After kidney ischemia/reperfusion (I/R) injury, monocytes home to the kidney and differentiate into activated macrophages. Whereas proinflammatory macrophages contribute to the initial kidney damage, an alternatively activated phenotype can promote normal renal repair. The microenvironment of the kidney during the repair phase mediates the transition of macrophage activation from a proinflammatory to a reparative phenotype. In this study, we show that macrophages isolated from murine kidneys during the tubular repair phase after I/R exhibit an alternative activation gene profile that differs from the canonical alternative activation induced by IL-4-stimulated STAT6 signaling. This unique activation profile can be reproduced in vitro by stimulation of bone marrow-derived macrophages with conditioned media from serum-starved mouse proximal tubule cells. Secreted tubular factors were found to activate macrophage STAT3 and STAT5 but not STAT6, leading to induction of the unique alternative activation pattern. Using STAT3-deficient bone marrow-derived macrophages and pharmacologic inhibition of STAT5, we found that tubular cell-mediated macrophage alternative activation is regulated by STAT5 activation. Both in vitro and after renal I/R, tubular cells expressed GM-CSF, a known STAT5 activator, and this pathway was required for in vitro alternative activation of macrophages by tubular cells. Furthermore, administration of a neutralizing antibody against GM-CSF after renal I/R attenuated kidney macrophage alternative activation and suppressed tubular proliferation. Taken together, these data show that tubular cells can instruct macrophage activation by secreting GM-CSF, leading to a unique macrophage reparative phenotype that supports tubular proliferation after sterile ischemic injury.
557 PHY906 is a traditional Chinese botanical formulation consisting of 4 different herbs, and it has been used for some 1800 years to treat gastrointestinal ailments, some of which are commonly observed side effects in cancer patients undergoing chemotherapy. We have previously reported that PHY906 reduced chemotherapy-induced toxicities including body weight loss and mortality, and that it also enhanced the antitumor efficacy of a broad-spectrum of anticancer agents, such as CPT-11, 5-FU, CPT-11/5-FU/LV, VP-16, and L-OddC in in vivo animal models. PHY906 has been co-administrated with either the oral 5-FU prodrug capecitabine or CPT-11 in human hepatocellular xenografts mouse model, and with gemcitabine in mouse pancreatic cancer model. We have shown that PHY906 significantly enhanced the therapeutic index of chemotherapeutic agents studied in both hepatocellular and pancreatic cancer models. Co-administration of PHY906 with either capecitabine or gemcitabine in animal models did not alter the pharmacokinetic profile of capecitabine, gemcitabine, or their respective metabolites. Our biochemical studies revealed that the PHY906 formulation possesses a wide range of pharmacological activities. The potential mechanism(s) of action of PHY906 include (1) enhancement of oral uptake of pharmacologically active agents with inhibition in intestinal CYP3A4; (2) inhibition of NF-κB activity; (3) inhibition of MMP activity; and (4) destruction of the integrity of sinusoids in hepatoma. These pre-clinical in vivo studies have provided rationale for developing PHY906 in the clinical setting. A phase I/IIA double-blind placebo-controlled, dose escalation study of PHY906 was initiated to evaluate the potential effect of PHY906 in modulating CPT-11-induced diarrhea associated with the bolus, weekly schedule of CPT-11/5-FU/LV in advanced colorectal cancer patients. A second phase I/II open-label dose escalation clinical trial has just been opened to patient accrual to evaluate the role of PHY906 in combination with capecitabine in the treatment of hepatocellular carcinoma.
Podocytes are specialized actin-rich epithelial cells that line the kidney glomerular filtration barrier. The interface between the podocyte and the glomerular basement membrane requires integrins, and defects in either α3 or β1 integrin, or the α3β1 ligand laminin result in nephrotic syndrome in murine models. The large cytoskeletal protein talin1 is not only pivotal for integrin activation, but also directly links integrins to the actin cytoskeleton. Here, we found that mice lacking talin1 specifically in podocytes display severe proteinuria, foot process effacement, and kidney failure. Loss of talin1 in podocytes caused only a modest reduction in β1 integrin activation, podocyte cell adhesion, and cell spreading; however, the actin cytoskeleton of podocytes was profoundly altered by the loss of talin1. Evaluation of murine models of glomerular injury and patients with nephrotic syndrome revealed that calpain-induced talin1 cleavage in podocytes might promote pathogenesis of nephrotic syndrome. Furthermore, pharmacologic inhibition of calpain activity following glomerular injury substantially reduced talin1 cleavage, albuminuria, and foot process effacement. Collectively, these findings indicate that podocyte talin1 is critical for maintaining the integrity of the glomerular filtration barrier and provide insight into the pathogenesis of nephrotic syndrome.
<p>PDF file - 128K, Fig. S1. BRCA1 and BRCA2-defective EOC cells are sensitive to triapine. Fig. S2. Triapine abrogates olaparib-induced RPA phosphorylation in SKOV-3 cells. Fig. S3. Depletion of Chk1 partially reverses the inhibitory effects of triapine on RPA32 and CtIP phosphorylation. Fig. S4. Depletion of CtIP by siRNA. Table S1. CI values of triapine and olaparib combination determined by MTS viability assay in NTC and BRCA1-kd SKOV3 cells. Table S2A. CI values of triapine and olaparib combination at a constant ratio in NTC and BRCA1-kd SKOV3 cells (Fig. 2A). Table S2B. CI values of triapine and olaparib combination in SKOV-3 cells (Fig. 2B). Table S2C. CI values of triapine and olaparib combination in BG-1 cells (Fig. 2C). Table S2D. CI values of triapine and olaparib combination in PEO4 cells (Fig. 2D).</p>
PHY906 is a Chinese medicine formulation prepared from four medicinal herbs for adjuvant cancer chemotherapy. In this paper, liquid chromatography/electrospray ionization time-of-flight mass spectrometry (LC/ESI-TOFMS) was used to clarify the chemical composition of PHY906. The aqueous extract of PHY906 was separated on a Waters Atlantis C(18) column, and was eluted with acetonitrile/0.05% (v/v) formic acid. The separated compounds were identified with pure standards, or tentatively characterized by analyzing their mass spectra recorded in both negative and positive ion polarity modes. Further structural information was obtained from in-source fragmentation. Based on the LC/MS analysis, we proposed the structures for 64 bioactive compounds, including flavonoids, triterpene saponins, and monoterpene glycosides. All the compounds identified from PHY906 were further assigned in the four individual herbs, and some of them are reported for the first time.
Platinum resistance may be attributable to inherent or acquired proficiency in homologous recombination repair (HRR) in epithelial ovarian cancer (EOC). The objective of this study was to evaluate the efficacy of the small molecule inhibitor triapine to disrupt HRR and sensitise BRCA wild-type EOC cells to platinum-based combination therapy in vitro and in vivo. The sensitivity of BRCA wild-type cancer cells to olaparib, cisplatin, carboplatin, doxorubicin, or etoposide in combination with triapine was evaluated by clonogenic survival assays. The effects of triapine on HRR activity in cells were measured with a DR-GFP reporter assay. The ability of triapine to enhance the effects of the carboplatin-doxil combination on EOC tumour growth delay was determined using a xenograft tumour mouse model. Platinum resistance is associated with wild-type BRCA status. Triapine inhibits HRR activity and enhances the sensitivity of BRCA wild-type cancer cells to cisplatin, olaparib, and doxorubicin. However, sequential combination of triapine and cisplatin is necessary to achieve synergism. Moreover, triapine potentiates platinum-based combination therapy against BRCA wild-type EOC cells and produces significant delay of EOC tumour growth. Triapine promises to augment the clinical efficacy of platinum-based combination regimens for treatment of platinum-resistant EOC with wild-type BRCA and proficient HRR activity.
Human Macrophage Migration Inhibitory Factor (MIF) is a trimeric cytokine implicated in a number of inflammatory and autoimmune diseases and cancer. We previously reported that the dye p425 (Chicago Sky Blue), which bound MIF at the interface of two MIF trimers covering the tautomerase and allosteric pockets, revealed a unique strategy to block MIF's pro-inflammatory activities. Structural liabilities, including the large size, precluded p425 as a medicinal chemistry lead for drug development. We report here a rational design strategy linking only the fragment of p425 that binds over the tautomerase pocket to the core of ibudilast, a known MIF allosteric site-specific inhibitor. The chimeric compound, termed L2-4048, was shown by X-ray crystallography to bind at the allosteric and tautomerase sites as anticipated. L2-4048 retained target binding and blocked MIF's tautomerase CD74 receptor binding, and pro-inflammatory activities. Our studies lay the foundation for the design and synthesis of smaller and more drug-like compounds that retain the MIF inhibitory properties of this chimera.
Polycystic kidney disease (PKD) exhibits an inflammatory component, but the contribution of inflammation to cyst progression is unknown. Macrophages promote the proliferation of tubular cells following ischemic injury, suggesting that they may have a role in cystogenesis. Furthermore, cultured Pkd1-deficient cells express the macrophage chemoattractants Mcp1 and Cxcl16 and stimulate macrophage migration. Here, in orthologous models of both PKD1 and PKD2, abnormally large numbers of alternatively activated macrophages surrounded the cysts. To determine whether pericystic macrophages contribute to the proliferation of cyst-lining cells, we depleted phagocytic cells from Pkd1(fl/fl);Pkhd1-Cre mice by treating with liposomal clodronate from postnatal day 10 until day 24. Compared with vehicle-treated controls, macrophage-depleted mice had a significantly lower cystic index, reduced proliferation of cyst-lining cells, better-preserved renal parenchyma, and improved renal function. In conclusion, these data suggest that macrophages home to cystic areas and contribute to cyst growth. Interruption of these homing and proliferative signals could have therapeutic potential for PKD.
