Seahorse technology showed an enhanced oxidative activity in cells with a competent ISG15 system. Our study demonstrates that ISG15 controls critical nodes in cardiac metabolism. ISG15 reduces the glucose demand, supports higher ATP production capacity in the heart, despite nutrient shortage in infection, and counteracts cardiac atrophy and dysfunction.

Effect and mechanism of Yueju Pills on breast cancer induced by DMBA in rats

This paper aims to explore the inhibitory effect of Yueju Pills on breast cancer and decipher the underlying mechanism. A total of 92 SPF-grade SD female rats were involved in this study, and 14 of them were randomly selected into control group. The remaining 78 rats were administrated with 7,12-dimethylbenzanthracene(DMBA) by gavage to establish the breast cancer model. The modeled rats were randomized into model, tamoxifen(1.9 mg·kg~(-1)·d~(-1)), and low-and high-dose(17, 34 g·kg~(-1)·d~(-1)) Yueju Pills groups. The mental state, food intake, and activities of the rats were observed daily, and the body weight was measured on alternate days. After 12 weeks of administration, the rats were sacrificed and the tumor weight was measured. The serum estrogen and progeste-rone levels were determined by enzyme-linked immunosorbent assay. The histopathological changes of the breast and tumor were observed by hematoxylin-eosin staining. Western blot was employed to measure the protein levels of glucose transporter 1(GLUT1), lactate dehydrogenase A(LDHA), phosphofructokinase muscle(PFKM), pyruvate kinase isozyme type M2(PKM2), hexokinase 2(HK2), nuclear factor-kappaB(NF-κB), and phosphorylated NF-κB. The intestinal microbiome was examined by 16S rRNA high-throughput sequencing. The results showed that compared with the model group, high and low-dose Yueju Pills showed the tumor inhibition rate of 15.8% and 64.5%, respectively, and the low dose group had stronger inhibitory effect. Compared with the control group, the model group presented elevated the levels of estrogen and progesterone in serum. The administration of Yueju Pills lowered such ele-vation, and the low-dose group showed stronger lowering effect(P<0.05). Compared with the model group, Yueju Pills reduced the glands with increased breast tissue, the degree of breast duct expansion, the number and area of acinar cavity, the secretions, and the layers of mammary epithelial cells. Furthermore, Yueju Pills down-regulated the expression of GLUT1, LDHA, PFKM, PKM2, HK2, and NF-κB(P<0.05) and altered the diversity, composition, structure, and abundance of intestinal flora. The results showed that Yueju Pills could inhibit breast cancer by regulating the secretion of estrogen and progesterone, glycolysis, inflammatory cytokines, and intestinal flora.

Transcriptome profiling of A549 non-small cell lung cancer cells in response to Trichinella spiralis muscle larvae excretory/secretory products.

Trichinella spiralis (T. spiralis) muscle-larva excretory/secretory products (ML-ESPs) is a complex array of proteins with antitumor activity. We previously demonstrated that ML-ESPs inhibit the proliferation of A549 non-small cell lung cancer (NSCLC) cell line. However, the mechanism of ML-ESPs against A549 cells, especially on the transcriptional level, remains unknow. In this study, we systematically investigated a global profile bioinformatics analysis of transcriptional response of A549 cells treated with ML-ESPs. And then, we further explored the transcriptional regulation of genes related to glucose metabolism in A549 cells by ML-ESPs. The results showed that ML-ESPs altered the expression of 2,860 genes (1,634 upregulated and 1,226 downregulated). GO and KEGG analysis demonstrated that differentially expressed genes (DEGs) were mainly associated with pathway in cancer and metabolic process. The downregulated genes interaction network of metabolic process is mainly associated with glucose metabolism. Furthermore, the expression of phosphofructokinase muscle (PFKM), phosphofructokinase liver (PFKL), enolase 2 (ENO2), lactate dehydrogenase B (LDHB), 6-phosphogluconolactonase (6PGL), ribulose-phosphate-3-epimerase (PRE), transketolase (TKT), transaldolase 1 (TALDO1), which genes mainly regulate glycolysis and pentose phosphate pathway (PPP), were suppressed by ML-ESPs. Interestingly, tricarboxylic acid cycle (TCA)-related genes, such as pyruvate dehydrogenase phosphatase 1 (PDP1), PDP2, aconitate hydratase 1 (ACO1) and oxoglutarate dehydrogenase (OGDH) were upregulated by ML-ESPs. In summary, the transcriptome profiling of A549 cells were significantly altered by ML-ESPs. And we also provide new insight into how ML-ESPs induced a transcriptional reprogramming of glucose metabolism-related genes in A549 cells.

Bone marrow mesenchymal stem cell-derived exosomal miR-21a-5p alleviates renal fibrosis by attenuating glycolysis by targeting PFKM

Renal fibrosis is a common pathological feature and outcome of almost all chronic kidney diseases, and it is characterized by metabolic reprogramming toward aerobic glycolysis. Mesenchymal stem cell-derived exosomes (MSC-Exos) have been proposed as a promising therapeutic approach for renal fibrosis. In this study, we investigated the effect of MSC-Exos on glycolysis and the underlying mechanisms. We demonstrated that MSC-Exos significantly ameliorated unilateral ureter obstruction (UUO)-induced renal fibrosis by inhibiting glycolysis in tubular epithelial cells (TECs). miRNA sequencing showed that miR-21a-5p was highly enriched in MSC-Exos. Mechanistically, miR-21a-5p repressed the expression of phosphofructokinase muscle isoform (PFKM), a rate-limiting enzyme of glycolysis, thereby attenuating glycolysis in TECs. Additionally, knockdown of miR-21a-5p abolished the renoprotective effect of MSC-Exos. These findings revealed a novel role for MSC-Exos in the suppression of glycolysis, providing a new insight into the treatment of renal fibrosis.

PFKM inhibits doxorubicin-induced cardiotoxicity by enhancing oxidative phosphorylation and glycolysis

Heart failure (HF) is a global pandemic which affects about 26 million people. PFKM (Phosphofructokinase, Muscle), catalyzing the phosphorylation of fructose-6-phosphate, plays a very important role in cardiovascular diseases. However, the effect of PFKM in glycolysis and HF remains to be elucidated. H9c2 rat cardiomyocyte cells were treated with doxorubicin (DOX) to establish injury models, and the cell viability, apoptosis and glycolysis were measured. Quantitative reverse transcription-polymerase chain reaction (RT-PCR) and immunoblotting were used for gene expression. DOX treatment significantly inhibited PFKM expression in H9c2 cells. Overexpression of PFKM inhibited DOX-induced cell apoptosis and DOX-decreased glycolysis and oxidative phosphorylation (OXPHOS), while silencing PFKM promoted cell apoptosis and inhibited glycolysis and OXPHOS in H9c2 cells. Moreover, PFKM regulated DOX-mediated cell viability and apoptosis through glycolysis pathway. Mechanism study showed that histone deacetylase 1 (HDAC1) inhibited H3K27ac-induced transcription of PFKM in DOX-treated cells and regulated glycolysis. PFKM could inhibit DOX-induced cardiotoxicity by enhancing OXPHOS and glycolysis, which might benefit us in developing novel therapeutics for prevention or treatment of HF.

An evaluation of the growth, blood biochemistry, hepatic glucose metabolism and hepatocyte apoptosis in the genetically improved farmed tilapia Oreochromis niloticus fed diets with distinct protein to corn starch ratios

This study was performed to evaluate the effects of dietary distinct protein to starch ratios on the growth, feed utilization, blood biochemistry, hepatic glucose metabolism and hepatocyte apoptosis of Nile tilapia Oreochromis niloticus. Three iso-lipidic (ca 6.56%) diets were formulated with elevated corn starch levels at the cost of protein provision, being designated as diets P40S5 (40.4% protein and 4.62% starch), P30S25 (30.4% protein and 24.2% starch) and P20S50 (19.9% protein and 48.6% starch). Tilapia juveniles (initial mean body weight, 32.0 g/fish) were assigned to 12 rectangular tanks (250 L each) with 20 fish per tank and fed the experimental diets to apparent satiation for 8 weeks. The results showed that either excess (40%) or insufficient (20%) amount of dietary protein impaired the growth of tilapia. Excess dietary protein led to unsatisfactory protein catabolism (lower protein efficiency ratio and protein productive value, and higher mRNA levels of hepatic glutamic-pyruvic transaminase 1 and glutamic-oxaloacetic transaminase paralogs), while insufficient amount of protein resulted in lower whole-body protein percentage. An increase in dietary starch from 5% or 25% to 50% was associated with enhanced lipid and glycogen storages in the liver of tilapia. Consistently, dietary starch reception up to 50% upregulated the expression of hepatic representative genes involved with glycolysis (glucokinase and phosphofructokinase muscle type a) and lipogenesis (acetyl-CoA carboxylase α and fatty acid synthase) while downregulated the expression of the gluconeogenic glucose-6-phosphatase catalytic subunit paralogs. TUNEL staining suggested that stronger apoptosis signal was recorded at the bile duct epithelium of the P20S50 fish. Concomitantly, plasma bile acid level nearly doubled, and an incidence (50%) of ‘green liver’ was observed in this treatment as compared with the P40S5 and P30S25 fish, indicating that bile acid metabolism disorder might be induced. Though plasma glucose (4.66–5.11 mmol/L) was not responsive to different treatments, advanced glycation end-products accumulated in the blood of the P20S50 fish, suggesting that 50% starch reception was probably associated with the progression of diabetes-associated pathologies. The overall results in this study suggested that tilapia might possess capability to tolerate dietary corn starch level up to 50% in a short-term exposure, whereas a long-term exposure might induce detrimental effects on liver functions.

Effect of lncRNA XLOC_005950 knockout by CRISPR/Cas9 gene editing on energy metabolism and proliferation in osteosarcoma MG63 cells mediated by hsa-miR-542-3p.

Cancer cells use glucose via glycolysis to maintain tumor cell proliferation. However, the effect of long non-coding RNAs (lncRNAs) on glycolysis in osteosarcoma (OS) cells remains unclear. The present study aimed to investigate the involvement of the lncRNA XLOC_005950/hsa-microRNA (miR)-542-3p/phosphofructokinase, muscle (PFKM) axis in the regulation of glucose metabolism, cell proliferation and apoptosis in the progression of OS. lncRNA XLOC_005950, hsa-miR-542-3p and PFKM expression in OS tissues and cells was detected via reverse transcription-quantitative PCR analysis. CRISPR/Cas9 gene editing was used to knockout lncRNA XLOC_005950 expression in MG63 cells. Cell Counting Kit-8 assay, flow cytometry, PFKM activity, and glucose and lactic acid content determination were performed to assess the effects of lncRNA XLOC_005950 knockout and overexpression of hsa-miR-542-3p on the phenotypes of OS cells. The dual-luciferase reporter assay was performed to confirm the targeting associations between lncRNA XLOC_005950, hsa-miR-542-3p and PFKM. The results demonstrated that lncRNA XLOC_005950 expression was upregulated in OS tissues and cells. Functional experiments indicated that lncRNA XLOC_005950 knockout decreased PFKM activity, the intracellular glucose and lactic acid content, and cell proliferation, while increasing apoptosis of OS cells. Furthermore, lncRNA XLOC_005950 knockout upregulated hsa-miR-542-3p expression and downregulated PFKM expression. Overexpression of hsa-miR-542-3p suppressed PFKM expression. Furthermore, lncRNA XLOC_005950, as the molecular sponge of miR-542-3p in OS, modulated the downstream target gene, PFKM. Taken together, the results of the present study suggest that lncRNA XLOC_005950 knockout may inhibit the progression of OS via hsa-miR-542-3p-mediated regulation of PFKM expression.

In silico analysis of SNPs in human phosphofructokinase, muscle (PFKM) gene: An apparent therapeutic target of aerobic glycolysis and cancer

Phosphofructokinase, muscle (PFKM), a key glycolytic regulatory enzyme, is a potential target for cancer therapeutic studies accredited to the employed inefficient phenomenon known as Warburg effect. PFKM is encoded by PFKM gene located at chromosome 12q13.11. Single nucleotide polymorphisms (SNPs) are known to profoundly affect gene expression and protein function. Therefore, the first attempt was made to computationally identify putative functional PFKM variants. These SNPs were further explored to find their probable association with different cancer types. A total of 9694 SNPs were retrieved from dbSNP database. Of which, only 85 validated SNPs with ≥10% minor allele frequency (MAF) were subjected to analysis by softwares including Ensembl Genome browser, FuncPred (SNPinfo), regulomeDB (v 2.0), SIFT and PolyPhen-2. The relative analysis of output obtained classified the selected-SNPs into 11 highly prioritized (HP), 20 moderately prioritized and 54 not/poorly prioritized SNPs. The 11 HP-SNPs were found to have the highest likelihood of being functionally important, evidenced by previous association of rs2269935, rs11168417, rs11609399 and rs2228500 HP-SNPs with cachexia, lung and breast cancer. The study warrants further experiments to confirm the predictive role of prioritized SNPs in cancer etiology and also provides directions to fellow researchers.

Glucose homeostasis and glucose tolerance were impaired with elevated lipid to starch ratios in practical diets for the omnivorous genetically improved farmed tilapia Oreochromis niloticus

The present study was performed to ascertain if glucose metabolism and glucose tolerance could be regulated by dietary lipid to starch ratios in the omnivorous genetically improved farmed tilapia Oreochromis niloticus. Three isonitrogenous (ca 34.5% protein) and isocaloric (ca 14.5 kJ/g) practical diets were formulated with elevated lipid at the cost of starch provision, being designated as diets L6S23 (5.55% lipid and 22.5% starch), L9S18 (8.77% lipid and 18.1% starch) and L12S13 (12.0% lipid and 13.8% starch). Tilapia juveniles (initial mean body weight: 23.0 g/fish) were allocated to 12 rectangular tanks (250 L) with 20 fish per tank, and were fed experimental diets to apparent satiation for 8 weeks. At the end of the trial, approximately 24 h after the last feeding, fish in each tank were bulk-weighed and counted. Nine fish per tank were sampled to analyze the whole-body composition (3 fish), blood chemistry (3 fish) and the expression of glucose-metabolic genes (3 fish). Thirty-six of the remaining fish per treatment were subjected to an acute glucose tolerance test. The results showed that the growth (weight gain) and feed utilization (feed and protein efficiency ratios) of tilapia were not affected by different treatments. Intraperitoneal fat ratio (1.32%) and lipid percentages in the liver (9.69%) and whole-body (10.2%) of the L12S13 fish were higher than those of the L6S23 fish (0.75%, 6.61% and 8.17%, respectively). The L9S18 (5.24 mmol/L) and L12S13 fish (5.59 mmol/L) obtained higher plasma glucose levels than the L6S23 fish (4.10 mmol/L). In the liver, the mRNA levels of representative genes involved in glycolysis (glucokinase) and glucose release (glucose-6-phosphatase catalytic subunit a2) were simultaneously up-regulated in the L12S13 fish as compared with the L6S23 fish, indicating that a futile cycle between glucose and glucose-6-phosphate was associated with elevated dietary lipid to starch ratios. In the white muscle, the mRNA levels of glucose transporter 1a (glut1a), glut4, hexokinase 1b, phosphofructokinase muscle type a (pfkma), pfkmb and glycogen synthase 1 were 0.44-, 0.71-, 0.58-, 0.51-, 0.72- and 0.53-fold lower in the L12S13 fish than in the L6S23 fish, suggesting that muscular glucose transport and utilization were depressed with elevated dietary lipid to starch ratios. Although it took similar time (3 h) to restore plasma glucose for all treatments, plasma glucose was lower in the L6S23 fish than in the L12S13 fish during 1–3 h after the glucose injection. Additionally, plasma glucose of the L9S18 and L12S13 fish decreased continually along the injection time (during 7–10 h) as compared with the L6S23 fish, which further proved that the regulatory mechanisms of glucose homeostasis were impaired in tilapia that were fed elevated lipid to starch ratios. Our data confirmed that the impairment of glucose homeostasis was also associated with high fat diet reception in the omnivorous tilapia.

Mycobacterium tuberculosis Limits Host Glycolysis and IL-1β by Restriction of PFK-M via MicroRNA-21

Increased glycolytic metabolism recently emerged as an essential process driving host defense against Mycobacterium tuberculosis (Mtb), but little is knownabout how this process is regulated during infection.Here, we observe repression of host glycolysis in Mtb-infected macrophages, which is dependent on sustained upregulation of anti-inflammatory microRNA-21 (miR-21) by proliferating mycobacteria. The dampening of glycolysis by miR-21 is mediated through targeting of phosphofructokinase muscle (PFK-M) isoform at the committed step of glycolysis, which facilitates bacterial growth by limiting pro-inflammatory mediators, chiefly interleukin-1β (IL-1β). Unlike other glycolytic genes, PFK-M expression and activity is repressed during Mtb infection through miR-21-mediated regulation, while other less-active isoenzymes dominate. Notably, interferon-γ (IFN-γ), which drives Mtb host defense, inhibits miR-21 expression, forcing an isoenzyme switch in the PFK complex, augmenting PFK-M expression and macrophage glycolysis. These findings place the targeting of PFK-M by miR-21 as a key node controlling macrophage immunometabolic function.

Molecular and metabolic adaption of glucose metabolism in the red and white muscle of the omnivorous GIFT tilapia Oreochromis niloticus to a glucose load

In this experiment, Genetically improved farmed Nile tilapia Oreochromis niloticus were intraperitoneally injected with 1 g glucose/kg of body weight or saline. Red and white muscle tissues were collected at 0, 1, 2, 4, 6 and 12 h after the glucose tolerance test (GTT) or saline injection, and the time course of changes in molecular and metabolic adaption of glucose metabolism of these two tissues were evaluated. The results showed that the expression of insulin-responsive glucose transporter 4 (glut4) was up-regulated at 4 h after the GTT in the red muscle, implying an increase of glucose uptake. However, the expression of glut4 in the white muscle did not change with glucose load. The glycolysis of red muscle in tilapia was stimulated during 2–4 h after the GTT, as the expression of hexokinase 1b (hk1b), hk2, phosphofructokinase muscle type a (pfkma) and pfkmb and the activity of HK and PFK increased. By contrast, only the expression of hk1b was up-regulated at 6 h after the GTT in the white muscle. The mRNA level of glycogen synthase 1 (gys1) and glycogen content increased at 2 and 6 h, respectively after the GTT in the red muscle, suggesting that glucose storage was provoked. However, glycogen content in the white muscle was not impacted by GTT. Lipogenesis was stimulated in the red muscle as reflected by up-regulated expression of acetyl-CoA carboxylase α (accα) (during 2–4 h) and accβ (during 4–12 h) with GTT. In the white muscle, however, the expression of accα was not changed, and mRNA level of accβ was not up-regulated until 6 h after the GTT. Taken together, it was concluded that the glycolytic and glycogen synthesis mechanisms in the red muscle were highly regulated by an acute glucose load while those in the white muscle were less responsive to this stimulus.

Abstract 5430: Evaluating a novel phytobiologic mixture against breast cancer cell lines: Effect on glycolytic enzymes and EMT gene expression profile

Abstract Background: Recently, there has been a growing evidence proposed regarding role of phytobiological compounds against various malignancies. One of the novel phytobiological mixture (PB), which is a mixture composed of the following: quercetin, resveratrol, cruciferex, epigallocatechin-3-gallate and curcumin at different proportions. To the best of our knowledge, this is the first study to evaluate the effects of a PB on breast cancer cell lines and its impact on the expression profile of these cells. Aim: Herein, we investigated the anti-proliferative effects of PB against two breast cancer cell lines: MDA-MB-231 and MCF-7, in addition, to investigating whether PB may alter the expression profile of glycolytic enzymes and epithelial mesenchymal transition (EMT) genes in these cell lines. Method: The anti-proliferative effect of PB was evaluated in vitro against MCF-7 and MDA-MB-231 cell lines using SRB assay. The effect of PB on the relative expression profile of some of the glycolytic enzymes and EMT genes was conducted using TaqMan gene expression assays via reverse transcription-quantitative PCR (RT-qPCR). Results: At 72 hrs, treatment with PB showed cytotoxic effects against MDA-MB-231 and MCF-7 cells with an IC50's of 25.9±3 µg/ml and 29.5± 0.9 µg/ml, respectively. At 24 hrs post PB treatment, there was a significant increase in the gene expression of most glycolytic enzymes such as; Phosphofructokinase muscle type (PFKM) (1.4± 0.1, p<0.01) and Pyruvate dehydrogenase kinase isoenzyme 1 (PDK-1) (1.6± 0.2 fold, p< 0.001). PB did not seem to alter the EMT gene expression profile with both cadherin 1 and 2 showing no significant changes. Interestingly, however, a significant up-regulation was seen with ZEB2 member of the zinc finger E-box binding homeobox 2, post PB treatment. Conclusion: In conclusion PB showed a promising anti-proliferative effect against two breast cancer cell lines. However, we postulate that the drug could be modulating glycolytic enzymes expression, which ultimately can affect key molecules in cell differentiation. Further studies are needed to elucidate the exact underlying mechanism and to better understand the most beneficial drug combinations that can work synergistically with PB to interfere with different tumor hallmarks. Citation Format: Aliaa A. Alamoudi, Afnan A. Alqarni, Ghada Ajabnoor, Aleksandra Niedwiecki, Matthias Rath, Steve M. Harakeh, Ahmed M. Al-Abd. Evaluating a novel phytobiologic mixture against breast cancer cell lines: Effect on glycolytic enzymes and EMT gene expression profile [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5430.

A Genome-wide Association Study of Early-Onset Breast Cancer IdentifiesPFKMas a Novel Breast Cancer Gene and Supports a Common Genetic Spectrum for Breast Cancer at Any Age

Early-onset breast cancer (EOBC) causes substantial loss of life and productivity, creating a major burden among women worldwide. We analyzed 1,265,548 Hapmap3 single-nucleotide polymorphisms (SNP) among a discovery set of 3,523 EOBC incident cases and 2,702 population control women ages ≤ 51 years. The SNPs with smallest P values were examined in a replication set of 3,470 EOBC cases and 5,475 control women. We also tested EOBC association with 19,684 genes by annotating each gene with putative functional SNPs, and then combining their P values to obtain a gene-based P value. We examined the gene with smallest P value for replication in 1,145 breast cancer cases and 1,142 control women. The combined discovery and replication sets identified 72 new SNPs associated with EOBC (P < 4 × 10(-8)) located in six genomic regions previously reported to contain SNPs associated largely with later-onset breast cancer (LOBC). SNP rs2229882 and 10 other SNPs on chromosome 5q11.2 remained associated (P < 6 × 10(-4)) after adjustment for the strongest published SNPs in the region. Thirty-two of the 82 currently known LOBC SNPs were associated with EOBC (P < 0.05). Low power is likely responsible for the remaining 50 unassociated known LOBC SNPs. The gene-based analysis identified an association between breast cancer and the phosphofructokinase-muscle (PFKM) gene on chromosome 12q13.11 that met the genome-wide gene-based threshold of 2.5 × 10(-6). In conclusion, EOBC and LOBC seem to have similar genetic etiologies; the 5q11.2 region may contain multiple distinct breast cancer loci; and the PFKM gene region is worthy of further investigation. These findings should enhance our understanding of the etiology of breast cancer.

Modulation of polyglutamine-induced cell death by genes identified by expression profiling.

The majority of triplet-repeat diseases are caused by mutated genes with an extended polyglutamine tract, exemplified by Huntington's disease (HD). In order to model HD pathogenesis in a controlled system, we developed stable PC12 cell lines that express exon 1 fragments of the huntingtin gene with 23 or 74 polyglutamines driven by an inducible doxycycline (dox)-sensitive promoter (HD-23Q or HD-74Q). We aimed to identify early perturbations induced by the mutation by studying expression levels of 1824 genes at 0, 5, 10 and 18 hours after induction, using adaptor-tagged competitive PCR (ATAC-PCR). At these time points, the cells show no appreciable death or mitochondrial impairment and very low inclusion levels. A total of 126 genes, including 69 known genes, exhibited statistically significant alterations in the HD-74Q cell lines but no changes in the HD-23Q lines. We tested 11 of these genes for their abilities to modulate polyglutamine-induced cell death in transiently transfected cell models. Five genes [glucose transporter 1 (Glut1), phosphofructokinase muscle isozyme (Pfkm), prostate glutathione-S -transferase 2 (Gstm2), RNA-binding motif protein 3 (Rbm3) and KRAB-A interacting protein 1 (Krip-1)] significantly suppressed cell death in both neuronal precursor and non-neuronal cell lines, suggesting that these transcriptional changes were relevant to polyglutamine pathology. The efficient recovery of functionally relevant genes supports the utility of gene expression profiling for discovering pathways related to pathogenesis, and the importance of analyzing molecular events in the early stages of disease.

Syntenic Assignment of Human Chromosome 1 Homologous Loci in the Bovine

Three mouse chromosomes (MMU 1, 3, and 4) carry homologs of human chromosome 1 (HSA 1) genes. A similar situation is found in the bovine, where five bovine chromosomes (BTA 2, 3, 5, 16, and unassigned syntenic group U25) contain homologs of HSA 1 loci. To evaluate further the syntenic relationship of HSA 1 homologs in cattle, 10 loci have been physically mapped through segregation analysis in bovine-rodent hybrid somatic cells. These loci, chosen for their location on HSA 1, are antithrombin 3 (AT3), renin (REN), complement component receptor 2 (CR2), phosphofructokinase muscle type (PFKM), Gardner-Rasheed feline sarcoma viral (v-fgr) oncogene homolog (FGR), alpha fucosidase (FUCA1), G-protein beta 1 subunit (GNB1), alpha 1A amylase, (AMY1), the neuroblastoma RAS viral (v-ras) oncogene homolog (NRAS), and alpha skeletal actin (ACTA1). AT3, REN, CR2, and GNB1 mapped to BTA 16, PFKM to BTA 5, AMY1A and NRAS to BTA 3, FGR and FUCA1 to BTA 2, and ACTA1 to BTA 28.