Catechol-O-Methyltransferase inhibition and alcohol use disorder: Evaluating the efficacy of tolcapone in ethanol-dependent rats.

Alcohol Use Disorder (AUD) is a significant public health issue in the United States. It affects millions of individuals and their families and contributes to substantial societal and economic burdens. Despite the availability of some pharmacological treatments, there is still a pressing need to develop more effective therapeutic strategies to address the diverse range of symptoms and challenges associated with AUD. Catechol-O-methyltransferase (COMT) inhibition recently emerged as a promising new approach to treating AUD due to its potential to improve cognitive effects commonly associated with AUD. Tolcapone, an FDA-approved COMT inhibitor, has shown some promise for treating AUD; however, its ability to decrease drinking in ethanol-dependent rats has not been well-established. In this study, we evaluated the effects of tolcapone on operant, oral ethanol self-administration in non-dependent and dependent rats, and in rats that self-administered oral saccharin. To induce dependence, rats underwent the chronic intermittent exposure to vapor model, and their drinking levels were assessed during acute withdrawal from ethanol. Our results demonstrated that tolcapone attenuated responding for ethanol in dependent rats only, without affecting self-administration in non-dependent rats or rats self-administering saccharin. Moreover, we found that tolcapone was differentially effective in different estrous phases in female rats. These findings suggest that COMT inhibition, specifically using tolcapone, may be a valuable pharmacotherapy for treating AUD, particularly in individuals who are physically dependent on alcohol. Further research is needed to elucidate the precise mechanisms underlying the observed effects and to assess the potential of COMT inhibitors in a broader population of individuals with AUD.

Efonidipine Exerts Cerebroprotective Effect by Down-regulation of TGF-ß/SMAD-2-Dependent Signaling Pathway in Diabetic Rats.

Calcium overload and hyperglycemia are risks of stroke onset in diabetics. Our study was designed to elucidate the beneficial role of calcium channel blockers by targeting voltage-gated calcium channels in diabetes-associated cerebrovascular complications. Diabetes was induced using the neonatal streptozotocin rat model. After confirmation of diabetes, middle cerebral artery occlusion (MCAO) was carried out. The pre-treatment with 1 mg/kg/day efonidipine was administered for the period of 4 weeks. After 24 h of ischemic induction surgery, the neurological score was determined, and blood was collected for determination of biochemical parameters. Treatment with efonidipine showed a significant reduction in post-ischemic brain infract volume, brain hemisphere weight difference, neurological score, Na+-K+ ATPase activity, serum CK-MB, and LDH levels in normoglycemic and hyperglycemic MCAO-induced animals. While no significant changes in glucose and lipid levels were observed by treatment, efonidipine significantly decreased the levels of malondialdehyde, acetylcholine esterase, and nitrite levels and increased the levels of antioxidant markers in both normoglycemic and hyperglycemic MCAO animals. TGF-ß and VEGF were found to be down-regulated after treatment with efonidipine in gene expression study. In conclusion, the study data supports the cerebroprotective role of efonidipine in diabetic animals possibly through TGF-ß/SMAD-2 signaling pathway.

Co-Operativity between MYC and BCL-2 Pro-Survival Proteins in Cancer.

B-Cell Lymphoma 2 (BCL-2), c-MYC and related proteins are arguably amongst the most widely studied in all of biology. Every year there are thousands of papers reporting on different aspects of their biochemistry, cellular and physiological mechanisms and functions. This plethora of literature can be attributed to both proteins playing essential roles in the normal functioning of a cell, and by extension a whole organism, but also due to their central role in disease, most notably, cancer. Many cancers arise due to genetic lesions resulting in deregulation of both proteins, and indeed the development and survival of tumours is often dependent on co-operativity between these protein families. In this review we will discuss the individual roles of both proteins in cancer, describe cancers where co-operativity between them has been well-characterised and finally, some strategies to target these proteins therapeutically.

The apoptosis inhibitor Bcl-xL controls breast cancer cell migration through mitochondria-dependent reactive oxygen species production.

The Bcl-xL apoptosis inhibitor plays a major role in vertebrate development. In addition to its effect on apoptosis, Bcl-xL is also involved in cell migration and mitochondrial metabolism. These effects may favour the onset and dissemination of metastasis. However, the underlying molecular mechanisms remain to be fully understood. Here we focus on the control of cell migration by Bcl-xL in the context of breast cancer cells. We show that Bcl-xL silencing led to migration defects in Hs578T and MDA-MB231 cells. These defects were rescued by re-expressing mitochondria-addressed, but not endoplasmic reticulum-addressed, Bcl-xL. The use of BH3 mimetics, such as ABT-737 and WEHI-539 confirmed that the effect of Bcl-xL on migration did not depend on interactions with BH3-containing death accelerators such as Bax or BH3-only proteins. In contrast, the use of a BH4 peptide that disrupts the Bcl-xL/VDAC1 complex supports that Bcl-xL by acting on VDAC1 permeability contributes to cell migration through the promotion of reactive oxygen species production by the electron transport chain. Collectively our data highlight the key role of Bcl-xL at the interface between cell metabolism, cell death, and cell migration, thus exposing the VDAC1/Bcl-xL interaction as a promising target for anti-tumour therapy in the context of metastatic breast cancer.

Targeting BCL2 in Chronic Lymphocytic Leukemia and Other Hematologic Malignancies.

Apoptosis, the process of programmed cell death, occurs normally during development and aging. Members of the B-cell lymphoma 2 (BCL2) family of proteins are central regulators of apoptosis, and resistance to apoptosis is one of the hallmarks of cancer. Targeting the apoptotic pathway via BCL2 inhibitors has been considered a promising treatment strategy in the past decade. Initial efforts with small molecule BH3 mimetics such as ABT-737 and ABT-263 (navitoclax) pioneered the development of the first-in-class Food and Drug Administration (FDA)-approved oral BCL2 inhibitor, venetoclax. Venetoclax was approved for the treatment of chronic lymphocytic leukemia and acute myeloid leukemia, and is now being studied in a number of hematologic malignancies. Several other inhibitors targeting different BCL2 family members are now in early stages of development.

Nutritional preconditioning induced by astragaloside â £ on isolated hearts and cardiomyocytes against myocardial ischemia injury via improving Bcl-2-mediated mitochondrial function.

Ischemic preconditioning and pharmacological preconditioning are common strategies to prevent lethal myocardial injury, especially nutritional preconditioning (NPC). In this study, we investigated the effects of astragaloside IV (Ast), as an NPC agent, on myocardium suffered anoxia/reoxygenation (A/R) injury. Rats received 5 mg/kg Ast daily for 3 weeks by intragastric administration. Then, hearts were harvested and underwent A/R treatment using a Langendorff apparatus. Ast- pretreatment significantly promoted functional recovery of the myocardium, reduced infarct size, and oxidative stress, and decreased the apoptotic index. Similar findings were demonstrated in H9c2 cardiomyocytes that were pretreated with Ast for 24 h. Moreover, Ast-pretreatment significantly upregulated Bcl-2 expression, especially in mitochondria. The effects of Ast treatment against A/R injury were also reflected by increased antioxidant potential, inhibited reactive oxygen species (ROS) burst, increased oxygen consumption rate, maintained mitochondrial membrane potential (MMP), inhibited mitochondrial permeability transition pore (mPTP) opening, and prevented apoptosis. Selective inhibition of Bcl-2 by ABT-737 decreased myocardial injury protection of Ast. Ast-pretreatment resulted in NPC- related effects against A/R, and mitochondria may be the target of a cascade of events elicited by upregulating Bcl-2 expression, promoting translocation of Bcl-2 into mitochondria, maintaining MMP, inhibiting ROS bursts, thereby leading to recovery of mitochondrial respiration, preventing mPTP opening, decreasing cytochrome C release, preventing apoptosis, and ultimately alleviating myocardial injury.

Synergistic antitumor effects of 9.2.27-PE38KDEL and ABT-737 in primary and metastatic brain tumors.

Standard treatment, unfortunately, yields a poor prognosis for patients with primary or metastatic cancers in the central nervous system, indicating a necessity for novel therapeutic agents. Immunotoxins (ITs) are a class of promising therapeutic candidates produced by fusing antibody fragments with toxin moieties. In this study, we investigated if inherent resistance to IT cytotoxicity can be overcome by rational combination with pro-apoptotic enhancers. Therefore, we combined ITs (9.2.27-PE38KDEL or Mel-14-PE38KDEL) targeting chondroitin sulfate proteoglycan 4 (CSPG4) with a panel of Bcl-2 family inhibitors (ABT-737, ABT-263, ABT-199 [Venetoclax], A-1155463, and S63845) against patient-derived glioblastoma, melanoma, and breast cancer cells/cell lines. In vitro cytotoxicity assays demonstrated that the addition of the ABT compounds, specifically ABT-737, sensitized the different tumors to IT treatment, and improved the IC50 values of 9.2.27-PE38KDEL up to >1,000-fold. Mechanistic studies using 9.2.27-PE38KDEL and ABT-737 revealed that increased levels of intracellular IT, processed (active) exotoxin, and PARP cleavage correlated with the enhanced sensitivity to the combination treatment. Furthermore, we confirmed the synergistic effect of 9.2.27-PE38KDEL and ABT-737 combination therapy in orthotopic GBM xenograft and cerebral melanoma metastasis models in nude mice. Our study defines strategies for overcoming IT resistance and enhancing specific antitumor cytotoxicity in primary and metastatic brain tumors.

Impaired immune surveillance accelerates accumulation of senescent cells and aging.

Cellular senescence is a stress response that imposes stable cell-cycle arrest in damaged cells, preventing their propagation in tissues. However, senescent cells accumulate in tissues in advanced age, where they might promote tissue degeneration and malignant transformation. The extent of immune-system involvement in regulating age-related accumulation of senescent cells, and its consequences, are unknown. Here we show that Prf1-/- mice with impaired cell cytotoxicity exhibit both higher senescent-cell tissue burden and chronic inflammation. They suffer from multiple age-related disorders and lower survival. Strikingly, pharmacological elimination of senescent-cells by ABT-737 partially alleviates accelerated aging phenotype in these mice. In LMNA+/G609G progeroid mice, impaired cell cytotoxicity further promotes senescent-cell accumulation and shortens lifespan. ABT-737 administration during the second half of life of these progeroid mice abrogates senescence signature and increases median survival. Our findings shed new light on mechanisms governing senescent-cell presence in aging, and could motivate new strategies for regenerative medicine.

CAPE-pNO2 attenuates diabetic cardiomyopathy through the NOX4/NF-κB pathway in STZ-induced diabetic mice.

Diabetic cardiomyopathy (DCM) is one of the most severe cardiovascular complications in diabetes. Caffeic acid para-nitro phenethyl ester (CAPE-pNO2) could ameliorate diabetic nephropathy in the diabetic mice in our previous study. This paper was aimed to investigate the effect of CAPE-pNO2 on DCM and its potential mechanism. The DCM mice were established by intraperitoneal injection with streptozotocin (STZ, 50 mg/kg) for 5 days. When the fasting blood glucose level remains above 11.1 mmol/L, treated the mice with CAPE and CAPE-pNO2 for 8 weeks, then the mice were executed, and the samples of blood and heart tissue were collected for the subsequent experiments. The results showed that CAPE-pNO2 can alleviate CK, LDH, TC and TG levels, as well as depress the activity of ROS by down-regulating the expression of NOX4 and improving SOD activity in the serum of STZ-induced DCM mice. Meanwhile, it can also reduce the content of MDA and inhibit lipid accumulation. Besides, CAPE-pNO2 could repress the expression of TNF-α, IL-1ß and IL-6 IL - 6 via the NOX4/NF-κB pathway to improve the development of inflammation. Furthermore, it can suppress the expression of collagen and fibronectin to inhibit myocardial fibrosis through the TGF-ß1/Smad pathway, and inhibit ECM deposition by regulating TGF-ß1 directly, as shown in cardiac tissue section. Importantly, the above results showed that CAPE-pNO2 had better effects of reversing pathological changes than CAPE in a significant difference of p < 0.05. In brief, CAPE-pNO2 can prevent the heart injury of DCM mice via the NOX4/NF-κB pathway, and shows the improvement effects of anti-fibrosis, anti-oxidative and anti-inflammatory.

Secreted Heat Shock Protein 90α Attenuated the Effect of Anticancer Drugs in Small-Cell Lung Cancer Cells Through AKT/GSK3ß/ß-Catenin Signaling.

Small-cell lung cancer (SCLC) represents the progressive form of lung cancer. Patients with SCLC have poor prognosis, partially due to drug resistance. Therefore, understanding the underlying mechanism for drug resistance in SCLC is needed to improve clinical outcomes. The concentrations of heat shock protein 90α (HSP90α) in medium were detected by enzyme-linked immunosorbent assay. The protein levels were detected by Western blot. Cell apoptosis was detected by propidium iodide staining in cell lines or terminal deoxynucleotidyl transferase dUTP nick end labeling staining in tumor sections. Doxorubicin (DOX) was administered into cultured cell lines or intraperitoneally injected into xenograft mouse to induce apoptosis. In SCLC cell lines, either DOX or ABT-737 increased extracellular HSP90α levels, which attenuated the percentage of apoptotic cells. Extracellular HSP90α activated Ak strain transforming (AKT) and ß-catenin signaling and inhibited glycogen synthase kinase 3ß (GSK3ß) signaling. In the xenograft mouse model, extracellular HSP90α promoted tumor development and inhibited apoptosis of tumor cells. Heat shock protein 90α attenuates the efficacy of anticancer drugs in SCLC cells through AKT/GSK3ß/ß-catenin signaling.

ABT737 reverses cisplatin resistance by targeting glucose metabolism of human ovarian cancer cells.

The poor prognosis and high mortality of patients with ovarian cancer result in part from their poor response to platinum-based chemotherapy. However, the precise mechanism behind cisplatin resistance is still not fully understood. In the present study, the authors explored the mechanism of resistance to cisplatin from the perspective of glucose metabolism in human ovarian cancer. The experiments using genetically matched ovarian cancer cell lines SKOV3 (cisplatin-sensitive) and SKOV3/DDP (cisplatin-resistant) in the present study provided some important findings. First, in comparison to SKOV3 cells, SKOV3/DDP cells exhibited decreased dependence on aerobic glycolysis and an increased demand for glucose. Secondly, the stable overexpression of Bcl­2 and ability to shift metabolism towards oxidative phosphorylation (OXPHOS) in SKOV3/DDP cells were associated with increased oxygen consumption. Furthermore, the metabolic characteristic of elevated OXPHOS primarily comprised most mitochondrial­derived reactive oxygen species (ROS) and, at least in part, contributed to the slight pro-oxidant state of SKOV3/DDP cells in turn. Thirdly, SKOV3/DDP cells reset the redox balance by overexpressing the key enzyme glucose 6-phosphate dehydrogenase (G6PD) of the pentose phosphate pathway to eliminate the cytotoxicity of highly elevated ROS. Furthermore, the inhibition of Bcl­2 reduced the OXPHOS and sensitivity of SKOV3/DDP cells to cisplatin in a selective manner. Furthermore, when combined with 2-deoxyglucose (2-DG), the anticancer effect of the Bcl­2 inhibitor ABT737 was greatly potentiated and hypoxia-inducible factor 1α (HIF­1α) appeared to be closely associated with Bcl­2 family members in the regulation of glucose metabolism. These results suggested that the special glucose metabolism in SKOV3/DDP cells might be selectively targeted by disrupting Bcl­2-dependent OXPHOS.

Anti-allergic and anti-inflammatory effects of the Bcl-2 inhibitor ABT-737 on experimental allergic rhinitis models.

The anti-cancer agent ABT-737 is designed specifically to inhibit anti-apoptotic proteins of the Bcl-2 family. The development of cancer has long been associated with inflammation. Here, we assess the anti-allergic and anti-inflammatory effects and the underlying molecular mechanisms of ABT-737 on allergic rhinitis (AR) using in vitro and in vivo models. In the in vitro model, the ABT-737 treatment diminished the levels of several inflammatory cytokines in this case vascular endothelial growth factor (VEGF), thymic stromal lymphopoietin (TSLP), interleukin (IL)-1ß, IL-6, and tumor necrosis factor-α (TNF-α) by inhibiting caspase-1 and NF-κB activation in an activated human mast cell line, here HMC-1 cells. These mechanistic observations were validated in ovalbumin-sensitized AR mice. In an AR animal model, ABT-737 significantly diminished clinical symptoms of AR and the levels of AR biomarkers, specifically IgE, histamine, hypoxia-inducible factor-1α, VEGF, TSLP, IL-1ß, IL-4, IL-5, IL-6, IL-13, TNF-α, intercellular adhesion molecule-1, and macrophage inflammatory protein-2. In addition, ABT-737 reduced the degree of caspase-1 activity compared to that in AR mice. Simultaneously, ABT-737 diminished the recruitment of mast cells and eosinophils into nasal mucosa tissues compared to the levels in AR mice. In conclusion, we identified new anti-allergic and anti-inflammatory effects of ABT-737. These results imply that ABT-737 can ameliorate allergic inflammatory diseases such as AR.

Quantitative assessment of the sensitivity of dormant AML cells to the BAD mimetics ABT-199 and ABT-737.

Cells from patients with acute myeloid leukemia (AML) that remain dormant and protected by stromal cells may escape effects of chemotherapy. We modeled dormancy in vitro and investigated the ability of Bcl-2 inhibitors ABT-199 and ABT-737 to overcome chemoprotection of dormant cells. CD34-enriched primary AML cells with aberrant leukemia-associated phenotypes (LAPs) were cultured on stromal cells. The chemosensitivity of dormant (PKH26high), CD34+, LAP+ cells was ascertained by 5-colour flow cytometric counting after 12 d. The PKH26high, CD34+, LAP + subset retained clonogenic capacity. The dormant fraction was completely resistant to Ara-C (p = .007). However, ABT-199 and ABT-737 were able to reduce the dormant fraction by 84% and 80%, respectively, of their effects on proliferating counterparts. In conclusion, we have elaborated a system for quantifying chemosensitivity in LAP+ dormant leukemia cells, thought to contribute to disease relapse, and shown sensitivity of dormant LAP+ cells to ABT-199 and ABT-737 in this system.

Effect of amlodipine, efonidipine, and trichlormethiazide on home blood pressure and upper-normal microalbuminuria assessed by casual spot urine test in essential hypertensive patients.

The aim of this study was to assess the effects of irbesartan alone and combined with amlodipine, efonidipine, or trichlormethiazide on blood pressure (BP) and urinary albumin (UA) excretion in hypertensive patients with microalbuminuria (30≤UA/creatinine (Cr) ratio [UACR] <300 mg/g Cr) and upper-normal microalbuminuria (10≤UACR<30 mg/g Cr). This randomized controlled trial enrolled 175 newly diagnosed and untreated hypertensive patients (home systolic blood pressure [SBP]≥135 mmHg; 10≤UACR<300 mg/g Cr of casual spot urine at the first visit to clinic). All patients were treated with irbesartan (week 0). Patients who failed to achieve home SBP ≤125 mmHg on 8-week irbesartan monotherapy (nonresponders, n = 115) were randomized into three additional drug treatment groups: trichlormethiazide (n = 42), efonidipine (n = 39), or amlodipine (n = 34). Irbesartan monotherapy decreased home SBP and first morning urine samples (morning UACR) for 8 weeks (p < 0.0001). At 8 weeks after randomization, all three additional drugs decreased home SBP (p < 0.0002) and trichlormethiazide significantly decreased morning UACR (p = 0.03). Amlodipine decreased morning UACR in patients with microalbuminuria based on casual spot urine samples (p = 0.048). However, multivariate analysis showed that only higher home SBP and UACR at week 8, but not any additional treatments, were significantly associated with UACR reduction between week 8 and week 16. In conclusion, crucial points of the effects of combination therapy on UACR were basal UACR and SBP levels. The effect of trichlormethiazide or amlodipine treatment in combination with irbesartan treatment on microalbuminuria needs to be reexamined based on a larger sample size after considering basal UACR and SBP levels.

Current perspectives in fragment-based lead discovery (FBLD).

It is over 20 years since the first fragment-based discovery projects were disclosed. The methods are now mature for most 'conventional' targets in drug discovery such as enzymes (kinases and proteases) but there has also been growing success on more challenging targets, such as disruption of protein-protein interactions. The main application is to identify tractable chemical startpoints that non-covalently modulate the activity of a biological molecule. In this essay, we overview current practice in the methods and discuss how they have had an impact in lead discovery - generating a large number of fragment-derived compounds that are in clinical trials and two medicines treating patients. In addition, we discuss some of the more recent applications of the methods in chemical biology - providing chemical tools to investigate biological molecules, mechanisms and systems.

Highlights from the Biennial International Congress on Schizophrenia Research (ICOSR), March 24-March 28, 2017.

The 2017 International Congress on Schizophrenia Research, held in San Diego, California (March 24-28, 2017), attracted over 900 attendees from 34 countries. With the gracious assistance of Congress president James Meador-Woodruff, we bring you the following reports on the prospects for new drugs to treat schizophrenia.

Tolcapone induces oxidative stress leading to apoptosis and inhibition of tumor growth in Neuroblastoma.

Catechol-O-methyltransferase (COMT) is an enzyme that inactivates dopamine and other catecholamines by O-methylation. Tolcapone, a drug commonly used in the treatment of Parkinson's disease, is a potent inhibitor of COMT and previous studies indicate that Tolcapone increases the bioavailability of dopamine in cells. In this study, we demonstrate that Tolcapone kills neuroblastoma (NB) cells in preclinical models by inhibition of COMT. Treating four established NB cells lines (SMS-KCNR, SH-SY5Y, BE(2)-C, CHLA-90) and two primary NB cell lines with Tolcapone for 48 h decreased cell viability in a dose-dependent manner, with IncuCyte imaging and Western blotting indicating that cell death was due to caspase-3-mediated apoptosis. Tolcapone also increased ROS while simultaneously decreasing ATP-per-cell in NB cells. Additionally, COMT was inhibited by siRNA in NB cells and showed similar increases in apoptotic markers compared to Tolcapone. In vivo xenograft models displayed inhibition of tumor growth and a significant decrease in time-to-event in mice treated with Tolcapone compared to untreated mice. These results indicate that Tolcapone is cytotoxic to neuroblastoma cells and invite further studies into Tolcapone as a promising novel therapy for the treatment of neuroblastoma.

Effects of iron overload, an iron chelator and a T-Type calcium channel blocker on cardiac mitochondrial biogenesis and mitochondrial dynamics in thalassemic mice.

Although cardiac mitochondrial dysfunction is involved in the pathophysiology of iron-overload cardiomyopathy, the precise mechanisms of iron-induced mitochondrial dysfunction, and the roles of the iron chelator deferiprone and the T-type calcium channel blocker efonidipine on cardiac mitochondrial biogenesis in thalassemic mice are still unknown. ß-thalassemic (HT) mice were fed with a normal diet (ND) or a high iron-diet (FE) for 90 days. Then, the FE-fed mice were treated with deferiprone (75mg/kg/day) or efonidipine (4mg/kg/day) for 30 days. The hearts were used to determine cardiac mitochondrial function, biogenesis, mitochondrial dynamics and protein expressions for oxidative phosphorylation (OXPHOS) and apoptosis. ND-fed HT mice had impaired heart rate variability (HRV), increased mitochondrial dynamic proteins and caspase-3, compared with ND-fed wild-type mice. Iron overload led to increased plasma non-transferrin bound iron, oxidative stress, and the impairments of HRV and left ventricular function, cardiac mitochondrial function and mitochondrial dynamics, and decreased complex IV in thalassemic mice. Our results suggested that deferiprone and efonidipine treatment showed similar benefit in attenuating cardiac iron deposit and oxidative stress, and improved cardiac mitochondrial function, leading to improved left ventricular function, without altering the cardiac mitochondrial biogenesis, and apoptosis proteins in iron-overload thalassemic mice.

Bcl-2 inhibitors reduce steroid-insensitive airway inflammation.

BACKGROUND: Asthmatic inflammation is dominated by accumulation of either eosinophils, neutrophils, or both in the airways. Disposal of these inflammatory cells is the key to disease control. Eosinophilic airway inflammation is responsive to corticosteroid treatment, whereas neutrophilic inflammation is resistant and increases the burden of global health care. Corticosteroid-resistant neutrophilic asthma remains mechanistically poorly understood and requires novel effective therapeutic strategies. OBJECTIVE: We sought to explore the underlying mechanisms of airway inflammation persistence, as well as corticosteroid resistance, and to investigate a new strategy of effective treatment against corticosteroid-insensitive neutrophilic asthma. METHODS: Mouse models of either eosinophil-dominated or neutrophil-dominated airway inflammation were used in this study to test corticosteroid sensitivity in vivo and in vitro. We also used vav-Bcl-2 transgenic mice to confirm the importance of granulocytes apoptosis in the clearance of airway inflammation. Finally, the Bcl-2 inhibitors ABT-737 or ABT-199 were tested for their therapeutic effects against eosinophilic or neutrophilic airway inflammation and airway hyperresponsiveness. RESULTS: Overexpression of Bcl-2 protein was found to be responsible for persistence of granulocytes in bronchoalveolar lavage fluid after allergic challenge. This was important because allergen-induced airway inflammation aggravated and persisted in vav-Bcl-2 transgenic mice, in which nucleated hematopoietic cells were overexpressed with Bcl-2 and resistant to apoptosis. The Bcl-2 inhibitors ABT-737 or ABT-199 play efficient roles in alleviation of either eosinophilic or corticosteroid-resistant neutrophilic airway inflammation by inducing apoptosis of immune cells, such as eosinophils, neutrophils, TH2 cells, TH17 cells, and dendritic cells. Moreover, these inhibitors were found to be more efficient than steroids to induce granulocyte apoptosis ex vivo from patients with severe asthma. CONCLUSION: Apoptosis of inflammatory cells is essential for clearance of allergen-induced airway inflammation. The Bcl-2 inhibitors ABT-737 or ABT-199 might be promising drugs for the treatment of airway inflammation, especially for corticosteroid-insensitive neutrophilic airway inflammation.

Impact of loss of BH3-only proteins on the development and treatment of MLL-fusion gene-driven AML in mice.

Inhibition of the apoptosis pathway controlled by opposing members of the Bcl-2 protein family plays a central role in cancer development and resistance to therapy. To investigate how pro-apoptotic Bcl-2 homology domain 3 (BH3)-only proteins impact on acute myeloid leukemia (AML), we generated mixed lineage leukemia (MLL)-AF9 and MLL-ENL AMLs from BH3-only gene knockout mice. Disease development was not accelerated by loss of Bim, Puma, Noxa, Bmf, or combinations thereof; hence these BH3-only proteins are apparently ineffectual as tumor suppressors in this model. We tested the sensitivity of MLL-AF9 AMLs of each genotype in vitro to standard chemotherapeutic drugs and to the proteasome inhibitor bortezomib, with or without the BH3 mimetic ABT-737. Loss of Puma and/or Noxa increased resistance to cytarabine, daunorubicin and etoposide, while loss of Bim protected against cytarabine and loss of Bmf had no impact. ABT-737 increased sensitivity to the genotoxic drugs but was not dependent on any BH3-only protein tested. The AML lines were very sensitive to bortezomib and loss of Noxa conveyed significant resistance. In vivo, several MLL-AF9 AMLs responded well to daunorubicin and this response was highly dependent on Puma and Noxa but not Bim. Combination therapy with ABT-737 provided little added benefit at the daunorubicin dose trialed. Bortezomib also extended survival of AML-bearing mice, albeit less than daunorubicin. In summary, our genetic studies reveal the importance of Puma and Noxa for the action of genotoxics currently used to treat MLL-driven AML and suggest that, while addition of ABT-737-like BH3 mimetics might enhance their efficacy, new Noxa-like BH3 mimetics targeting Mcl-1 might have greater potential.