NQO1 Mediates Lenvatinib Resistance by Regulating ROS-induced Apoptosis in Hepatocellular Carcinoma.

OBJECTIVE: Hepatocellular carcinoma (HCC) is the third leading cause of cancer-associated death worldwide. As a first-line drug for advanced HCC treatment, lenvatinib faces a significant hurdle due to the development of both intrinsic and acquired resistance among patients, and the underlying mechanism remains largely unknown. The present study aims to identify the pivotal gene responsible for lenvatinib resistance in HCC, explore the potential molecular mechanism, and propose combinatorial therapeutic targets for HCC management. METHODS: Cell viability and colony formation assays were conducted to evaluate the sensitivity of cells to lenvatinib and dicoumarol. RNA-Seq was used to determine the differences in transcriptome between parental cells and lenvatinib-resistant (LR) cells. The upregulated genes were analyzed by GO and KEGG analyses. Then, qPCR and Western blotting were employed to determine the relative gene expression levels. Afterwards, the intracellular reactive oxygen species (ROS) and apoptosis were detected by flow cytometry. RESULTS: PLC-LR and Hep3B-LR were established. There was a total of 116 significantly upregulated genes common to both LR cell lines. The GO and KEGG analyses indicated that these genes were involved in oxidoreductase and dehydrogenase activities, and reactive oxygen species pathways. Notably, NAD(P)H:quinone oxidoreductase 1 (NQO1) was highly expressed in LR cells, and was involved in the lenvatinib resistance. The high expression of NQO1 decreased the production of ROS induced by lenvatinib, and subsequently suppressed the apoptosis. The combination of lenvatinib and NQO1 inhibitor, dicoumarol, reversed the resistance of LR cells. CONCLUSION: The high NQO1 expression in HCC cells impedes the lenvatinib-induced apoptosis by regulating the ROS levels, thereby promoting lenvatinib resistance in HCC cells.

Sustained Release of Dicumarol via Novel Grafted Polymer in Electrospun Nanofiber Membrane for Treatment of Peritendinous Adhesion.

The prevention and treatment of post-traumatic peritendinous adhesion (PA) have always been a great difficulty for orthopedic surgeons. Current treatments include resecting surgery, non-steroidal anti-inflammatory drugs (NSAIDs) usage and implantable membranes, often target single disease pathogenic processes, resulting in unfavorable therapeutic outcomes. Here a polylactic acid (PLA)-dicumarol conjugates-electrospun nanofiber membrane (ENM) (PCD) is generated, which can achieve spatial accuracy and temporal sustainability in drug release. It is further demonstrated that PCD possesses a significantly higher and more sustainable drug release profile than traditional drug-loading ENM. By providing a physical barrier and continuous releasing of dicumarol, PCD implantation significantly reduces tissue adhesion by 25%, decreases fibroblasts activity and inhibits key fibrogenic cytokine transforming growth factor beta (TGFß) production by 30%, and improves the biomechanical tendon property by 14.69%. Mechanistically, PCD potently inhibits the connexin43 (Cx43) and thereby tunes down the fibroblastic TGFß/Smad3 signaling pathway. Thus, this approach leverages the anti-adhesion effect of dicumarol and drug release properties of grafted copolymer ENM by esters to provide a promising therapeutic strategy for patients who suffer from PA.

ATP-releasing SWELL1 channel in spinal microglia contributes to neuropathic pain.

Following peripheral nerve injury, extracellular adenosine 5'-triphosphate (ATP)-mediated purinergic signaling is crucial for spinal cord microglia activation and neuropathic pain. However, the mechanisms of ATP release remain poorly understood. Here, we show that volume-regulated anion channel (VRAC) is an ATP-releasing channel and is activated by inflammatory mediator sphingosine-1-phosphate (S1P) in microglia. Mice with microglia-specific deletion of Swell1 (also known as Lrrc8a), a VRAC essential subunit, had reduced peripheral nerve injury-induced increase in extracellular ATP in spinal cord. The mutant mice also exhibited decreased spinal microgliosis, dorsal horn neuronal hyperactivity, and both evoked and spontaneous neuropathic pain-like behaviors. We further performed high-throughput screens and identified an FDA-approved drug dicumarol as a novel and potent VRAC inhibitor. Intrathecal administration of dicumarol alleviated nerve injury-induced mechanical allodynia in mice. Our findings suggest that ATP-releasing VRAC in microglia is a key spinal cord determinant of neuropathic pain and a potential therapeutic target for this debilitating disease.

Structure-based screening and biological validation of the anti-thrombotic drug-dicoumarol as a novel and potent PPARγ-modulating ligand.

PPARγ full agonists, thiazolidinediones (TZDs), have been known as a class of most effective drugs for the treatment of type 2 diabetes mellitus (T2DM). However, recently their therapeutic benefits have been compromised by several undesirable side effects. In this study, a host-based repurposing strategy and in combination with comprehensive biological evaluations were synergistically employed to seek for potent PPARγ ligands, which led to the identification of an anti-thrombotic drug, dicoumarol (Dic), as the novel and safer selectively PPARγ modulator (SPPARγM) with advantages over current TZD drugs. The results in vitro showed that Dic had a potent binding affinity and weakly agonistic activity for PPARγ and its downstream key genes. Moreover, in diabetic model, it significantly reduced blood glucose without leading to the weight gain of both body and main organ tissues. Further mechanistic investigations revealed that Dic possessed such desired pharmacological properties mainly through effectively inhibiting the phosphorylation of PPARγ-Ser273 and selectively regulating the expressions of insulin-sensitive and resistance genes. Finally, the docking studies on the analysis of the potent binding mode of Dic with PPARγ revealed a remarkable difference on interaction region compared with other developed PPARγ agonists, which not only gave a proof of concept for the abovementioned mechanism but also provided the molecular basis for the discrimination of Dic from other PPARγ ligands, especially TZD drugs. Taken together, our findings suggested that Dic could serve as a new and promising candidate with good therapeutic index for treating T2DM, especially for those T2DM patients with thrombosis.

A pharmacological review of dicoumarol: An old natural anticoagulant agent.

Dicoumarol is an oral anticoagulant agent prescribed in clinical for decades. It is a natural hydroxycoumarin discovered from the spoilage of Melilotus officinalis (L.) Pall and is originally discovered as a rodenticide. Due to its structural similarity to that of vitamin K, it significantly inhibits vitamin K epoxide reductase and acts as a vitamin K antagonist. Dicoumarol is mainly used as an anticoagulant to prevent thrombogenesis and to cure vascular thrombosis. Other biological activities besides anticoagulants such as anticancer, antimicrobial, antiviral, etc., have also been documented. The side effects of dicoumarol raise safety concerns for clinical application. In this review, the physicochemical property, the pharmacological activities, the side effects, and the pharmacokinetics of dicoumarol were summarized, aiming to provide a whole picture of the "old" anticoagulant.

Inhibition of pyruvate dehydrogenase kinase­1 by dicoumarol enhances the sensitivity of hepatocellular carcinoma cells to oxaliplatin via metabolic reprogramming.

The Warburg effect is a unique metabolic feature of the majority of tumor cells and is closely related to chemotherapeutic resistance. Pyruvate dehydrogenase kinase 1 (PDK1) is considered a 'switch' that controls the fate of pyruvate in glucose metabolism. However, to date, to the best of our knowledge, there are only a few studies to available which had studied the reduction of chemotherapeutic resistance via the metabolic reprogramming of tumor cells with PDK1 as a target. In the present study, it was found dicoumarol (DIC) reduced the phosphorylation of pyruvate dehydrogenase (PDH) by inhibiting the activity of PDK1, which converted the metabolism of human hepatocellular carcinoma (HCC) cells to oxidative phosphorylation, leading to an increase in mitochondrial reactive oxygen species ROS (mtROS) and a decrease in mitochondrial membrane potential (MMP), thereby increasing the apoptosis induced by oxaliplatin (OXA). Furthermore, the present study elucidated that the targeting of PDK1 may be a potential strategy for targeting metabolism in the chemotherapy of HCC. In addition, DIC as an 'old drug' exhibits novel efficacy, bringing new hope for antitumor therapy.

Impact of atrial fibrillation in critically ill patients admitted to a stepdown unit.

BACKGROUND: Limited data are available on the clinical course of patients with history of atrial fibrillation (AF) when admitted in an intensive care environment. We aimed to describe the occurrence of major adverse events in AF patients admitted to a stepdown care unit (SDU) and to analyse clinical factors associated with outcomes, impact of dicumarolic oral anticoagulant (OAC) therapy impact and performance of clinical risk scores in this setting. MATERIALS AND METHODS: Single-centre, observational retrospective analysis on a population of subjects with AF history admitted to a SDU. Therapeutic failure (composite of transfer to ICU or death) was considered the main study outcome. Occurrence of stroke and major bleeding (MH) was considered as secondary outcomes. The performance of clinical risk scores was evaluated. RESULTS: A total of 1430 consecutive patients were enrolled. 194 (13.6%) reported the main outcome. Using multivariate logistic regression, age (odds ratio [OR]: 1.03, 95% confidence interval [CI]: 1.01-1.05), acute coronary syndrome (OR:3.10, 95% CI: 1.88-5.12), cardiogenic shock (OR:10.06, 95% CI: 5.37-18.84), septic shock (OR:5.19,95%CI:3.29-18.84), acute respiratory failure (OR:2.49, 95% CI: 1.67-3.64) and OAC use (OR: 1.61, 95% CI: 1.02-2.55) were independently associated with main outcome. OAC prescription was associated with stroke risk reduction and to both MH and main outcome risk increase. CHA2 DS2 -VASc (c-index: 0.545, P = .117 for stroke) and HAS-BLED (c-index:0.503, P = .900 for MH) did not significantly predict events occurrence. CONCLUSIONS: In critically ill AF patients admitted to a SDU, adverse outcomes are highly prevalent. OAC use is associated to an increased risk of therapeutic failure, clinical scores seem unhelpful in predicting stroke and MH, suggesting a highly individualized approach in AF management in this setting.

Advanced Drug-Eluting Poly (Vinyl Chloride) Surfaces Deposited by Spin Coating.

Background and objectives: Medical devices such as catheters are used on a large scale to treat heart and cardiovascular diseases. Unfortunately, they present some important drawbacks (structure failure, calcifications, infections, thrombosis, etc.), with the main side effects occurring due to adhesion and proliferation of bacteria and living cells on the surface of the implanted devices. The aim of this work is to modify the surface of polyvinyl chloride (PVC), an affordable biocompatible material, in order to reduce these aforementioned side effects. Materials and Methods: The surface of PVC was modified by depositing a thin layer also of PVC that incorporates an active substance, dicoumarol (a well-known anticoagulant), by spin coating process. The modified surfaces were analyzed by Fourier-transform infrared (FT-IR) microscopy, Fourier-transform infrared (FT-IR) spectroscopy, Ultraviolet-visible spectroscopy (UV-VIS), and Scanning electron microscopy (SEM) in order to determine the surface morphology and behavior. The samples were tested for Gram-positive (S. aureus ATCC 25923) and Gram-negative (P. aeruginosa ATCC 27853) standard strains from American Type Culture Collection (ATCC). Results: The material obtained had a smooth surface with a uniform distribution of dicoumarol, which is released depending on the deposition parameters. The concentration of dicoumarol at the surface of the material and also the release rate is important for the applications for which the surface modification was designed. PVC modified using the proposed method showed a good ability to prevent salt deposition and decreased the protein adhesion, and the resistance to bacterial adherence was improved compared with standard PVC.

Targeting PSG1 to enhance chemotherapeutic efficacy: new application for anti-coagulant the dicumarol.

Chemotherapeutic response is critical for the successful treatment and good prognosis in cancer patients. In this study, we analysed the gene expression profiles of preoperative samples from oestrogen receptor (ER)-negative breast cancer patients with different responses to taxane-anthracycline-based (TA-based) chemotherapy, and identified a group of genes that was predictive. Pregnancy specific beta-1-glycoprotein 1 (PSG1) played a central role within signalling pathways of these genes. Inhibiting PSG1 can effectively reduce chemoresistance via a transforming growth factor-ß (TGF-ß)-related pathway in ER-negative breast cancer cells. Drug screening then identified dicumarol (DCM) to target the PSG1 and inhibit chemoresistance to TA-based chemotherapy in vitro, in vivo, and in clinical samples. Taken together, this study highlights PSG1 as an important mediator of chemoresistance, whose effect could be diminished by DCM.

[Perception of physicians on factors that influence the choice of a dicoumarin or a new oral anticoagulant in patients with non-valvular atrial fibrillation]. / Percepción de los médicos sobre los factores que influyen en la elección de un dicumarínico o de un nuevo anticoagulante oral en pacientes con fibrilación auricular no valvular.

AIMS: Recent studies have demonstrated the efficacy and safety of new oral anticoagulant drugs for the prevention of thromboembolic events in patients with non-valvular atrial fibrillation. Our aim was to evaluate the factors that can influence physicians in their choice between a classic and a new anticoagulant in these patients. DESIGN: Several variables of interest were discussed and analysed using a WorkmatTM methodology. SITES: Six regional meetings were held in Spain (East, Catalonia, Andalusia-Extremadura, Madrid, North-east, and North of Spain). PARTICIPANTS: Meetings were attended by 39 specialists (cardiologists, neurologists, haematologists, internists, and emergency and Primary Care physicians). MEASUREMENTS: Each participant graded their level of agreement, with a score from 1 to 10, on every analysed variable. RESULTS: A new anticoagulant drug was preferred in patients with previous failure of dicoumarin therapy (9.7±0.5), high haemorrhagic risk (8.7±1), prior bleeding (7.8±1.5), and high thrombotic risk (7.7±1.2). Dicoumarins were preferred in cases of severe (1.2±0.4) or moderate (4.2±2.5) kidney failure, good control with dicoumarins (2.3±1.5), cognitive impairment (3.2±3), and low haemorrhagic risk (4.3±3). Age, sex, weight, cost of drug, polymedication, and low thrombotic risk achieved intermediate scores. There were no differences between the different specialists or Spanish regions. CONCLUSIONS: The presence of a high thrombotic or haemorrhagic risk and the failure of previous dicoumarin therapy lead to choosing a new oral anticoagulant in patients with non-valvular atrial fibrillation, while kidney failure, cognitive impairment, good control with dicoumarins, and a low bleeding risk predispose to selecting a classic dicoumarin anticoagulant.

[Fournier's disease. Clinical case]. / La malattia di Fournier. Caso clinico.

Fournier's disease is a necrotizing fasciitis that prefers the male and which is located in the perineal area. In view of possible complications related to sepsis, systemic organ impairment and high mortality, is now considered a urological emergency. The main causes are infections of urethral and anorectal tract, immunodepression syndromes, diabetes mellitus and trauma. Infections are mixed with aerobic and anaerobic germs responsible of necrosis of tissue disorders due to the phenomena of necrotizing immunovasculitis disease. We present a case of Fournier's disease in perineal area, with gangrenosum necrotic evolution treated with antibiotic therapy, curettage of the necrotic tissue and local disinfection with saline and antiseptics solution. A total parenteral nutrition to ensure maximum perineal decontamination has been made. The extensive loss of substance scrotal was rebuilt in 25- day while the anal wound has healed spontaneously after complete surgical curettage.

Volumetric capnography: in the diagnostic work-up of chronic thromboembolic disease.

The morbidity and mortality of pulmonary embolism (PE) have been found to be related to early diagnosis and appropriate treatment. The examinations used to diagnose PE are expensive and not always easily accessible. These options include noninvasive examinations, such as clinical pretests, ELISA D-dimer (DD) tests, and volumetric capnography (VCap). We report the case of a patient whose diagnosis of PE was made via pulmonary arteriography. The clinical pretest revealed a moderate probability of the patient having PE, and the DD result was negative; however, the VCap associated with arterial blood gases result was positive. The patient underwent all noninvasive exams following admission to hospital and again eight months after discharge. Results gained from invasive tests were similar to those produced by image exams, highlighting the importance of VCap as an important noninvasive tool.

[Monitoring age-dependent effect of anticoagulation treatment in patients with atrial fibrillation]. / Sledování ucinnosti antikoagulacní lécby u nemocných s fibrilací síní v závislosti na veku.

Oral anticoagulation treatment with dicumarol preparations (warfarin sodium) is the standard in patients with atrial fibrillation. The effect of treatment depends on many factors, especially in elderly patients. In the study, we assessed the effect of treatment in patients with atrial fibrillation hospitalized in our cardiology ward from 2004 to 2005, in the form of a telephone survey (who controlled the treatment--general practitioner or internist?, the last 2 INR results, complications). INR 2.0-3.5 is considered an efficient therapeutic range. The proportion of permanently correctly anticoagulated patients is approximately 47% across the whole age range, the hypothesis of lower efficiency of treatment in elderly patients does not apply (48% of efficiently anticoagulated patients younger than 75 years vs. 46% of older patients--however, the study does not include polymorbid patients who could not take warfarin at all!) The fact whether a patient is monitored by a general practitioner or an outpatient specialist does not make any difference (49% of anticoagulated patients monitored by a general practitioner vs. 52% of patients monitored by an internist). The percentage of severe complications is relatively low (3.4%).

Mechanical valve replacement in congenital heart defects in the era of international normalized ratio self-management.

As the number of patients with congenital heart defects requiring heart valve replacement increases, the need for durable valve substitutes with good hemodynamic performance and a low incidence of complications becomes more apparent. The use of porcine xenografts is burdened with early fibrocalcific degeneration, whereas the use of mechanical heart valves led to an increased number of thromboembolic events, especially when implanted in the right side of the heart. We report on our experiences implanting bileaflet heart valves in congenital heart defects since the introduction of international normalized ratio (INR) self-management. The data of 68 long-term survivors (33 males, 35 females) who underwent mechanical heart valve replacement in congenital heart defect were reviewed. Patient age at the time of valve replacement ranged from 5 months to 61 years (mean 21 years). Underlying diagnoses were tetralogy of Fallot (n=33), morbus Ebstein (n=4), atrioventricular canal (n=13), truncus arteriosus communis (n=5), transposition of the great arteries (n=10), and congenitally corrected transposition of the great arteries (n=3). In all patients, bileaflet valves were implanted (St. Jude Medical n=40, Carbomedics n=28). Anticoagulation was performed using dicumarol (Marcumar) and INR self-management in all cases. The mean follow-up was 72 months (range 6-132 months; 409 patient-years). Valve thrombosis developed in 3 of 68 patients (4.4%, all three had tetralogy of Fallot, mean age 9.8 years) after a mean follow-up of 3.5 years. In two of these three patients, re-pulmonary valve replacement was necessary, whereas the third patient was treated by thrombolysis. From our experience, we conclude that mechanical heart valve replacement is a good therapy option with a low complication rate for patients with congenital heart defects requiring valve replacement, especially when INR self-management is performed.

Heat-induced up-regulation of NAD(P)H:quinone oxidoreductase potentiates anticancer effects of beta-lapachone.

PURPOSE: The purpose of the present study was to evaluate the efficacy of mild hyperthermia to potentiate the anticancer effects of beta-lapachone (3,4-dihydro-2,2-dimethyl-2H-naphthol[1,2-b]pyran-5,6-dione) by up-regulating NAD(P)H:quinone oxidoreductase (NQO1) in cancer cells. EXPERIMENTAL DESIGN: Effects of beta-lapachone alone or in combination with mild heating on the clonogenic survival of FSaII fibrosarcoma cells of C3H mice and A549 human lung tumor cells in vitro was determined. Effects of heating on the NQO1 level in the cancer cells in vitro were assessed using Western blot analysis for NQO1 expression, biochemical determination of NQO1 activity, and immunofluorescence microscopy for NQO1 expression. Growth of FSaII tumors in the hind legs of C3H mice was determined after treating the host mice with i.p. injection of 45 mg/kg beta-lapachone followed by heating the tumors at 42 degrees C for 1 hour every other day for four times. RESULTS: Incubation of FSaII tumor cells and A549 tumor cells with beta-lapachone at 37 degrees C reduced clonogenic survival of the cells in dose-dependent and incubation time-dependent manner. NQO1 level in the cancer cells in vitro increased within 1 hour after heating at 42 degrees C for 1 hour and remained elevated for >72 hours. The clonogenic cell death caused by beta-lapachone increased in parallel with the increase in NQO1 levels in heated cells. Heating FSaII tumors in the legs of C3H mice enhanced the effect of i.p.-injected beta-lapachone in suppressing tumor growth. CONCLUSION: We observed for the first time that mild heat shock up-regulates NQO1 in tumor cells. The heat-induced up-regulation of NQO1 enhanced the anticancer effects of beta-lapachone in vitro and in vivo.

[Recurrent vascular access trombosis associated with the prothrombin mutation G20210A in a adult patient in haemodialysis]. / Trombosis recurrente del acceso vascular asociada a la mutación G202 10A del gen de la protrombina en un paciente adulto en hemodiálisis.

Vascular access-related complications are a frequent cause of morbidity in haemodialysis patients and generate high costs. We present the case of an adult patient with end-stage renal disease and recurrent vascular access thrombosis associated with the prothrombin mutation G20210A and renal graft intolerance. The clinical expression of this heterozygous gene mutation may have been favoured by inflammatory state, frequent in dialysis patients. In this patient, the inflammatory response associated with the renal graft intolerance would have favored the development of recurrent vascular access thrombosis in a adult heterozygous for prothrombin mutation G20210A. In the case of early dysfunction of haemodialysis vascular access and after ruling out technical problems, it is convenient to carry out a screening for thrombophilia.

[Recurrent hemoptysis secondary to an aortobronchial fistula]. / Hemoptisis recurrente secundaria a una fístula aortobronquial.

Aortobronchial fistula is a rare but serious cause of hemoptysis. It can develop from an aneurysm of the descending thoracic aorta in the context of infections or it may appear as a sequel of surgical repair of congenital heart defects. Presenting symptoms include mild bronchial hemorrhages and recurrent chest pain, culminating in a normally fatal massive hemorrhage. Diagnosis by imaging is not always conclusive and clinical suspicion based on medical history is essential. Surgical placement of an endovascular stent graft is the treatment of choice. Post-surgical prognosis is good although there is a risk of recurrence in the case of superinfection.

Dietary induction of NQO1 increases the antitumour activity of mitomycin C in human colon tumours in vivo.

The bioreductive antitumour agent, mitomycin C (MMC), requires activation by reductive enzymes like NAD(P)H:quinone oxidoreductase 1 (NQO1). We used a novel approach to increase MMC efficacy by selectively inducing NQO1 in tumour cells in vivo. CD-1 nude mice were implanted with HCT116 cells, and fed control diet or diet containing 0.3% of the NQO1 inducer, dimethyl fumarate (DMF). The mice were then treated with saline, 2.0, 3.5 or 2.0 mg kg(-1) MMC and dicoumarol, an NQO1 inhibitor. The DMF diet increased NQO1 activity by 2.5-fold in the tumours, but had no effect in marrow cells. Mice given control diet/2.0 mg kg(-1) MMC had tumours with the same volume as control mice; however, mice given DMF diet/2.0 mg kg(-1) MMC had significantly smaller tumours. Tumour volumes in mice given DMF/2.0 mg kg(-1) MMC were similar to those in mice given control diet/3.5 mg kg(-1) MMC. Tumour inhibition was partially reversed in mice given DMF/2.0 mg kg(-1) MMC and dicoumarol. DMF diet/2.0 mg kg(-1) MMC treatment did not increase myelosuppression and did not produce any organ toxicity. These results provide strong evidence that dietary inducers of NQO1 can increase the antitumour activity of bioreductive agents like MMC without increasing toxicity.

[Superior vena cava thrombosis in a patient on hemodialysis]. / Trombosis de la vena cava superior en un paciente en hemodiálisis.

We present a patient with end-stage renal disease on maintenace hemodialysis through a permanent catheter (Permcath) on the right subclavian vein. One month after the catheter placement the patient exhibited a superior vena cava syndrome due to a pericatheter thrombosis. The patient was initially managed with anticoagulation with early clinical improvement. Nevertheless, the reappearance of the symptoms forced the removal of the catheter and percutaneous angioplasty of the superior vena cava. After those measures and anticoagulation with coumarin the patient remains stable with complete clinical resolution and angiographical improvement.