Exposure of human immune cells, to the antiretrovirals efavirenz and lopinavir, leads to lower glucose uptake and altered bioenergetic cell profiles through interactions with SLC2A1.

SLC2A1 mediates glucose cellular uptake; key to appropriate immune function. Our previous work has shown efavirenz and lopinavir exposure inhibits T cell and macrophage responses, to known agonists, likely via interactions with glucose transporters. Using human cell lines as a model, we assessed glucose uptake and subsequent bioenergetic profiles, linked to immunological responses. Glucose uptake was measured using 2-deoxyglucose as a surrogate for endogenous glucose, using commercially available reagents. mRNA expression of SLC transporters was investigated using qPCR TaqMan™ gene expression assay. Bioenergetic assessment, on THP-1 cells, utilised the Agilent Seahorse XF Mito Stress test. In silico analysis of potential interactions between SLC2A1 and antiretrovirals was investigated using bioinformatic techniques. Efavirenz and lopinavir exposure was associated with significantly lower glucose accumulation, most notably in THP-1 cells (up to 90% lower and 70% lower with efavirenz and lopinavir, respectively). Bioenergetic assessment showed differences in the rate of ATP production (JATP); efavirenz (4 µg/mL), was shown to reduce JATP by 87% whereas lopinavir (10 µg/mL), was shown to increase the overall JATP by 77%. Putative in silico analysis indicated the antiretrovirals, apart from efavirenz, associated with the binding site of highest binding affinity to SLC2A1, similar to that of glucose. Our data suggest a role for efavirenz and lopinavir in the alteration of glucose accumulation with subsequent alteration of bioenergetic profiles, supporting our hypothesis for their inhibitory effect on immune cell activation. Clarification of the implications of this data, for in vivo immunological responses, is now warranted to define possible consequences for these, and similar, therapeutics.

Prevalence and predictors of glucose metabolism disorders among People Living with HIV on combination antiretroviral therapy.

OBJECTIVE: We investigated prevalence and predictors of glucose metabolism disorders (GMDs) among People Living with HIV (PLWH) on efavirenz- and atazanavir/ritonavir-based combination antiretroviral therapy (cART). METHODS: This cross-sectional study involved adult PLWH on efavirenz- (n = 240) and atazanavir/ritonavir-based (n = 111) cART. The prevalence of GMDs was determined by fasting serum glucose, insulin, and homeostasis model assessment. A logistic regression model was used to determine predictors. RESULTS: The overall prevalence of GMDs for all regimens was 27.6% (97/351) [95% CI 23.0-32.6%] s, with 31.1% (75/240) [95% CI 25.4-37.5%] for efavirenz-based and 19.8% (22/111) [95% CI 12.9-28.5%)] for atazanavir/ritonavir-based cART group. The prevalence of impaired fasting glycemia was significantly higher (p = 0.026) in the efavirenz- [(15.4%) (37/240); 95%CI (11.1-20.6%)] than atazanavir/ritonavir-based [(7.2%) (8/111), (95%CI (3.2-13.7%)] cART. However, no significant difference was observed in the prevalence of diabetes mellitus and insulin resistance between the two regimens. Age ≥46 years old and specific type of ARV contained in cART, such as TDF, were independent predictors of GMD in both groups. Whereas the male gender and BMI category were predictors of GMDs among EFV-based cART group, AZT- and ABC- containing regimens and triglyceride levels were predictors in the ATV/r-based group. CONCLUSIONS: GMDs were highly prevalent among adults on EFV- than ATV/r-based cARTs. Age ≥46 years and TDF-containing cARTs are common predictors in both regimens. Close monitoring for impaired fasting glucose during long-term EFV-based cART is recommended for early diagnosis of type-2 diabetes and management.

De novo electrocardiographic abnormalities in persons living with HIV.

Persons living with HIV (PLWH) may have increased incidence of cardiovascular events and longer QTc intervals than uninfected persons. We aimed to investigate the incidence and risk factors of de novo major electrocardiogram (ECG) abnormalities and QTc prolongation in well-treated PLWH. We included virologically suppressed PLWH without major ECG abnormalities, who attended the 2-year follow-up in the Copenhagen comorbidity in HIV infection (COCOMO) study. ECGs were categorized according to Minnesota Code Manual. We defined de novo major ECG abnormalities as new major Minnesota Code Manual abnormalities. Prolonged QTc was defined as QTc > 460 ms in females and QTc > 450 ms in males. Of 667 PLWH without major ECG abnormalities at baseline, 34 (5%) developed de novo major ECG abnormalities after a median of 2.3 years. After adjustment, age (RR: 1.57 [1.08-2.28] per decade older), being underweight (RR: 5.79 [1.70-19.71]), current smoking (RR: 2.34 [1.06-5.16]), diabetes (RR: 3.89 [1.72-8.80]) and protease inhibitor use (RR: 2.45 [1.27-4.74) were associated with higher risk of getting de novo major ECG abnormalities. Of PLWH without prolonged QTc at baseline, only 11 (1.6%) participants developed de novo prolonged QTc. Five percent of well-treated PLWH acquired de novo major ECG abnormalities and protease inhibitor use was associated with more than twice the risk of de novo major ECG abnormalities. De novo prolonged QTc was rare and did not seem to constitute a problem in well-treated PLWH.

Discovery of Potent Antiallergic Agents Based on an o-Aminopyridinyl Alkynyl Scaffold.

Effective therapeutic agents are highly desired for immune-mediated allergic diseases. Herein, we report the design, synthesis, and structure-activity relationship of an o-aminopyridinyl alkyne series as novel orally bioavailable antiallergic agents, which was identified through phenotypic screening. Compound optimization yielded a highly potent compound 36, which effectively suppressed mast cell degranulation in a dose-dependent manner (IC50, 2.54 nM for RBL-2H3 cells; 48.28 nM for peritoneal mast cells (PMCs)) with a good therapeutic index. It also regulated the activation of FcεRI-mediated downstream signaling proteins in IgE/Ag-stimulated RBL-2H3 cells. In addition, 36 exhibited excellent in vivo pharmacokinetic properties and antiallergic efficacy in both passive systemic anaphylaxis (PSA) and house dust mite (HDM)-induced murine models of pulmonary allergic inflammation. Furthermore, preliminary analysis of the kinases profile identified Src-family kinases as potential targets for 36. Compound 36 may serve as a new valuable lead compound for future antiallergic drug discovery.

Point-of-Care Detection of Nonadherence to Antiretroviral Treatment for HIV-1 in Resource-Limited Settings Using Drug Level Testing for Efavirenz, Lopinavir, and Dolutegravir: A Validation and Pharmacokinetic Simulation Study.

BACKGROUND: Virological failure during antiretroviral treatment (ART) may indicate the presence of drug resistance, but may also originate from nonadherence. Qualitative detection of ART components using drug level testing may be used to differentiate between these scenarios. We aimed to validate and implement qualitative point-of-care drug level tests for efavirenz (EFV), lopinavir (LPV), and dolutegravir (DTG) in rural South Africa. METHODS: Qualitative performance of immunoassays for EFV, LPV, and DTG was assessed by calculating limit of detection (LoD), region of uncertainty, and qualitative agreement with a reference test. Minimum duration of nonadherence resulting in a negative drug level test was assessed by simulation of treatment cessation using validated population pharmacokinetic models. RESULTS: LoD was 0.05 mg/L for EFV, 0.06 mg/L for LPV, and 0.02 mg/L for DTG. Region of uncertainty was 0.01-0.06 mg/L for EFV, 0.01-0.07 mg/L for LPV, and 0.01-0.02 mg/L for DTG. Qualitative agreement with reference testing at the LoD in patient samples was 95.2% (79/83) for EFV, 99.3% (140/141) for LPV, and 100% (118/118) for DTG. After simulated treatment cessation, median time to undetectability below LoD was 7 days [interquartile range (IQR) 4-13] for EFV, 30 hours (IQR 24-36) for LPV, and 6 days (IQR 4-7) for DTG. CONCLUSIONS: We demonstrate that qualitative ART drug level testing using immunoassays is feasible in a rural resource-limited setting. Implementation of this technology enables reliable detection of recent nonadherence and may allow for rapid and cost-effective differentiation between patients in need for adherence counseling and patients who require drug resistance testing or alternative treatment.

Design and synthesis of Grp94 selective inhibitors based on Phe199 induced fit mechanism and their anti-inflammatory effects.

Glucose-regulated protein 94 (Grp94), a member of the Heat shock protein 90 (Hsp90) family, is implicated in many human diseases, including cancer, neurodegeneration, inflammatory, and infectious diseases. Here, we describe our effort to design and develop a new series of Grp94 inhibitors based on Phe199 induced fit mechanism. Using an alkynyl-containing inhibitor as a starting point, we developed compound 4, which showed potent inhibitory activity toward Grp94 in a fluorescence polarization-based assay. With improved physicochemical properties and suitable pharmacokinetic properties, compound 4 was advanced into in vivo bioactivity evaluation. In a dextran sulfate sodium (DSS)-induced mouse model of ulcerative colitis (UC), compound 4 showed anti-inflammatory property and reduced the levels of pro-inflammatory cytokines (TNF-α and IL-6). Together, these findings provide evidence that this approach may be promising for further Grp94 drug development efforts.

Efavirenz as a potential drug for the treatment of triple-negative breast cancers.

PURPOSE: In contrast to hormone receptor driven breast cancer, patients presenting with triple-negative breast cancer (TNBC) often have limited drug treatment options. Efavirenz, a non-nucleoside reverse transcriptase (RT) inhibitor targets abnormally overexpressed long interspersed nuclear element 1 (LINE-1) RT and has been shown to be a promising anticancer agent for treating prostate and pancreatic cancers. However, its effectiveness in treating patients with TNBC has not been comprehensively examined. METHODS: In this study, the effect of Efavirenz on several TNBC cell lines was investigated by examining several cellular characteristics including viability, cell division and death, changes in cell morphology as well as the expression of LINE-1. RESULTS: The results show that in a range of TNBC cell lines, Efavirenz causes cell death, retards cell proliferation and changes cell morphology to an epithelial-like phenotype. In addition, it is the first time that a whole-genome RNA sequence analysis has identified the fatty acid metabolism pathway as a key regulator in this Efavirenz-induced anticancer process. CONCLUSION: In summary, we propose Efavirenz is a potential anti-TNBC drug and that its mode of action can be linked to the fatty acid metabolism pathway.

The Early Results of Vertebral Pathological Compression Fracture of Extra- nodal Lymphoma with HIV-positive Patients Treated by Percutaneous Kyphoplasty.

BACKGROUND: Vertebral pathological compression fracture involving extra-nodal lymphoma impacts negatively on the quality of life of HIV-positive patients. The choice of a safe and effective approach to palliative care in this condition remains a challenge. OBJECTIVE: The purpose of this study was to investigate the safety and efficacy of percutaneous kyphoplasty (PKP) in the treatment of vertebral pathological compression fracture of extra-nodal lymphoma in HIV-positive patients. METHODS: A retrospective analysis, from January 2016 to August 2019, was performed on 7 HIVpositive patients, 3 males and 4 females, with extra-nodal lymphoma with a vertebral pathological compression fracture. The patients were treated using percutaneous kyphoplasty in our hospital. Preoperative assessment of the patients was conducted regarding their hematological profile, biochemical indicators, liver and kidney function, blood coagulation function, CD4+T lymphocyte count and viral load. Subsequently, the patients were placed on highly active antiretroviral therapy (HAART) and rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone (RCHOP) regimen. Besides, antibiotics, nutritional support and immune-modulating drugs were also administered, rationally. Postoperatively, the height of the anterior edge of the injured vertebrae, Visual Analogue Scale (VAS) and Oswestry Disability Index (ODI) values were evaluated. Patients were also monitored for any complications related to the operation. RESULTS: The average CD4+T cell count for the patients was 164 (range 114 ~247 / ul), while the viral load was 26,269 (range 5,765 ~82,321 copies/ul). All patients received nutritional and immune support and registered significant improvements in the levels of ALB and Hb (P<0.05). In all cases, the operation was uneventful with neither cement leakage nor toxic reactions observed. Similarly, no opportunistic infections, other complications or deaths were reported. The height of the anterior vertebral body and the ODI score of the injured vertebrae were significantly improved immediately after surgery (P<0.05). Compared to the preoperative VAS (7.71±1.11), postoperative values were significantly reduced immediately after surgery (3.85±0.90) and at 2 weeks, 1 month and 6 months post-surgery: 2.71±0.76, 3.29±1.11, and 4.00±0.82, respectively (P<0.01). CONCLUSION: Supported with appropriate perioperative treatment measures, PKP is safe and effective in the treatment of pathological vertebral compression fracture due to extra-nodal lymphoma in HIV-positive patients.

Stable and Well-Organized Near-Infrared Platinum(II)-Acetylide-Based Metallacycles-Mediated Cancer Phototherapy.

The development of metallacycles with high stability and intense near-infrared (NIR) absorption is important for biomedical applications. However, very few molecular design strategies have been developed on such metallacycles. Herein, we report a new series of stable and well-defined NIR-absorbing metallacycles (M1-M3) through the Pt-acetylide coordination with highly efficient photoconversion performance for cancer phototherapy. The metallacycles showed high stability and strong NIR absorption, and the absorption peaks were red shifted approximately 30 nm in comparison with their corresponding precursors. The introduction of Pt into metallacycles promotes significant photoconversions, including the singlet-to-triplet and nonradiative transitions. Moreover, the fabricated M3 nanoparticles (M3-NPs) showed favorable photoconversions into both thermal effect and singlet oxygen generation upon NIR irradiation, achieving tumor ablation. This novel design of Pt-acetylide metallacycles possesses not only complex topological architectures but also a valuable paradigm for precise cancer phototherapy, which is important for grafting stimuli-responsive functional groups into metallacycles for the development of high-performance biomedical supramolecular materials.

Dual disruption of aldehyde dehydrogenases 1 and 3 promotes functional changes in the glutathione redox system and enhances chemosensitivity in nonsmall cell lung cancer.

Aldehyde dehydrogenases (ALDHs) are multifunctional enzymes that oxidize diverse endogenous and exogenous aldehydes. We conducted a meta-analysis based on The Cancer Genome Atlas and Gene Expression Omnibus data and detected genetic alterations in ALDH1A1, ALDH1A3, or ALDH3A1, 86% of which were gene amplification or mRNA upregulation, in 31% of nonsmall cell lung cancers (NSCLCs). The expression of these isoenzymes impacted chemoresistance and shortened survival times in patients. We hypothesized that these enzymes provide an oxidative advantage for the persistence of NSCLC. To test this hypothesis, we used genetic and pharmacological approaches with DIMATE, an irreversible inhibitor of ALDH1/3. DIMATE showed cytotoxicity in 73% of NSCLC cell lines tested and demonstrated antitumor activity in orthotopic xenografts via hydroxynonenal-protein adduct accumulation, GSTO1-mediated depletion of glutathione and increased H2O2. Consistent with this result, ALDH1/3 disruption synergized with ROS-inducing agents or glutathione synthesis inhibitors to trigger cell death. In lung cancer xenografts with high to moderate cisplatin resistance, combination treatment with DIMATE promoted strong synergistic responses with tumor regression. These results indicate that NSCLCs with increased expression of ALDH1A1, ALDH1A3, or ALDH3A1 may be targeted by strategies involving inhibitors of these isoenzymes as monotherapy or in combination with chemotherapy to overcome patient-specific drug resistance.

Novel Class of Colony-Stimulating Factor 1 Receptor Kinase Inhibitors Based on an o-Aminopyridyl Alkynyl Scaffold as Potential Treatment for Inflammatory Disorders.

Colony-stimulating factor 1 receptor (CSF-1R) is involved in inflammatory disorders as well as in many types of cancer. Based on high-throughput screening and docking results, we performed a detailed structure-activity-relationship study, leading to the discovery of a new series of compounds with nanomolar IC50 values against CSF-1R without the inhibition of fibroblast growth factor receptors. One of the most promising hits, compound 29, potently inhibited CSF-1R kinase with an IC50 value of 0.7 nM, while it showed no inhibition to the same family member FMS-like tyrosine kinase 3. Compound 29 displayed excellent anti-inflammatory effects against RAW264.7 macrophages indicated by significant inhibition against the activation of the CSF-1R pathway with low cytotoxicity. In addition, compound 29 exhibited strong in vivo anti-inflammatory efficacy alongside favorable drug characteristics. This novel compound 29 may serve as a new drug candidate with promising applications in inflammatory disorders.

Propargylglycine decreases neuro-immune interaction inducing pain response in temporomandibular joint inflammation model.

The mechanisms underlying temporomandibular disorders following orofacial pain remain unclear. Hydrogen sulfide (H2S), a newly identified gasotransmitter, has been reported to modulate inflammation. Cystathionine γ-lyase (CSE) is responsible for the systemical production of H2S, which exerts both pro- and antinociceptive effects through inflammation. In the current study, we investigated whether the endogenous H2S production pathway contributes to arousal and maintenance of orofacial inflammatory pain, through the investigation of the effects of a CSE inhibitor, propargyglycine (PAG), in a rat CFA (Complete Freund Adjuvant)-induced temporomandibular inflammation model to mimic persistent pain in the orofacial region. For this, rats received either CFA or saline in the temporomandibular joints (TMJs), and after 3 or 14 days, they received a single injection of PAG or saline and were evaluated for nociception with the von Frey and formalin test. Also, pro-inflammatory cytokines, tumor necrosis factor-α (TNF-α), and interleukin-1ß (IL-1ß) were analyzed in TMJs and trigeminal ganglion (TG). In this last one, glial cells reactivity was also verified. Endogenous H2S production rate were measured in both, TMJ and TG. Our results indicated decreased allodynia and hyperalgesic responses in rats submitted to CFA after injection of PAG. Moreover, PAG inhibited leucocyte migration to temporomandibular synovial fluid after 3 and 14 days of inflammation. PAG was able to reduce levels of CBS, CSE, TNF-α, and IL-1ß in the TMJ and TG, after 13 days of CFA injection. The observed increased activation of glial cells in the trigeminal ganglia on the 14th day of inflammation can be prevented by the highest dose of PAG. Finally, CBS and CSE expression, and endogenous H2S production rate in the TMJ and TG was found higher in rats with persistent temporomandibular inflammation compared to rats injected with saline and PAG was able to prevent this elevation. Our results elucidated the molecular mechanisms by which H2S exerts its pro-inflammatory and pro-nociceptive role in the orofacial region by alterations in both local tissue and TG.

The therapeutic potential of cystathionine gamma-lyase in temporomandibular inflammation-induced orofacial hypernociception.

Hydrogen sulfide (H2S) is an endogenous neuromodulator produced mainly by the enzyme cystathionine gamma-lyase (CSE) in peripheral tissues. A pronociceptive role of endogenously produced H2S has been previously reported by our group in a model of orofacial inflammatory pain. Using the established persistent orofacial pain rat model induced by complete Freund's adjuvant (CFA) injection into temporomandibular joint (TMJ), we have now investigated the putative role of endogenous H2S modulating hypernociceptive responses. Additionally, plasmatic extravasation on TMJ was measured following different treatments by Evans blue dye quantification. Thus, rats were submitted to Von Frey and Formalin tests in orofacial region before and after pharmacological inhibition of the CSE-H2S system combined or not with CFA-induced TMJ inflammation. Pretreatment with CSE inhibitor, propargylglycine (PAG; 88.4 µmol/kg) reduced temporomandibular inflammatory pain when injected locally as well as systemically. In particular, local PAG injection seems to be more effective for hypernociceptive responses in orofacial persistent inflammation since its action is evidenced in the majority analyzed periods of the inflammatory process compared to its systemic use. Moreover, local injection seems to act on temporomandibular vascular permeability, evidenced by decreased plasmatic extravasation induced by local PAG administration. Our data are consistent with the notion that the endogenous synthetized gas H2S modulates persistent orofacial pain responses revealing the pharmacological importance of the CSE inhibitor as a possible therapeutic target for their control.

AZD5582, an IAP antagonist that leads to apoptosis in head and neck squamous cell carcinoma cell lines and is eligible for combination with irradiation.

CONCLUSION: On the one hand, AZD5582, an inhibitor of inhibitor of apoptosis family proteins (IAP), leads to cellular growth arrest and induction of apoptosis in head and neck squamous cell carcinoma (HNSCC) cell lines. On the other hand, it is a viable candidate for combination therapy with irradiation. OBJECTIVES: The aim and purpose of this study was to evaluate the effects of AZD5582 on HNSCC cell lines. METHODS: HNSCC cell lines SCC25, Cal27, and FaDu were used for all cell culture experiments. Proliferation assays were used to assess a potential inhibitory effect of AZD5582 and a combination therapy of the IAP inhibitor and irradiation. Colony forming assays were used to determine long-term effects of a combined treatment. Apoptosis was measured via flow cytometry and wound-healing assays were performed. RESULTS: All three cell lines showed a dose-dependent cytotoxic effect after treatment with AZD5582. It was possible to observe a synergistic and additive effect after short-term treatment of AZD5582 and irradiation in Ca27 and FaDu cells, respectively. All test cell lines showed a significant inhibition of colony formation after combined treatment. Treatment of AZD5582 resulted in apoptosis in SCC25, Cal27, and FaDu cells.

The apoptotic and autophagic properties of two natural occurring prodrugs, hyperoside and hypoxoside, against pancreatic cancer cell lines.

Pancreatic cancer is only the 12th most common cancer, but the fourth leading cause of cancer-related deaths in the world. This is due to late prognosis, poor response to chemotherapy and early metastases. Natural prodrugs may play an important role in the treatment of pancreatic cancer. The main aim of this study was to determine the cytotoxicity of five natural prodrugs, namely harpagoside, hyperoside, hypoxoside, oleuropein and polydatin, by investigating apoptosis and autophagy as possible mechanism/s of action. Hypoxoside and hyperoside have shown selective cytotoxicity at IC50 values of ∼25 and 50µM against INS-1 and MIA PaCa-2 pancreatic cancer cells, respectively. Hypoxoside and hyperoside induced G2/M phase arrest and caspase-3 activation in INS-1 and MIA PaCa-2 cells, respectively. Hoechst/phalloidin staining confirmed morphological changes, including condensed chromatin multinucleation, membrane blebbing and loss of cytoskeletal arrangement in INS-1 and MIA PaCa-2 cells. Acridine orange staining was absent in INS-1 (hypoxoside) and MIA PaCa-2 (hyperoside) treated cells, whereas LC3B expression was not significantly increased. INS-1 and MIA PaCa-2 treated cells favour the cell death pathway, apoptosis, over the cell survival pathway, autophagy.

Effects of 2-Octynyladenosine (YT-146) on Mitochondrial Function in Ischemic/Reperfused Rat Hearts.

This study investigated the effects of an adenosine receptor agonist, 2-octynyladenosine (YT-146), on mitochondrial function in ischemic and ischemic/reperfused hearts. Isolated rat hearts were perfused in the Langendorff manner with a constant flow rate, and exposed to 30 min of ischemia followed by 60 min of reperfusion. Preischemic treatment with YT-146 significantly improved postischemic recovery of left ventricular developed pressure. The high-energy phosphate content in reperfused hearts treated with YT-146 was also more greatly restored than in untreated hearts. YT-146 treatment attenuated the Na(+) content of a mitochondria-enriched fraction, but not the myocardial Na(+) content, at the end of ischemia. These results suggest that preischemic YT-146 treatment preserves the energy-producing ability of mitochondria during ischemia in the Na(+)-accumulated myocardium. YT-146 also attenuated both the sodium lactate-induced decrease in mitochondrial energy-producing ability and the increase in mitochondrial Na(+) concentration in the myocardial skinned fibers. YT-146 may attenuate Na(+) influx to myocardial mitochondria in ischemic cardiac cells, resulting in both preservation of the ability of mitochondria to produce energy and enhancement of the contractile recovery in reperfused hearts. Our findings suggest that the cardioprotective effects of YT-146 against ischemia/reperfusion injury are at least partially due to the preservation of mitochondrial function in the ischemic myocardium.

A novel small-molecule IAP antagonist, AZD5582, draws Mcl-1 down-regulation for induction of apoptosis through targeting of cIAP1 and XIAP in human pancreatic cancer.

Inhibitor of apoptosis proteins (IAPs) plays an important role in controlling cancer cell survival. IAPs have therefore attracted considerable attention as potential targets in anticancer therapy. In this study, we investigated the anti-tumor effect of AZD5582, a novel small-molecule IAP inhibitor, in human pancreatic cancer cells. Treating human pancreatic cancer cells with AZD5582 differentially induced apoptosis, dependent on the expression of p-Akt and p-XIAP. Moreover, the knockdown of endogenous Akt or XIAP via RNA interference in pancreatic cancer cells, which are resistant to AZD5582, resulted in increased sensitivity to AZD5582, whereas ectopically expressing Akt or XIAP led to resistance to AZD5582. Additionally, AZD5582 targeted cIAP1 to induce TNF-α-induced apoptosis. More importantly, AZD5582 induced a decrease of Mcl-1 protein, a member of the Bcl-2 family, but not that of Bcl-2 and Bcl-xL. Interestingly, ectopically expressing XIAP and cIAP1 inhibited the AZD5582-induced decrease of Mcl-1 protein, which suggests that AZD5582 elicits Mcl-1 decrease for apoptosis induction by targeting of XIAP and cIAP1. Taken together, these results indicate that sensitivity to AZD5582 is determined by p-Akt-inducible XIAP phosphorylation and by targeting cIAP1. Furthermore, Mcl-1 in pancreatic cancer may act as a potent marker to analyze the therapeutic effects of AZD5582.

Hydrogen sulfide in posthemorrhagic shock mesenteric lymph drainage alleviates kidney injury in rats.

Posthemorrhagic shock mesenteric lymph (PHSML) is a key factor in multiple organ injury following hemorrhagic shock. We investigated the role of hydrogen sulfide (H2S) in PHSML drainage in alleviating acute kidney injury (AKI) by administering D,L-propargylglycine (PPG) and sodium hydrosulfide hydrate (NaHS) to 12 specific pathogen-free male Wistar rats with PHSML drainage. A hemorrhagic shock model was established in 4 experimental groups: shock, shock+drainage, shock+drainage+PPG (45 mg/kg, 0.5 h prehemorrhage), and shock+drainage+NaHS (28 µmol/kg, 0.5 h prehemorrhage). Fluid resuscitation was performed after 1 h of hypotension, and PHMSL was drained in the last three groups for 3 h after resuscitation. Renal function and histomorphology were assessed along with levels of H2S, cystathionine-γ-lyase (CSE), Toll-like receptor 4 (TLR4), interleukin (IL)-10, IL-12, and tumor necrosis factor (TNF)-α in renal tissue. Hemorrhagic shock induced AKI with increased urea and creatinine levels in plasma and higher H2S, CSE, TLR4, IL-10, IL-12, and TNF-α levels in renal tissue. PHSML drainage significantly reduced urea, creatinine, H2S, CSE, and TNF-α but not TLR4, IL-10, or IL-12. PPG decreased creatinine, H2S, IL-10, and TNF-α levels, but this effect was reversed by NaHS administration. In conclusion, PHSML drainage alleviated AKI following hemorrhagic shock by preventing increases in H2S and H2S-mediated inflammation.

Inecalcitol, an analog of 1,25D3, displays enhanced antitumor activity through the induction of apoptosis in a squamous cell carcinoma model system.

Epidemiological data suggest an important role of vitamin D signaling in cancer development and progression, and experimental studies demonstrate that the active vitamin D metabolite 1α, 25-dihydroxyvitamin D3 (1,25D3) has broad spectrum antitumor activity. Hypercalcemia has often been suggested to limit the clinical application of these data. The 14-epi-analog of 1,25D3, inecalcitol [19-nor-14-epi-23-yne-1,25-(OH)2D3; TX522], was developed to have superagonistic antitumor activities but low hypercalcemia potential. We examined the antitumor activity of inecalcitol and the underlying mechanisms in a murine squamous cell carcinoma (SCC) model system. In vitro, compared with 1,25D3, inecalcitol showed enhanced vitamin D receptor (VDR)-mediated transcriptional activity. Inecalcitol suppressed SCC cell proliferation in a dose-dependent manner with an IC50 value 30 times lower than that of 1,25D3. Both inecalcitol and 1,25D3 induced a comparable level of G0/G1 cell cycle arrest in SCC cells. The level of apoptosis induced by inecalcitol was markedly higher than that of 1,25D3. Apoptosis was mediated through the activation of the caspase 8/10- caspase 3 pathway. Further, inecalcitol markedly inhibited the mRNA and protein expression of c-IAP1 and XIAP compared with 1,25D3. In vivo, inecalcitol inhibits SCC tumor growth in a dose-dependent fashion. Notably, inecalcitol induced a significantly higher level of apoptosis in the SCC xenograft model. While in vitro inecalcitol demonstrates apparent enhanced VDR binding and antiproliferative effects compared to 1,25D3, in vivo these advantages disappear; at doses of inecalcitol that have equivalent antitumor effects, similar hypercalcemia is seen. This may be explained by the pharmacokinetics of 1,25D3 vs. inecalcitol and attributed to the much shorter serum half-life of inecalcitol.We show that inecalcitol has potent antitumor activity in the SCC model system, and this is associated with a strong induction of apoptosis. These findings support the further development of inecalcitol in cancer treatment.

Inecalcitol, an analog of 1α,25(OH)(2) D(3) , induces growth arrest of androgen-dependent prostate cancer cells.

19-nor-14-epi-23-yne-1,25(OH)(2) D(3) (inecalcitol) is a unique vitamin D(3) analog. We evaluated the activity of inecalcitol in a human prostate cancer model system. The analog was 11-fold more potent than 1,25(OH)(2) D(3) in causing 50% clonal growth inhibition of androgen-sensitive human prostate cancer LNCaP cells. Inecalcitol, more than 1,25(OH)(2) D(3) , reduced in a dose-dependent manner the expression levels of the transcription factor ETS variant 1 and the serine/threonine protein kinase Pim-1, both of which are upregulated in prostate cancer. Remarkably, dose challenge experiments revealed that inecalcitol maximal tolerated dose (MTD) by intraperitoneal (i.p.) administration was 30 µg/mouse (1,300 µg/kg) three times per week, while we previously found that the MTD of 1,25(OH)(2) D(3) is 0.0625 µg/mouse; therefore, inecalcitol is 480 times less hypercalcemic than 1,25(OH)(2) D(3) . Pharmacokinetic studies showed that plasma half-life of inecalcitol were 18.3 min in mice. A xenograft model of LNCaP cells was developed in immunodeficient mice treated with inecalcitol. The tumors of the diluent-treated control mice increased in size but those in the inecalcitol treatment group did not grow. Our data suggest that inecalcitol inhibits androgen-responsive prostate cancer growth in vivo and should be examined either alone or with other chemotherapy in clinical trials in individuals with rising serum prostate-specific antigen after receiving either surgery or irradiation therapy with curative intent.