Activation loop targeting strategy for design of receptor-interacting protein kinase 2 (RIPK2) inhibitors

Development of selective kinase inhibitors remains a challenge due to considerable amino acid sequence similarity among family members particularly in the ATP binding site. Targeting the activation loop might offer improved inhibitor selectivity since this region of kinases is less conserved. However, the strategy presents difficulties due to activation loop flexibility. Herein, we report the design of receptor-interacting protein kinase 2 (RIPK2) inhibitors based on pan-kinase inhibitor regorafenib that aim to engage basic activation loop residues Lys169 or Arg171. We report development of CSR35 that displayed >10-fold selective inhibition of RIPK2 versus VEGFR2, the target of regorafenib. A co-crystal structure of CSR35 with RIPK2 revealed a resolved activation loop with an ionic interaction between the carboxylic acid installed in the inhibitor and the side-chain of Lys169. Our data provides principle feasibility of developing activation loop targeting type II inhibitors as a complementary strategy for achieving improved selectivity.     

Functional analysis of Cullin 3 E3 ligases in tumorigenesis

Cullin 3-RING ligases (CRL3) play pivotal roles in the regulation of various physiological and pathological processes, including neoplastic events. The substrate adaptors of CRL3 typically contain a BTB domain that mediates the interaction between Cullin 3 and target substrates to promote their ubiquitination and subsequent degradation. The biological implications of CRL3 adaptor proteins have been well described where they have been found to play a role as either an oncogene, tumor suppressor, or can mediate either of these effects in a context-dependent manner. Among the extensively studied CRL3-based E3 ligases, the role of the adaptor protein SPOP (speckle type BTB/POZ protein) in tumorigenesis appears to be tissue or cellular context dependent. Specifically, SPOP acts as a tumor suppressor via destabilizing downstream oncoproteins in many malignancies, especially in prostate cancer. However, SPOP has largely an oncogenic role in kidney cancer. Keap1, another well-characterized CRL3 adaptor protein, likely serves as a tumor suppressor within diverse malignancies, mainly due to its specific turnover of its downstream oncogenic substrate, NRF2 (nuclear factor erythroid 2-related factor 2). In accordance with the physiological role the various CRL3 adaptors exhibit, several pharmacological agents have been developed to disrupt its E3 ligase activity, therefore blocking its potential oncogenic activity to mitigate tumorigenesis.     

Human placenta-derived mesenchymal stem cells loaded on linear ordered collagen scaffold improves functional recovery after completely transected spinal cord injury in canine

Traumatic spinal cord injury(SCI) is a major challenge in the clinic. In this study, we sought to examine the synergistic effects of linear ordered collagen scaffold(LOCS) and human placenta-derived mesenchymal stem cells(hPMSCs) when transplanted into completely transected beagle dogs. After 36 weeks observation, we found that LOCS+hPMSCs implants promoted better hindlimb locomotor recovery than was observed in the non-treatment(control) group and LOCS group. Histological analysis showed that the regenerated tissue after treatment was well integrated with the host tissue, and dramatically reduced the volume of cystic and chondroitin sulfate proteoglycans(CSPGs) expression. Furthermore, the LOCS+hPMSCs group also showed more neuron-specific βIII-tubulin(Tuj-1)-and NeuN-positive neurons in the lesion area, as well as axonal regeneration, remyelination and synapse formation in the lesion site. Additionally, dogs in the LOCS+hPMSCs group experienced enhanced sprouting of both ascending(CGRP-positive) sensory fibers and descending(5-HT-and TH-positive) motor fibers at the lesion area. All these data together suggested that the combined treatment had beneficial effects on neuronal regeneration and functional improvement in a canine complete transection model. Therefore, LOCS+hPMSCs implantation holds a great promise for bridging the nerve defect and may be clinically useful in the near future.     

Oplopane Sesquiterpenes from Ligularia knorringiana and Their Anti‐Complementary Activity

Three new oplopane sesquiterpenes, knorringianalarins D – F ( 1 – 3 , respectively), and five known analogues ( 4 – 8 , respectively), were isolated from the roots and rhizomes of Ligularia knorringiana. The structures of three new compounds were identified as 4‐acetoxy‐11α,12‐epoxy‐2β‐hydroxy‐3β‐(2‐methylbutyryloxy)‐9α‐(4‐methylsenecioyloxy)oplop‐10(14)‐ene ( 1 ), 3β,4‐diacetoxy‐9α‐(4‐acetoxy‐4‐methylsenecioyloxy)‐11α,12‐epoxy‐8α‐(2‐methylbutyryloxy)oplop‐10(14)‐ene ( 2 ), and (1R,5R,6R,7R,9R)‐5,9,11‐trihydroxy‐4,15‐dinoroplop‐10(14)‐en‐3‐one ( 3 ) based on spectroscopic methods including 1D‐ and 2D‐NMR, mass spectrometry, and CD spectroscopy techniques. All compounds were evaluated for their anti‐complementary activity on the classical pathway of the complement system in vitro. Among which, three oplopane sesquiterpenes ( 3 , 7 , and 8 ) exhibited better anti‐complementary effects with CH₅₀ values ranging from 0.33 to 0.89 mm , which are plausible candidates for developing potent anti‐complementary agents.     

Inhibition of EGF expression and NF-κB activity by treatment with quercetin leads to suppression of angiogenesis in nasopharyngeal carcinoma

The present study was performed to investigate the effect of quercetin on nasopharyngeal carcinoma (NPC) angiogenesis. The real-time RT-PCR and enzyme-linked immunosorbent assays (ELISA) were performed to analyze the expression levels of vascular endothelial growth factor (VEGF) in nasopharyngeal carcinoma cell lines prior to and after the quercetin treatment. Effect of quercetin on the rate of cell proliferation was measured by MTT assay. It was observed that quercetin treatment at a concentration of 10 mg/mL reduced the rate of NPC039 cell viability to 36% compared to control after 24 h. The expression of VEGF and activity of NF-κB was also markedly reduced. The ability of tube formation in HUVECs was inhibited significantly on exposure to quercetin compared to the untreated cells. Therefore, quercetin plays an important role in the inhibition of NPC039 nasopharyngeal carcinoma and can be of therapeutic importance.