Upregulation of HAS2 promotes glioma cell proliferation and chemoresistance via c-myc.

Glioblastoma multiforme (GBM) is the most common and aggressive primary malignant human brain tumor. Although comprehensive therapies, including chemotherapy and radiotherapy following surgery, have shown promise in prolonging survival, the prognosis for GBM patients remains poor, with an overall survival rate of only 14.6 months. Chemoresistance is a major obstacle to successful treatment and contributes to relapse and poor survival rates in glioma patients. Therefore, there is an urgent need for novel strategies to overcome chemoresistance and improve treatment outcomes for human glioma patients. Recent studies have shown that the tumor microenvironment plays a key role in chemoresistance. Our study demonstrates that upregulation of HAS2 and subsequent hyaluronan secretion promotes glioma cell proliferation, invasion, and chemoresistance in vitro and in vivo through the c-myc pathway. Targeting HAS2 sensitizes glioma cells to chemotherapeutic agents. Additionally, we found that hypoxia-inducible factor HIF1α regulates HAS2 expression. Together, our findings provide insights into the dysregulation of HAS2 and its role in chemoresistance and suggest potential therapeutic strategies for GBM.

Pan-cancer analysis reveals the potential of hyaluronate synthase as therapeutic targets in human tumors.

Hyaluronic acid (HA) is a crucial component of the extracellular matrix, and its level of accumulation is related to the progression of various malignant tumors. In this study, a pan-cancer analysis of the three enzymes called hyaluronan synthases (HAS1, HAS2, and HAS3) that produce HA was performed. The study comprehensively describes the characteristics of HAS1, HAS2, and HAS3 in cancers using public databases and tools, to identify the potential biological pathways involved at the molecular, protein, cellular, and clinical sample levels. The analysis showed that dysregulation of the three genes often occurs in cancer, contributing to cancer progression, metastasis, and prognosis. Overexpression of HAS2 promotes secretion of HA in GBM and enhances cell proliferation and migration. The common and specific functions of HAS in certain diseases have important research implications for the treatment and prognosis of tumors.

Induction of cancer neoantigens facilitates development of clinically relevant models for the study of pancreatic cancer immunobiology.

Neoantigen burden and CD8 T cell infiltrate are associated with clinical outcome in pancreatic ductal adenocarcinoma (PDAC). A shortcoming of many genetic models of PDAC is the lack of neoantigen burden and limited T cell infiltrate. The goal of the present study was to develop clinically relevant models of PDAC by inducing cancer neoantigens in KP2, a cell line derived from the KPC model of PDAC. KP2 was treated with oxaliplatin and olaparib (OXPARPi), and a resistant cell line was subsequently cloned to generate multiple genetically distinct cell lines (KP2-OXPARPi clones). Clones A and E are sensitive to immune checkpoint inhibition (ICI), exhibit relatively high T cell infiltration, and have significant upregulation of genes involved in antigen presentation, T cell differentiation, and chemokine signaling pathways. Clone B is resistant to ICI and is similar to the parental KP2 cell line in terms of relatively low T cell infiltration and no upregulation of genes involved in the pathways noted above. Tumor/normal exome sequencing and in silico neoantigen prediction confirms successful generation of cancer neoantigens in the KP2-OXPARPi clones and the relative lack of cancer neoantigens in the parental KP2 cell line. Neoantigen vaccine experiments demonstrate that a subset of candidate neoantigens are immunogenic and neoantigen synthetic long peptide vaccines can restrain Clone E tumor growth. Compared to existing models, the KP2-OXPARPi clones better capture the diverse immunobiology of human PDAC and may serve as models for future investigations in cancer immunotherapies and strategies targeting cancer neoantigens in PDAC.

Structural insights into the substrate binding sites of O-carbamoyltransferase VtdB from Streptomyces sp. NO1W98.

The O-carbamoyltransferase VtdB catalyzes the carbamoylation of venturicidin B, which is essential for the biosynthesis of the antibiotic venturicidin A. Here, the crystal structures of VtdB and VtdB in complex with the intermediate carbamoyladenylate (VtdBCAO) were determined at resolutions of 2.99 Å and 2.90 Å, respectively. The structures resemble the conserved YrdC-like and specific Kae1-like domains. A magnesium ion and the intermediate carbamoyladenylate were also observed in the Kae1-like domain of VtdB. The structure of VtdBCAO in complex with the substrate venturicidin B was modeled by a molecular docking method to better understand the substrate binding mode, revealing a novel venturicidin B binding pocket.

IRAK4 Signaling Drives Resistance to Checkpoint Immunotherapy in Pancreatic Ductal Adenocarcinoma.

BACKGROUND & AIMS: Checkpoint immunotherapy is largely ineffective in pancreatic ductal adenocarcinoma (PDAC). The innate immune nuclear factor (NF)-κB pathway promotes PDAC cell survival and stromal fibrosis, and is driven by Interleukin-1 Receptor Associated Kinase-4 (IRAK4), but its impact on tumor immunity has not been directly investigated. METHODS: We interrogated The Cancer Genome Atlas data to identify the correlation between NF-κB and T cell signature, and a PDAC tissue microarray (TMA) to correlate IRAK4 activity with CD8+ T cell abundance. We performed RNA sequencing (RNA-seq) on IRAK4-deleted PDAC cells, and single-cell RNA-seq on autochthonous KPC (p48-Cre/TP53f/f/LSL-KRASG12D) mice treated with an IRAK4 inhibitor. We generated conditional IRAK4-deleted KPC mice and complementarily used IRAK4 inhibitors to determine the impact of IRAK4 on T cell immunity. RESULTS: We found positive correlation between NF-κB activity, IRAK4 and T cell exhaustion from The Cancer Genome Atlas. We observed inverse correlation between phosphorylated IRAK4 and CD8+ T cell abundance in a PDAC tissue microarray. Loss of IRAK4 abrogates NF-κB activity, several immunosuppressive factors, checkpoint ligands, and hyaluronan synthase 2, all of which drive T cell dysfunction. Accordingly, conditional deletion or pharmacologic inhibition of IRAK4 markedly decreased tumor desmoplasia and increased the abundance and activity of infiltrative CD4+ and CD8+ T cells in KPC tumors. Single-cell RNA-seq showed myeloid and fibroblast reprogramming toward acute inflammatory responses following IRAK4 inhibition. These changes set the stage for successful combination of IRAK4 inhibitors with checkpoint immunotherapy, resulting in excellent tumor control and markedly prolonged survival of KPC mice. CONCLUSION: IRAK4 drives T cell dysfunction in PDAC and is a novel, promising immunotherapeutic target.

The MK2/Hsp27 axis is a major survival mechanism for pancreatic ductal adenocarcinoma under genotoxic stress.

Combination chemotherapies remain the cornerstone treatment for pancreatic ductal adenocarcinoma (PDAC), but de novo and acquired resistance is common. In this study, we aimed to identify and characterize resistance mechanisms to a FIRINOX chemotherapy regimen (a combination of 5-fluorouracil, irinotecan, and oxaliplatin) because it is the most aggressive regimen currently used clinically for patients with PDAC. Using an unbiased reverse-phase protein array, we detected phospho-activation of heat shock protein 27 (Hsp27) as the most up-regulated event after FIRINOX treatment in PDAC cells. Silencing HSP27 by RNA interference or by a small-molecule inhibitor enhanced apoptosis caused by FIRINOX in vitro. Mechanistically, FIRINOX up-regulated tumor necrosis factor­α (TNFα), causing autocrine phosphorylation and activation of transforming growth factor­ß­activated kinase 1 (TAK1), MAPK activated protein kinase 2 (MAPKAPK2 or MK2), and, ultimately, Hsp27. Targeting MK2, the kinase that directly phosphorylates Hsp27, abrogated Hsp27 activation, sensitized PDAC cells to apoptosis, and suppressed SN-38­induced protective autophagy in vitro, in part by blocking phospho-activation of Beclin1. In an autochthonous PDAC mouse model, the MK2 inhibitor ATI-450 decreased PDAC development and progression. When combined with FIRINOX, ATI-450 eliminated most PDAC foci and marked prolonged mouse survival without causing additional toxicity. Last, we found that high phospho-MK2 expression in tumors was associated with poorer survival of patients with PDAC. Our study identified MK2 as a mediator of genotoxic stress­induced activation of prosurvival pathways and provides preclinical support for combining an MK2 inhibitor with FIRINOX-based chemotherapies to treat PDAC.

TPL2 enforces RAS-induced inflammatory signaling and is activated by point mutations.

NF-κB transcription factors, driven by the IRAK/IKK cascade, confer treatment resistance in pancreatic ductal adenocarcinoma (PDAC), a cancer characterized by near-universal KRAS mutation. Through reverse-phase protein array and RNA sequencing we discovered that IRAK4 also contributes substantially to MAPK activation in KRAS-mutant PDAC. IRAK4 ablation completely blocked RAS-induced transformation of human and murine cells. Mechanistically, expression of mutant KRAS stimulated an inflammatory, autocrine IL-1ß signaling loop that activated IRAK4 and the MAPK pathway. Downstream of IRAK4, we uncovered TPL2 (also known as MAP3K8 or COT) as the essential kinase that propels both MAPK and NF-κB cascades. Inhibition of TPL2 blocked both MAPK and NF-κB signaling, and suppressed KRAS-mutant cell growth. To counter chemotherapy-induced genotoxic stress, PDAC cells upregulated TLR9, which activated prosurvival IRAK4/TPL2 signaling. Accordingly, a TPL2 inhibitor synergized with chemotherapy to curb PDAC growth in vivo. Finally, from TCGA we characterized 2 MAP3K8 point mutations that hyperactivate MAPK and NF-κB cascades by impeding TPL2 protein degradation. Cancer cell lines naturally harboring these MAP3K8 mutations are strikingly sensitive to TPL2 inhibition, underscoring the need to identify these potentially targetable mutations in patients. Overall, our study establishes TPL2 as a promising therapeutic target in RAS- and MAP3K8-mutant cancers and strongly prompts development of TPL2 inhibitors for preclinical and clinical studies.

IRAK4 mediates colitis-induced tumorigenesis and chemoresistance in colorectal cancer.

Aberrant activation of the NF-κB transcription factors underlies chemoresistance in various cancer types, including colorectal cancer (CRC). Targeting the activating mechanisms, particularly with inhibitors to the upstream IκB kinase (IKK) complex, is a promising strategy to augment the effect of chemotherapy. However, clinical success has been limited, largely because of low specificity and toxicities of tested compounds. In solid cancers, the IKKs are driven predominantly by the Toll-like receptor (TLR)/IL-1 receptor family members, which signal through the IL-1 receptor-associated kinases (IRAKs), with isoform 4 (IRAK4) being the most critical. The pathogenic role and therapeutic value of IRAK4 in CRC have not been investigated. We found that IRAK4 inhibition significantly abrogates colitis-induced neoplasm in APCMin/+ mice, and bone marrow transplant experiments showed an essential role of IRAK4 in immune cells during neoplastic progression. Chemotherapy significantly enhances IRAK4 and NF-κB activity in CRC cells through upregulating TLR9 expression, which can in turn be suppressed by IRAK4 and IKK inhibitors, suggesting a feed-forward pathway that protects CRC cells from chemotherapy. Lastly, increased tumor phospho-IRAK4 staining or IRAK4 mRNA expression is associated with significantly worse survival in CRC patients. Our results support targeting IRAK4 to improve the effects of chemotherapy and outcomes in CRC.

Author Correction: TGF-ß1-induced miR-503 controls cell growth and apoptosis by targeting PDCD4 in glioblastoma cells.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.

Concurrent HER or PI3K Inhibition Potentiates the Antitumor Effect of the ERK Inhibitor Ulixertinib in Preclinical Pancreatic Cancer Models.

Effective treatment for pancreatic ductal adenocarcinoma (PDAC) is an urgent, unmet medical need. Targeting KRAS, the oncogene that is present in >95% of PDAC, is a heavily pursued strategy, but remains unsuccessful in the clinic. Therefore, targeting key effector cascades of KRAS oncoprotein, particularly the mitogenic RAF-MEK-ERK pathway, represents the next best strategy. However, RAF or MEK inhibitors have failed to show clinical efficacy in PDAC. Several studies have shown that cancer cells treated with RAF or MEK inhibitors adopt multiple mechanisms to reactivate ERK signaling. Therefore, development of ERK-specific inhibitors carries the promise to effectively abrogate this pathway. Ulixertinib (or BVD-523) is a first-in-class ERK-specific inhibitor that has demonstrated promising antitumor activity in a phase I clinical trial for advanced solid tumors with NRAS and BRAF mutations, providing a strong rationale to test this inhibitor in PDAC. In this study, we show that ulixertinib effectively inhibits in vitro growth of multiple PDAC lines and potentiates the cytotoxic effect of gemcitabine. Moreover, we found that PDAC cells treated with ulixertinib upregulates the parallel PI3K-AKT pathway through activating the HER/ErbB family proteins. Concurrent inhibition of PI3K or HER proteins synergizes with ulixertinib in suppressing PDAC cell growth in vitro and in vivo Overall, our study provides the preclinical rationale for testing combinations of ulixertinib with chemotherapy or PI3K and HER inhibitors in PDAC patients. Mol Cancer Ther; 17(10); 2144-55. ©2018 AACR.

Tumor-Stroma IL1ß-IRAK4 Feedforward Circuitry Drives Tumor Fibrosis, Chemoresistance, and Poor Prognosis in Pancreatic Cancer.

Targeting the desmoplastic stroma of pancreatic ductal adenocarcinoma (PDAC) holds promise to augment the effect of chemotherapy, but success in the clinic has thus far been limited. Preclinical mouse models suggest that near-depletion of cancer-associated fibroblasts (CAF) carries a risk of accelerating PDAC progression, underscoring the need to concurrently target key signaling mechanisms that drive the malignant attributes of both CAF and PDAC cells. We previously reported that inhibition of IL1 receptor-associated kinase 4 (IRAK4) suppresses NFκB activity and promotes response to chemotherapy in PDAC cells. In this study, we report that CAF in PDAC tumors robustly express activated IRAK4 and NFκB. IRAK4 expression in CAF promoted NFκB activity, drove tumor fibrosis, and supported PDAC cell proliferation, survival, and chemoresistance. Cytokine array analysis of CAF and microarray analysis of PDAC cells identified IL1ß as a key cytokine that activated IRAK4 in CAF. Targeting IRAK4 or IL1ß rendered PDAC tumors less fibrotic and more sensitive to gemcitabine. In clinical specimens of human PDAC, high stromal IL1ß expression associated strongly with poor overall survival. Together, our studies establish a tumor-stroma IL1ß-IRAK4 feedforward signal that can be therapeutically disrupted to increase chemotherapeutic efficacy in PDAC.Significance: Targeting the IL1ß-IRAK4 signaling pathway potentiates the effect of chemotherapy in pancreatic cancer. Cancer Res; 78(7); 1700-12. ©2018 AACR.

TGF-â1-induced miR-503 controls cell growth and apoptosis by targeting PDCD4 in glioblastoma cells.

Aberrant expression of microRNAs hae been shown to be closely associated with glioblastoma cell proliferation, apoptosis and drug resistance. However, mechanisms underlying the role of mcroRNAs in glioblastoma cell growth and apoptosis are not fully understood. In this study, we report that miR-503 is overexpressed in glioblastoma tissue compared with normal human brain tissue. Mechanistically, miR-503 can be induced by TGF-â1 at the transcriptional level by binding the smad2/3 binding elements in the promoter. Ectopic overexpression of miR-503 promotes cell growth and inhibits apoptosis by targeting PDCD4. In contrast, inhibition of miR-503 reduces cell growth. Furthermore, miR-503 inhibitor augments the growth inhibitory effect of temozolomide in glioblastoma cells. These results establish miR-503 as a promising molecular target for glioblastoma therapy.

The RNA-seq analysis suggests a potential multi-component complement system in oyster Crassostrea gigas.

The complement system is one of the major effector mechanisms of immune system, playing essential roles in both the innate and adaptive immune responses. In the present study, the counterparts of vertebrate complement components were identified by screening the sequenced genome of Crassostrea gigas, resulting in the identification of 792 gene models containing complement-related domains. The transcriptome of haemocytes at 6, 12 and 24 h post lipopolysaccharides (LPS) stimulation showed differential expression of 77 C1q domain containing proteins, 53 C-type lectins and 42 fibrinogen-related proteins. mRNAs encoding 18 serine protease domain-containing (SPC) proteins, 4 MACPF-domain containing proteins and 11 C3 receptor-like proteins were up-regulated upon LPS stimulation, and CgC3 mRNA was significantly increased at 12 h. The presence of CgC3 was confirmed in cell free plasma and was present in three subunit chains as expected for the processed mature protein. The complement related PRRs with coiled coil regions and SPC proteins with CUB domains may function in the activation of CgC3, whereas, the C3-like receptors with integrin-α/ß domain mediated the phagocytosis of C3-labled pathogens. These PRRs appear to serve as opsonins to promote phagocytosis of opsonized pathogens. The overall results suggested the existence of a potential multi-component complement system in C. gigas.

Constitutive IRAK4 Activation Underlies Poor Prognosis and Chemoresistance in Pancreatic Ductal Adenocarcinoma.

Purpose: Aberrant activation of the NF-κB transcription factors underlies the aggressive behavior and poor outcome of pancreatic ductal adenocarcinoma (PDAC). However, clinically effective and safe NF-κB inhibitors are not yet available. Because NF-κB transcription factors can be activated by the interleukin-1 receptor-associated kinases (IRAKs) downstream of the Toll-like receptors (TLRs), but has not been explored in PDAC, we sought to investigate the role of IRAKs in the pathobiology of PDAC.Experimental Design: We examined the phosphorylation status of IRAK4 (p-IRAK4), the master regulator of TLR signaling, in PDAC cell lines, in surgical samples and commercial tissue microarray. We then performed functional studies using small-molecule IRAK1/4 inhibitor, RNA-interference, and CRISPR/Cas9n techniques to delineate the role of IRAK4 in NF-κB activity, chemoresistance, cytokine production, and growth of PDAC cells in vitro and in vivoResults: p-IRAK4 staining was detectable in the majority of PDAC lines and about 60% of human PDAC samples. The presence of p-IRAK4 strongly correlated with phospho-NF-κB/p65 staining in PDAC samples and is predictive of postoperative relapse and poor overall survival. Inhibition of IRAK4 potently reduced NF-κB activity, anchorage-independent growth, chemoresistance, and secretion of proinflammatory cytokines from PDAC cells. Both pharmacologic suppression and genetic ablation of IRAK4 greatly abolished PDAC growth in mice and augmented the therapeutic effect of gemcitabine by promoting apoptosis, reducing tumor cell proliferation and tumor fibrosis.Conclusions: Our data established IRAK4 as a novel therapeutic target for PDAC treatment. Development of potent IRAK4 inhibitors is needed for clinical testing. Clin Cancer Res; 23(7); 1748-59. ©2016 AACR.

Nuclear PKM2 contributes to gefitinib resistance via upregulation of STAT3 activation in colorectal cancer.

Gefitinib (Iressa, ZD-1839), a small molecule tyrosine kinase inhibitor (TKI) of the epidermal growth factor receptor (EGFR) pathway, is currently under investigation in clinical trials for the treatment of colorectal cancer (CRC). However, as known, some patients develop resistance to TKIs, and the mechanisms mediating intrinsic resistance to EGFR-TKIs in CRC have not been fully characterized. Resistance to EGFR inhibitors reportedly involves activation of signal transducer and activator of transcription 3 (STAT3) in glioma and lung cancer. Here, we demonstrated that the nuclear pyruvate kinase isoform M2 (PKM2) levels were positively correlated with gefitinib resistance in CRC cells. The overexpression of nuclear PKM2 in HT29 cells decreased the effect of gefitinib therapy, whereas PKM2 knockdown increased gefitinib efficacy. Furthermore, the activation of STAT3 by nuclear PKM2 was associated with gefitinib resistance. Inhibition of STAT3 by Stattic, a STAT3-specific inhibitor, or STAT3-specific siRNA sensitized resistant cells to gefitinib. These results suggest that nuclear PKM2 modulates the sensitivity of CRC cells to gefitinib and indicate that small molecule pharmacological disruption of nuclear PKM2 association with STAT3 is a potential avenue for overcoming EGFR-TKI resistance in CRC patients.

A novel multi-domain C1qDC protein from Zhikong scallop Chlamys farreri provides new insights into the function of invertebrate C1qDC proteins.

The C1q domain containing (C1qDC) proteins are a family of proteins possessing globular C1q (gC1q) domains, and they rely on this domain to recognize various ligands such as PAMPs, immunoglobulins, ligands on apoptotic cell. In the present study, a novel multi-domain C1qDC protein (CfC1qDC-2) was identified from scallop Chlamys farreri, and its full length cDNA was composed of 1648 bp, encoding a signal peptide and three typical gC1q domains. BLAST analysis revealed significant sequence similarity between CfC1qDC-2 and C1qDC proteins from mollusks. Three gC1q domains were predicted in its tertiary structure to form a tightly packed bell-shaped trimer, and each one adopted a typical 10-stranded sandwich fold with a jelly-roll topology and contained six aromatic amino acids forming the hydrophobic core. The mRNA transcripts of CfC1qDC-2 were mainly detected in the tissues of hepatopancreas and gonad of adult scallops, and the expression level was up-regulated in hemocytes after stimulated by LPS, PGN and ß-glucan. During the embryonic development of scallop, the mRNA transcripts of CfC1qDC-2 were presented in all the detected stages, and the expression level was up-regulated from D-hinged larvae and reached the highest at eye-spot larvae. The recombinant protein of MBP-CfC1qDC-2 (rCfC1qDC-2) could bind various PAMPs including LPS, PGN, LTA, ß-glucan, mannan as well as polyI:C, and different microorganisms including three Gram-negative bacteria, three Gram-positive bacteria and two yeasts, as well as scallop apoptotic cells. Meanwhile, rCfC1qDC-2 could interact with human heat-aggregated IgG and IgM, and inhibit the C1q-dependent hemolysis of rabbit serum. All these results indicated that CfC1qDC-2 could recognize not only PAMPs as a PRR, but also the apoptotic cells. Moreover, the similar structures and functions shared by CfC1qDC-2 and complement C1q provided a new insight into the evolution of C1qDC proteins in complement system.

A C1q domain containing protein from Crassostrea gigas serves as pattern recognition receptor and opsonin with high binding affinity to LPS.

C1q proteins serve as pattern recognition receptors and involve in the pathogen recognition and complement pathway activation. In the present study, a novel C1q domain containing protein from Crassostrea gigas (designated CgC1qDC-1) was isolated by liposaccharide-Sepharose 6B affinity chromatography. The coding sequence of CgC1qDC-1 gene was determined by performing a homologous search of eight tryptic peptides identified by MALDI-TOF/TOF-MS against the genome of C. gigas. The coding sequence of CgC1qDC-1 was of 387 bp encoding a polypeptide of 128 amino acids containing a typical globular C1q domain. The globular C1q domain possessed eight ß strands with a jelly-roll topology structure, which was similar to the structure of human gC1q domain. The mRNA transcripts of CgC1qDC-1 were dominantly expressed in mantle and hemocytes, while low expressed in hepatopancreas, gonad, gill and muscle. The expression level of CgC1qDC-1 increased drastically at 6 h after Vibrio splendidus stimulation, and then gradually fell to the normal level at about 24 h. ELISA assay quantified that CgC1qDC-1 bound to LPS with high binding affinity (Kd = 0.09 × 10(-6) M). Moreover, CgC1qDC-1 significantly enhanced the phagocytosis of oyster hemocytes towards Gram-negative bacteria Escherichia coli and V. splendidus. These results collectively indicated that CgC1qDC-1 could serve as pattern recognition receptor and opsonin in the innate immune response against invading Gram-negative bacteria.

Metabolic reprogramming of cancer-associated fibroblasts by IDH3α downregulation.

Cancer-associated fibroblasts (CAFs) provide critical metabolites for tumor growth and undergo metabolic reprogramming to support glycolysis. However, the molecular mechanisms responsible for this change remain unclear. Here, we report that TGF-ß1- or PDGF-induced CAFs switch from oxidative phosphorylation to aerobic glycolysis. We identify downregulation of isocitrate dehydrogenase 3α (IDH3α) as a marker for this switch. Furthermore, miR-424 downregulates IDH3α during CAF formation. Downregulation of IDH3α decreases the effective level of α-ketoglutarate (α-KG) by reducing the ratio of α-KG to fumarate and succinate, resulting in PHD2 inhibition and HIF-1α protein stabilization. The accumulation of HIF-1α, in turn, promotes glycolysis by increasing the uptake of glucose, upregulating expression of glycolytic enzymes under normoxic conditions, and inhibiting oxidative phosphorylation by upregulating NDUFA4L2. CAFs from tumor samples exhibit low levels of IDH3α, and overexpression of IDH3α prevents transformation of fibroblasts into CAFs. Our studies reveal IDH3α to be a critical metabolic switch in CAFs.

Hexokinase 2 regulates G1/S checkpoint through CDK2 in cancer-associated fibroblasts.

Hexokinase 2 (HK2), a pivotal glycolytic enzyme, is often overexpressed in tumor cells and contributes to glycolysis. Emerging evidence has suggested that glycolysis is also enhanced in cancer-associated fibroblasts (CAF). However, it is not clear whether HK2 is involved in enhanced glycolysis in CAFs or what role HK2 plays in the CAFs. In this study, both time course experiments and dose response experiments demonstrated that the protein and mRNA levels of HK2 increase in CAF cells, according to western blot and quantitative PCR analyses, respectively. Additionally, miR-182 targets the 3' UTR of HK2, and its overexpression results in the degradation of HK2 mRNA, which eventually reduces the level of HK2 protein. On the other hand, knockdown of miR-182 increased the expression of HK2. Most importantly, HK2 regulated the protein level and T14 phosphorylation of CDK2, and knockdown of HK2 resulted in a G1 phase cell cycle arrest. These observations suggest that HK2 plays important roles in glycolysis regulation and in cell cycle checkpoint activation.

miR-181a mediates metabolic shift in colon cancer cells via the PTEN/AKT pathway.

Cancer cell metabolism is often characterized by a shift from an oxidative to a glycolytic bioenergetics pathway, a phenomenon known as the warburg effect. Whether the deregulation of miRNAs contributes to the warburg effect remains largely unknown. Here we show that miR-181a expression is increased and thus induces a metabolic shift in colon cancer cells. miR-181a performs this function by inhibiting the expression of PTEN, leading to an increase of phosphorylated AKT which triggers metabolic shift. The increase of lactate production induced by miR-181a results in the rapid growth of cancer cells. These results identify miR-181a as a molecular switch involved in the orchestration of the warburg effect in colon cancer cells via the PTEN/AKT pathway.