SMYD2 regulates vascular smooth muscle cell phenotypic switching and intimal hyperplasia via interaction with myocardin.

The SET and MYND domain-containing protein 2 (SMYD2) is a histone lysine methyltransferase that has been reported to regulate carcinogenesis and inflammation. However, its role in vascular smooth muscle cell (VSMC) homeostasis and vascular diseases has not been determined. Here, we investigated the role of SMYD2 in VSMC phenotypic modulation and vascular intimal hyperplasia and elucidated the underlying mechanism. We observed that SMYD2 expression was downregulated in injured carotid arteries in mice and phenotypically modulated VSMCs in vitro. Using an SMC-specific SMYD2 knockout mouse model, we found that SMYD2 ablation in VSMCs exacerbated neointima formation after vascular injury in vivo. Conversely, SMYD2 overexpression inhibited VSMC proliferation and migration in vitro and attenuated arterial narrowing in injured vessels in mice. SMYD2 downregulation promoted VSMC phenotypic switching accompanied with enhanced proliferation and migration. Mechanistically, genome-wide transcriptome analysis and loss/gain-of-function studies revealed that SMYD2 up-regulated VSMC contractile gene expression and suppressed VSMC proliferation and migration, in part, by promoting expression and transactivation of the master transcription cofactor myocardin. In addition, myocardin directly interacted with SMYD2, thereby facilitating SMYD2 recruitment to the CArG regions of SMC contractile gene promoters and leading to an open chromatin status around SMC contractile gene promoters via SMYD2-mediated H3K4 methylation. Hence, we conclude that SMYD2 is a novel regulator of VSMC contractile phenotype and intimal hyperplasia via a myocardin-dependent epigenetic regulatory mechanism.

High-throughput screens identify a lipid nanoparticle that preferentially delivers mRNA to human tumors in vivo.

Lipid nanoparticles (LNPs) are a clinically relevant way to deliver therapeutic mRNA to hepatocytes in patients. However, LNP-mRNA delivery to end-stage solid tumors such as head and neck squamous cell carcinoma (HNSCC) remains more challenging. While scientists have used in vitro assays to evaluate potential nanoparticles for HNSCC delivery, high-throughput delivery assays performed directly in vivo have not been reported. Here we use a high-throughput LNP assay to evaluate how 94 chemically distinct nanoparticles delivered nucleic acids to HNSCC solid tumors in vivo. DNA barcodes were used to identify LNPHNSCC, a novel LNP for systemic delivery to HNSCC solid tumors. Importantly, LNPHNSCC retains tropism to HNSCC solid tumors while minimizing off-target delivery to the liver.

SMYD2 Regulates Vascular Smooth Muscle Cell Phenotypic Switching and Intimal Hyperplasia via Interaction with Myocardin.

The SET and MYND domain-containing protein 2 (SMYD2) is a histone lysine methyltransferase that has been reported to regulate carcinogenesis and inflammation. However, its role in vascular smooth muscle cell (VSMC) homeostasis and vascular diseases has not been determined. Here, we investigated the role of SMYD2 in VSMC phenotypic modulation and vascular intimal hyperplasia and elucidated the underlying mechanism. We observed that SMYD2 expression was downregulated in injured carotid arteries in mice and phenotypically modulated VSMCs in vitro. Using a SMC-specific Smyd2 knockout mouse model, we found that Smyd2 ablation in VSMCs exacerbates neointima formation after vascular injury in vivo. Conversely, Smyd2 overexpression inhibits VSMC proliferation and migration in vitro and attenuates arterial narrowing in injured vessels in mice. Smyd2 downregulation promotes VSMC phenotypic switching accompanied with enhanced proliferation and migration. Mechanistically, genome-wide transcriptome analysis and loss/gain-of-function studies revealed that SMYD2 up-regulates VSMC contractile gene expression and suppresses VSMC proliferation and migration, in part, by promoting expression and transactivation of the master transcription cofactor myocardin. In addition, myocardin directly interacts with SMYD2, thereby facilitating SMYD2 recruitment to the CArG regions of SMC contractile gene promoters and leading to an open chromatin status around SMC contractile gene promoters via SMYD2-mediated H3K4 methylation. Hence, we conclude that SMYD2 is a novel regulator of VSMC contractile phenotype and intimal hyperplasia via a myocardin-dependent epigenetic regulatory mechanism and may be a potential therapeutic target for occlusive vascular diseases.

Loss of the endocytic tumor suppressor HD-PTP phenocopies LKB1 and promotes RAS-driven oncogenesis.

Oncogenic RAS mutations drive aggressive cancers that are difficult to treat in the clinic, and while direct inhibition of the most common KRAS variant in lung adenocarcinoma (G12C) is undergoing clinical evaluation, a wide spectrum of oncogenic RAS variants together make up a large percentage of untargetable lung and GI cancers. Here we report that loss-of-function alterations (mutations and deep deletions) in the gene that encodes HD-PTP (PTPN23) occur in up to 14% of lung cancers in the ORIEN Avatar lung cancer cohort, associate with adenosquamous histology, and occur alongside an altered spectrum of KRAS alleles. Furthermore, we show that in publicly available early-stage NSCLC studies loss of HD-PTP is mutually exclusive with loss of LKB1, which suggests they restrict a common oncogenic pathway in early lung tumorigenesis. In support of this, knockdown of HD-PTP in RAS-transformed lung cancer cells is sufficient to promote FAK-dependent invasion. Lastly, knockdown of the Drosophila homolog of HD-PTP (dHD-PTP/Myopic) synergizes to promote RAS-dependent neoplastic progression. Our findings highlight a novel tumor suppressor that can restrict RAS-driven lung cancer oncogenesis and identify a targetable pathway for personalized therapeutic approaches for adenosquamous lung cancer.

Plasma Metabolomics Reveals ß-Glucan Improves Muscle Strength and Exercise Capacity in Athletes.

The present study aimed to assess the changes in muscle strength and plasma metabolites in athletes with ß-glucan supplementation. A total of 29 athletes who met the inclusion criteria were recruited for this study (ChiCTR2200058091) and were randomly divided into a placebo group (n = 14) and ß-glucan group (n = 15). During the trial, the experimental group received ß-glucan supplementation (2 g/d ß-glucan) for 4 weeks and the control group received an equal dose of placebo supplementation (0 g/d ß-glucan), with both groups maintaining their regular diet and exercise habits during the trial. The athletes' exercise performance, muscle strength, and plasma metabolome changes were analyzed after 4 weeks of ß-glucan supplementation. The results showed a significant increase in mean grip strength (kg), right hand grip strength (kg), left triceps strength (kg), and upper limb muscle mass (kg) in the experimental group after the 4-week intervention compared to the preintervention period (p < 0.05). A comparison of the difference between the two groups after the intervention showed that there were significant differences between the control group and the experimental group in mean grip strength (kg) and right-hand grip strength (kg) (p < 0.05). Athletes in the experimental group showed significant improvements in 1 min double rocking jump (pcs), VO2max (ml/kg-min) (p < 0.05). The ß-glucan intake increased the creatine-related pathway metabolites in plasma. Overall, these results suggest that 4 weeks of ß-glucan supplementation can improve muscle strength in athletes, with the potential to increase aerobic endurance and enhance immune function, possibly by affecting creatine-related pathways.

Epithelial SMYD5 Exaggerates IBD by Down-regulating Mitochondrial Functions via Post-Translational Control of PGC-1α Stability.

BACKGROUND & AIMS: The expression and role of methyltransferase SET and MYND domain-containing protein 5 (SMYD5) in inflammatory bowel disease (IBD) is completely unknown. Here, we investigated the role and underlying mechanism of epithelial SMYD5 in IBD pathogenesis and progression. METHODS: The expression levels of SMYD5 and the mitochondrial transcriptional coactivator peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) were examined by Western blot, immunofluorescence staining, and immunohistochemistry in intestinal epithelial cells (IECs) and in colon tissues from human IBD patients and colitic mice. Mice with Smyd5 conditional knockout in IECs and littermate controls were subjected to dextran sulfate sodium-induced colitis and the disease severity was assessed. SMYD5-regulated mitochondrial biogenesis was examined by quantitative reverse-transcription polymerase chain reaction and transmission electron microscopy, and the mitochondrial oxygen consumption rate was measured in a Seahorse Analyzer system (Agilent, Santa Clara, CA). SMYD5 and PGC-1α interaction was determined by co-immunoprecipitation assay. PGC-1α degradation and turnover (half-life) were analyzed by cycloheximide chase assay. SMYD5-mediated PGC-1α methylation was assessed via in vitro methylation assay followed by mass spectrometry for identification of methylated lysine residues. RESULTS: Up-regulated SMYD5 and down-regulated PGC-1α were observed in intestinal epithelia from IBD patients and colitic mice. Smyd5 depletion in IECs protected mice from dextran sulfate sodium-induced colitis. SMYD5 was critically involved in regulating mitochondrial biology such as mitochondrial biogenesis, respiration, and apoptosis. Mechanistically, SMYD5 regulates mitochondrial functions in a PGC-1α-dependent manner. Furthermore, SMYD5 mediates lysine methylation of PGC-1α and subsequently facilitates its ubiquitination and degradation. CONCLUSIONS: SMYD5 attenuates mitochondrial functions in IECs and promotes IBD progression by enhancing PGC-1α degradation in a methylation-dependent manner. Strategies to decrease SMYD5 expression and/or increase PGC-1α expression in IECs might be a promising therapeutic approach to treat IBD patients.

The Efficacy of Tai Chi and Stretching Exercises Based on a Smartphone Application for Patients With Parkinson's Disease: A Protocol for a Randomized Controlled Trial.

Introduction: Parkinson's disease (PD) is a common neurodegenerative disease that seriously impairs patients' quality of life, and increases the burden of patients and caregivers. Both drugs and exercise can alleviate its motor and non-motor symptoms, improving the quality of life for PD patients. Telehealth, an increasingly popular tool, makes rehabilitation accessible at home, overcoming the inconvenience of traffic and scheduling. Care-PD is a phone application designed for rehabilitation training, which provides Tai Chi and stretching exercises through tutorial videos as well as an online evaluation system. In this protocol, we will explore the efficacy of Tai Chi and stretching exercises as a PD rehabilitation therapy based on the smartphone application Care-PD. Methods and Analysis: A double-blind, parallel randomized controlled trial will be conducted in this study. The recruitment, intervention, and evaluation processes will be implemented through the Care-PD application. Persons with PD will fill out questionnaires on Activities of Daily Living (ADL), upload the latest case report, and sign the informed consent form in the application. Afterward, doctors and researchers will screen and enroll 180 participants who will be randomly (1:1:1) assigned to Tai Chi group, stretching exercises group, or control group. The subjects will participate in a 1-h exercise session three times per week for 12 weeks, ending with another 4 weeks of follow-up study. Each exercise session includes 10 min of warm-up, 45 min of exercise, and 5 min of cool-down. The primary outcomes are Motor Aspects of Experiences of Daily Living and the 39-item Parkinson's disease Questionnaire. The secondary outcomes include the 9-item Wearing-Off Questionnaire, the Freezing of Gait Questionnaire, the Caregiver Strain Index, Non-motor Experiences of Daily Living, ADL, and Morse Fall Scale. All assessments will be performed at baseline, week 12 and 16. Discussion: Care-PD integrates subject recruitment, intervention, and evaluation, providing a new perspective on clinical rehabilitation for persons with PD. This study will evaluate the efficacy of Tai Chi and stretching exercises on patients' quality of life and disease progression based on a smartphone application. We aim to provide a new rehabilitation training platform for persons with PD. Ethics and Dissemination: This study was approved by the Scientific Research Ethics Committee (102772020RT132) of Shanghai University of Sport. Data collection begins after the approval of the ethics committee. The participants must sign an informed consent form before enrollment. The results will be published in relevant journals, seminars, and be disseminated among rehabilitation practitioners and patients with PD. Clinical Trial Registration: Chinese Clinical Trial Registry, identifier [ChiCTR2100042096]. Registered on January 13, 2021.

The Effect of Low and Moderate Exercise on Hyperuricemia: Protocol for a Randomized Controlled Study.

Background: Hyperuricemia (HUA) is a metabolic disease by purine metabolism disorders. It is a risk factor for many chronic diseases, including diabetes, hypertension, and heart disease. Studies have shown that exercise can effectively reduce serum uric acid (SUA), but the optimal exercise dose, intensity, and mode of exercise for improving HUA have not been verified in clinical studies. Therefore, this study aims to explore the effect of different exercise intensities in improving SUA of patients with HUA. Methods and Analysis: A randomized, single-blind, parallel controlled trial will be conducted in this study. 186 HUA patients who meet the inclusion criteria will be randomly divided into a 1:1:1 ratio (1): control group (2), low-intensity exercise group (brisk walking, 57-63% maximum heart rate, 150 min/week, 12 months), and (3) moderate-intensity exercise group (jogging, 64-76% maximum heart rate, 150 min/week, 12 months). The three groups of subjects will receive the same health education and prohibition of high-purine diet during the intervention period. The primary outcomes will be SUA concentration, SUA concentration change (mg/dL), SUA change rate (%), and the proportion of HUA patients. Secondary outcomes will include anthropometric parameters (body weight, waist circumference, hip circumference, BMI); physiological indicators (blood pressure, grip, vital capacity, maximum oxygen); biochemical indicators (blood lipid, blood sugar, liver enzyme, creatinine, and blood urea nitrogen). Each group of patients will go through an assessment at baseline, 3rd, 6th, and 12th months. Discussion: This study will evaluate the effect of 12-month low-intensity exercise and moderate-intensity exercise on HUA patients. We hypothesize that both low-intensity and moderate-intensity exercise would improve HUA as compared with no-exercise control, and that moderate-intensity exercise would be more effective than low-intensity exercise in improving HUA. These results can provide a basis for the current physical activity guidelines for HUA's healthy lifestyle management. Ethics and Dissemination: This study has been approved by the Ethical Review Committee of the Shanghai University of Sport (approval number: 102772020RT005). Informed consent will be obtained from all participants or their guardians. The authors intend to submit the study findings to peer-reviewed journals or academic conferences to be published. Clinical Trial Registration: Chinese Clinical Trial Registry, identifier ChiCTR2100042643.

A Novel Management Platform Based on Personalized Home Care Pathways for Medicine Management and Rehabilitation of Persons With Parkinson's Disease-Requirements and Implementation Plan of the Care-PD Program.

As the percentage of the aging population increases, the incidence of Parkinson's disease (PD) in China is increasing year by year. PD is both a public health and social problem facing the government and society as a whole. Persons with PD need reasonable medication management and rehabilitation strategies after a clear diagnosis. A proper home care plan can effectively slow the progression of PD. However, people with PD lack an effective way to manage their illnesses and cannot achieve the recommended clinical path in a family environment. Medication management, condition monitoring, and rehabilitation training are important components of the home care plan for PD. Persons with PD require strategies that delay the development of the disease and to adhere to treatment, which would contribute to improving their quality of life. Thus, we developed a small program called Care-PD to build a medicine management and service platform for PD. The development of Care-PD is a multi-dimensional model designed for PD, which is funded by the National Key R&D Program of China (No. 2018YFC1314700), and includes services such as medication management, symptom monitoring, professional counseling, home life, and community communication. Care-PD can become a key technology that increases the compliance of persons with PD with home care plans and improve measures to control the disease. In this article, we describe the medication management and services for PD based on the Care-PD program and its structure. The small program will improve the adverse conditions faced by persons with PD by combining the latest technology and clinical approaches. Meanwhile, we describe a verification strategy to evaluate the effectiveness of the Care-PD program as a comprehensive management strategy for PD.

Functional Validation of Phytoene Synthase and Lycopene ε-Cyclase Genes for High Lycopene Content in Autumn Olive Fruit (Elaeagnus umbellata).

Lycopene is the most potent antioxidant among all carotenoids and is beneficial to human health. A ripe fruit of autumn olive (Elaeagnus umbellata Thunb.) accumulates a high level of lycopene, which is 5-20 times higher than that in an ordinary tomato fruit. During fruit ripening of autumn olive, only phytoene synthase (EutPSY) expression pattern shows a tight positive correlation with the increased lycopene content observed at four ripening stages, while the lycopene ε-cyclase (EutLCYe) transcript could not be detected throughout fruit ripening. Here, we investigated whether the two genes are important targets for engineering lycopene biosynthesis. The full-length cDNAs of EutPSY and EutLCYe were first isolated. Fruit-specific overexpression of EutPSY in tomato fruits resulted in elevated contents of lycopene and ß-carotene through feedforward regulation of carotenogenic genes, i.e., downregulation of SlLCYe and upregulation of SlLCYb and SlCYCB. These fruits were decreased in ethylene production throughout ripening. Transcript levels of genes for system-2 ethylene synthesis (SlACS2, SlACS4, SlACO1, and SlACO3), perception (SlNR/ETR3 and SlETR4), and response (SlE4 and SlE8) were also inhibited in EutPSY-overexpressing fruits. Repressing ethylene synthesis and signaling transduction delayed fruit climacteric ripening of transgenic tomato plants. Additionally, RNAi suppression of SlLCYe enhanced ß-carotene but not lycopene accumulation through altered expression of carotenogenic genes in transgenic tomato fruits by both feedforward and feedback regulatory mechanisms. Ethylene production in SlLCYe-RNAi fruits decreased, thereby delaying fruit ripening. Collectively, these results confirmed that transcriptional regulation of EutPSY and EutLCYe plays a crucial role and a part in massive lycopene accumulation in autumn olive fruits, respectively. EutPSY overexpression enhanced lycopene accumulation in tomato fruits independently of the ethylene pathway but did not influence the size and weight of tomato fruits. EutPSY can be used as an effective strategy capable of elevating the lycopene content in fruits for improving quality.

Hepatic NPC1L1 overexpression attenuates alcoholic autophagy in mice.

Alcohol consumption causes liver steatosis in humans. Metabolic disorders of lipids are one of the factors that cause liver steatosis in hepatocytes. Hepatic Niemann­Pick C1­like 1 (NPC1L1) regulates lipid homeostasis in mammals. The relationship between NPC1L1 and autophagy in those with a history of alcohol abuse is unclear. The present study aimed to investigate the function of NPC1L1 in the activation of hepatic autophagy in a mouse model with a human (h)NPC1L1 transgene under alcohol feeding conditions. The mice expressing hNPC1L1 (Ad­L1) or controls (Ad­null) were created by retro­orbital adenovirus injection. The Ad­L1 and Ad­null mice were fed with alcohol or a non­alcoholic diet to mimic chronic alcohol consumption in humans. Hepatic autophagy was demonstrated in isolated primary hepatocytes by monitoring autophagic vacuoles under fluorescence microscopy, and by western blotting for autophagic makers. Isolated hepatocytes from the livers of Ad­L1 mice were treated with different doses of ezetimibe to study the restoration of autophagy. Chronic alcohol feeding caused liver injury and steatosis, shown by significantly higher levels of plasma alanine transaminase and aspartate transaminase activity, and by hematoxylin and eosin staining in Ad­L1 and Ad­null mice. Compared to Ad­null control mice, the microtubule­associated proteins 1A/1B light chain 3 (LC3) particles in the isolated hepatocytes of Ad­L1 mice were decreased, both under alcohol and non­alcoholic feeding. The ratio of LC3II/LC3I was significantly decreased, and the level of p62/sequestosome­1 protein was significantly increased in Ad­L1 mice compared with Ad­null mice after alcohol feeding. Levels of LC3II protein were statistically increased in hepatocytes isolated from Ad­L1 mice with ezetimibe treatment. The increase in LC3II expression was dose dependent. Within the tested range, it reached its highest level at 40 µM. The livers of Ad­L1 mice represent a more human­like state for the study of hepatic autophagy. Hepatic expression of human NPC1L1 resulted in an inhibition of autophagy; it may contribute to alcoholic fatty liver disease in humans.

Stem/Progenitor Cells and Their Therapeutic Application in Cardiovascular Disease.

Cardiovascular disease is the leading cause of death in the world. The stem/progenitor cell-based therapy has emerged as a promising approach for the treatment of a variety of cardiovascular diseases including myocardial infarction, stroke, peripheral arterial disease, and diabetes. An increasing number of evidence has shown that stem/progenitor cell transplantation could replenish damaged cells, improve cardiac and vascular functions, and repair injured tissues in many pre-clinical studies and clinical trials. In this review, we have outlined the major types of stem/progenitor cells, and summarized the studies in applying these cells, especially endothelial stem/progenitor cells and their derivatives, in the treatment of cardiovascular disease. Here the strategies used to improve the stem/progenitor cell-based therapies in cardiovascular disease and the challenges with these therapies in clinical applications are also reviewed.

Disruption of GPR35 Exacerbates Dextran Sulfate Sodium-Induced Colitis in Mice.

BACKGROUND: G protein-coupled receptor 35 (GPR35) is an orphan receptor and is vastly expressed in immune cells and gastrointestinal cells, suggesting the potential physiological importance of GPR35 in these cells. Here, we tested the hypothesis that the lack of GPR35 expression in the colon mucosa exacerbates the severity of dextran sulfate sodium (DSS)-induced experimental colitis in mice. METHODS: Colitis was induced in GPR35 wild-type (GPR35+/+) and GPR35 knockout (GPR35-/-) mice through the administration of DSS in drinking water for 5 days followed by regular facility water for 1 day. Induction of colitis was evaluated by measuring relative body weight loss, clinical illness scores, and morphological changes in the colon. Abolition of Gpr35 gene expression in the colon mucosa of GPR35-/- mice was confirmed by quantitative real-time PCR (qPCR). Gene expressions of inflammatory and tissue remodeling cytokines were detected by qPCR. Human colorectal epithelial Caco cells were transfected with siRNA against GPR35 before treated with 1% DSS in vitro. Protein expressions were measured using Western blot. RESULTS: GPR35-/- mice receiving DSS showed a significantly worsened colitis disease with profound loss of body weight and a considerable amount of severe clinical illness compared to GPR35+/+ mice that received DSS. The histology of colon sections from GPR35-/- mice showed extensive pathological changes including submucosal edema, diffuse ulcerations, and evidence of complete loss of crypts compared to wild-type mice. The mean histopathological score was significantly higher in GPR35-/- mice as compared to GPR35+/+ mice. The qPCR data revealed significant expression of pro-inflammatory and tissue remodeling cytokines in GPR35-/- colon mucosa, including IL-1ß, CXCL1, CXCL2, CCL2, HMGB1, TGFß1, TGFß3, MMP1/9/12. The protein expressions of Zonula occludens-1, E-cadherin, Claudin1 were decreased upon knocking down GPR35 with or without 1% DSS treatment. CONCLUSIONS: Our experimental data suggest that lack of GPR35 resulted in worsened disease outcome in DSS-induced experimental colitis, indicating that GPR35 could play a crucial role in protecting from colonic inflammation and serve as a therapeutic target.

PNT2258, a novel deoxyribonucleic acid inhibitor, induces cell cycle arrest and apoptosis via a distinct mechanism of action: a new class of drug for non-Hodgkin's lymphoma.

Current therapy for BCL-2-associated tumors such as Non-Hodgkin Lymphomas (NHL) is inadequate. The DNAi PNT2258, a 24 base single-stranded phosphodiester DNA oligodeoxynucleotide (PNT100) encapsulated in a protective liposome, was precisely designed to treat cancers that over-express BCL-2. PNT2258 strongly inhibited BCL-2 promoter activity, confirming its predicted mechanism of action. BCL-2 mRNA and protein expression were significantly downregulated in a follicular small cleaved cell lymphoma (WSU-FSCCL) cell line. 2.5µM PNT2258 induced an initial S- phase arrest followed by a gradual increase in the sub-G0 (apoptosis) compartment and a reciprocal progressive decrease of the S phase. Terminal deoxynucleotidyl transferase (TdT)-positive populations and cleaved caspase-3 and PARP were also increased. The data are consistent with the idea that BCL-2 inhibition by PNT2258 activates apoptotic pathways in WSU-FSCCL cells. This is the first report to address the distinct mechanism of action underlying the anti-BCL-2 functions of PNT2258. Growth inhibition in two other cell lines, WSU-DLCL2 and WSU-WM, supports broad applicability of BCL-2 DNAi to treatment of B-cell NHL.

Emerging role of cystic fibrosis transmembrane conductance regulator - an epithelial chloride channel in gastrointestinal cancers.

Cystic fibrosis transmembrane conductance regulator (CFTR), a glycoprotein with 1480 amino acids, has been well established as a chloride channel mainly expressed in the epithelial cells of various tissues and organs such as lungs, sweat glands, gastrointestinal system, and reproductive organs. Although defective CFTR leads to cystic fibrosis, a common genetic disorder in the Caucasian population, there is accumulating evidence that suggests a novel role of CFTR in various cancers, especially in gastroenterological cancers, such as pancreatic cancer and colon cancer. In this review, we summarize the emerging findings that link CFTR with various cancers, with focus on the association between CFTR defects and gastrointestinal cancers as well as the underlying mechanisms. Further study of CFTR in cancer biology may help pave a new way for the diagnosis and treatment of gastrointestinal cancers.

Dysregulated Chemokine Signaling in Cystic Fibrosis Lung Disease: A Potential Therapeutic Target.

CF lung disease is characterized by a chronic and non-resolving activation of the innate immune system with excessive release of chemokines/cytokines including IL-8 and persistent infiltration of immune cells, mainly neutrophils, into the airways. Chronic infection and impaired immune response eventually lead to pulmonary damage characterized by bronchiectasis, emphysema, and lung fibrosis. As a complete knowledge of the pathways responsible for the exaggerated inflammatory response in CF lung disease is lacking, understanding these pathways could reveal new therapeutic targets, and lead to novel treatments. Therefore, there is a strong rationale for the identification of mechanisms and pathways underlying the exaggerated inflammatory response in CF lung disease. This article reviews the role of inflammation in the pathogenesis of CF lung disease, with a focus on the dysregulated signaling involved in the overexpression of chemokine IL-8 and excessive recruitment of neutrophils in CF airways. The findings suggest that targeting the exaggerated IL-8/IL-8 receptor (mainly CXCR2) signaling pathway in immune cells (especially neutrophils) may represent a potential therapeutic strategy for CF lung disease.

A critical role of CXCR2 PDZ-mediated interactions in endothelial progenitor cell homing and angiogenesis.

Bone marrow-derived endothelial progenitor cells (EPCs) contribute to neovessel formation in response to growth factors, cytokines, and chemokines. Chemokine receptor CXCR2 and its cognate ligands are reported to mediate EPC recruitment and angiogenesis. CXCR2 possesses a consensus PSD-95/DlgA/ZO-1 (PDZ) motif which has been reported to modulate cellular signaling and functions. Here we examined the potential role of the PDZ motif in CXCR2-mediated EPC motility and angiogenesis. We observed that exogenous CXCR2 C-tail significantly inhibited in vitro EPC migratory responses and angiogenic activities, as well as in vivo EPC angiogenesis. However, the CXCR2 C-tail that lacks the PDZ motif (ΔTTL) did not cause any significant changes of these functions in EPCs. In addition, using biochemical assays, we demonstrated that the PDZ scaffold protein NHERF1 specifically interacted with CXCR2 and its downstream effector, PLC-ß3, in EPCs. This suggests that NHERF1 might cluster CXCR2 and its relevant signaling molecules into a macromolecular signaling complex modulating EPC cellular functions. Taken together, our data revealed a critical role of a PDZ-based CXCR2 macromolecular complex in EPC homing and angiogenesis, suggesting that targeting this complex might be a novel and effective strategy to treat angiogenesis-dependent diseases.

Crystal structure of the NHERF1 PDZ2 domain in complex with the chemokine receptor CXCR2 reveals probable modes of PDZ2 dimerization.

The formation of CXCR2-NHERF1-PLCß2 macromolecular complex in neutrophils regulates CXCR2 signaling and plays a key role in neutrophil chemotaxis and transepithelial neutrophilic migration. However, NHERF1 by itself, with only two PDZ domains, has a limited capacity in scaffolding the multiprotein-complex formation. Here we report the crystal structure of the NHERF1 PDZ2 domain in complex with the C-terminal CXCR2 sequence. The structure reveals that the PDZ2-CXCR2 binding specificity is achieved by numerous hydrogen bonds and hydrophobic contacts with the last four CXCR2 residues contributing to specific interactions. The structure also reveals two probable modes of PDZ2 dimerization where the two canonical ligand-binding pockets are well separated and orientated in a unique parallel fashion. This study provides not only the structural basis for the PDZ-mediated NHERF1-CXCR2 interaction, but also an additional example of how PDZ domains may dimerize, which both could prove valuable in understanding NHERF1 complex-scaffolding function in neutrophils.

Crystallographic analysis of NHERF1-PLCß3 interaction provides structural basis for CXCR2 signaling in pancreatic cancer.

The formation of CXCR2-NHERF1-PLCß3 macromolecular complex in pancreatic cancer cells regulates CXCR2 signaling activity and plays an important role in tumor proliferation and invasion. We previously have shown that disruption of the NHERF1-mediated CXCR2-PLCß3 interaction abolishes the CXCR2 signaling cascade and inhibits pancreatic tumor growth in vitro and in vivo. Here we report the crystal structure of the NHERF1 PDZ1 domain in complex with the C-terminal PLCß3 sequence. The structure reveals that the PDZ1-PLCß3 binding specificity is achieved by numerous hydrogen bonds and hydrophobic contacts with the last four PLCß3 residues contributing to specific interactions. We also show that PLCß3 can bind both NHERF1 PDZ1 and PDZ2 in pancreatic cancer cells, consistent with the observation that the peptide binding pockets of these PDZ domains are highly structurally conserved. This study provides an understanding of the structural basis for the PDZ-mediated NHERF1-PLCß3 interaction that could prove valuable in selective drug design against CXCR2-related cancers.

New conformational state of NHERF1-CXCR2 signaling complex captured by crystal lattice trapping.

NHERF1 is a PDZ adaptor protein that scaffolds the assembly of diverse signaling complexes and has been implicated in many cancers. However, little is known about the mechanism responsible for its scaffolding promiscuity or its ability to bind to multiple targets. Computational studies have indicated that PDZ promiscuity may be attributed to its conformational dynamics, but experimental evidence for this relationship remains very limited. Here we examine the conformational flexibility of the NHERF1 PDZ1 domain using crystal lattice trapping via solving PDZ1 structure of a new crystal form. The structure, together with prior PDZ1 structures of a different space group, reveals that 4 of 11 ligand-interacting residues undergo significant crystal packing-induced structural changes. Most of these residues correspond to the residues involved in allosteric transition when a peptide ligand binds. In addition, a subtle difference in ligand conformations causes the same peptide to bind in slightly different modes in different crystal forms. These findings indicate that substantial structural flexibility is present in the PDZ1 peptide-binding pocket, and the structural substate trapped in the present crystal form can be utilized to represent the conformational space accessible to the protein. Such knowledge will be critical for drug design against the NHERF1 PDZ1 domain, highlighting the continued need for experimentally determined PDZ1-ligand complexes.