17 research outputs found
Targeting LOXL2 for cardiac interstitial fibrosis and heart failure treatment
Interstitial fibrosis plays a key role in the development and progression of heart failure. Here, we show that an enzyme that crosslinks collagen-Lysyl oxidase-like 2 (Loxl2)-is essential for interstitial fibrosis and mechanical dysfunction of pathologically stressed hearts. In mice, cardiac stress activates fibroblasts to express and secrete Loxl2 into the interstitium, triggering fibrosis, systolic and diastolic dysfunction of stressed hearts. Antibody-mediated inhibition or genetic disruption of Loxl2 greatly reduces stress-induced cardiac fibrosis and chamber dilatation, improving systolic and diastolic functions. Loxl2 stimulates cardiac fibroblasts through PI3K/AKT to produce TGF-β2, promoting fibroblast-to-myofibroblast transformation; Loxl2 also acts downstream of TGF-β2 to stimulate myofibroblast migration. In diseased human hearts, LOXL2 is upregulated in cardiac interstitium; its levels correlate with collagen crosslinking and cardiac dysfunction. LOXL2 is also elevated in the serum of heart failure (HF) patients, correlating with other HF biomarkers, suggesting a conserved LOXL2-mediated mechanism of human HF
High-Content, High-Throughput Analysis of Cell Cycle Perturbations Induced by the HSP90 Inhibitor XL888
BACKGROUND: Many proteins that are dysregulated or mutated in cancer cells rely on the molecular chaperone HSP90 for their proper folding and activity, which has led to considerable interest in HSP90 as a cancer drug target. The diverse array of HSP90 client proteins encompasses oncogenic drivers, cell cycle components, and a variety of regulatory factors, so inhibition of HSP90 perturbs multiple cellular processes, including mitogenic signaling and cell cycle control. Although many reports have investigated HSP90 inhibition in the context of the cell cycle, no large-scale studies have examined potential correlations between cell genotype and the cell cycle phenotypes of HSP90 inhibition. METHODOLOGY/PRINCIPAL FINDINGS: To address this question, we developed a novel high-content, high-throughput cell cycle assay and profiled the effects of two distinct small molecule HSP90 inhibitors (XL888 and 17-AAG [17-allylamino-17-demethoxygeldanamycin]) in a large, genetically diverse panel of cancer cell lines. The cell cycle phenotypes of both inhibitors were strikingly similar and fell into three classes: accumulation in M-phase, G2-phase, or G1-phase. Accumulation in M-phase was the most prominent phenotype and notably, was also correlated with TP53 mutant status. We additionally observed unexpected complexity in the response of the cell cycle-associated client PLK1 to HSP90 inhibition, and we suggest that inhibitor-induced PLK1 depletion may contribute to the striking metaphase arrest phenotype seen in many of the M-arrested cell lines. CONCLUSIONS/SIGNIFICANCE: Our analysis of the cell cycle phenotypes induced by HSP90 inhibition in 25 cancer cell lines revealed that the phenotypic response was highly dependent on cellular genotype as well as on the concentration of HSP90 inhibitor and the time of treatment. M-phase arrest correlated with the presence of TP53 mutations, while G2 or G1 arrest was more commonly seen in cells bearing wt TP53. We draw upon previous literature to suggest an integrated model that accounts for these varying observations
Effects and Interferences of Emicizumab, a Humanised Bispecific Antibody Mimicking Activated Factor VIII Cofactor Function, on Coagulation Assays
Correction: High-Content, High-Throughput Analysis of Cell Cycle Perturbations Induced by the HSP90 Inhibitor XL888.
Correction: High-Content, High-Throughput Analysis of Cell Cycle Perturbations Induced by the HSP90 Inhibitor XL888
Correction: High-Content, High-Throughput Analysis of Cell Cycle Perturbations Induced by the HSP90 Inhibitor XL888
Selective Allosteric Inhibition of MMP9 Is Efficacious in Preclinical Models of Ulcerative Colitis and Colorectal Cancer
<div><p>Expression of matrix metalloproteinase 9 (MMP9) is elevated in a variety of inflammatory and oncology indications, including ulcerative colitis and colorectal cancer. MMP9 is a downstream effector and an upstream mediator of pathways involved in growth and inflammation, and has long been viewed as a promising therapeutic target. However, previous efforts to target matrix metalloproteinases (MMPs), including MMP9, have utilized broad-spectrum or semi-selective inhibitors. While some of these drugs showed signs of efficacy in patients, all MMP-targeted inhibitors have been hampered by dose-limiting toxicity or insufficient clinical benefit, likely due to their lack of specificity. Here, we show that selective inhibition of MMP9 did not induce musculoskeletal syndrome (a characteristic toxicity of pan-MMP inhibitors) in a rat model, but did reduce disease severity in a dextran sodium sulfate-induced mouse model of ulcerative colitis. We also found that MMP9 inhibition decreased tumor growth and metastases incidence in a surgical orthotopic xenograft model of colorectal carcinoma, and that inhibition of either tumor- or stroma-derived MMP9 was sufficient to reduce primary tumor growth. Collectively, these data suggest that selective MMP9 inhibition is a promising therapeutic strategy for treatment of inflammatory and oncology indications in which MMP9 is upregulated and is associated with disease pathology, such as ulcerative colitis and colorectal cancer. In addition, we report the development of a potent and highly selective allosteric MMP9 inhibitor, the humanized monoclonal antibody GS-5745, which can be used to evaluate the therapeutic potential of MMP9 inhibition in patients.</p></div
MMP9 expression in human CRC and in an orthotopic xenograft mouse model of CRC.
<p>IHC analysis of HCT116-derived xenograft tumors (A, B) or of human CRC tumors (D, E). MMP9 staining from various cellular sources is highlighted as follows: blue arrows, tumor cells; yellow arrows, inflammatory cells; white arrows, stromal cells such as fibroblasts or smooth muscle cells. (A, B) Immunohistochemical staining for MMP9 in HCT116-derived tumors at 200x (A) or 400x (B) magnification. (D, E) Immunohistochemical staining for MMP9 in a human colorectal carcinoma at 200x (D) or 400x (E) magnification. Panels C (HCT116-derived tumors) and F (human CRC) show tissue sections that were incubated with secondary antibody only and demonstrate the absence of non-specific secondary antibody binding (200x magnification).</p
Efficacy of MMP9-targeting antibody in mouse DSS model of colitis.
<p>Treatment was initiated after establishment of colitis (Day 6): Vehicle control, IgG control (30 mg/kg), and AB0046 (30 mg/kg) were dosed every three days, and entanercept (10 mg/kg) was dosed every two days (A) The area under the curve (AUC) was calculated for daily body weight changes in each animal by the trapezoidal rule method;</p><p></p><p></p><p><mi>A</mi><mi>r</mi><mi>e</mi><mi>a</mi><mo>=</mo></p><p><mo>(</mo></p><p></p><p><mi>t</mi><mn>2</mn></p><mo>−</mo><p><mi>t</mi><mn>1</mn></p><p></p><mo>)</mo><p></p><p><mo>[</mo></p><p></p><p></p><p></p><p></p><p><mo>∫</mo></p><p></p><p><mo>(</mo></p><p></p><p><mi>t</mi><mn>1</mn></p><p></p><mo>)</mo><p></p><mo>+</mo><p></p><p><mo>∫</mo></p><p></p><p><mo>(</mo></p><p></p><p><mi>t</mi><mn>2</mn></p><p></p><mo>)</mo><p></p><p></p><p></p><p></p><p></p><p></p><p></p><p></p><mn>2</mn><p></p><p></p><mo>]</mo><p></p><p></p><p></p><p></p>. (B) The incidence of diarrhea was recorded daily and the AUC calculation was performed as above. (C) Endoscopic evaluation was performed on all groups at study termination. Scoring was based on the single most severe lesion observed in the distal 5 cm of colon. (D) Blinded histopathological analysis was performed on colons excised at study termination. The degree of inflammation (primarily macrophages and neutrophils), edema, and necrosis was scored. (E) Images representative of study groups (40X magnification) were taken by a pathologist and highlight areas of inflammation/mucosal necrosis (black arrows) and edema (blue arrows), which are reduced in AB0046 and etanercept-treated animals. Statistical significance was assessed by one-way ANOVA with Dunnett’s Multiple Comparison post-test. P value designations are as follows: * < 0.05, ** < 0.01, *** <0.001, **** < 0.0001.<p></p