Design and synthesis of dual cathepsin L and S inhibitors and antimetastatic activity evaluation in pancreatic cancer cells.

Currently, the migration and invasion of cancer cells remain the main factors of poor prognosis in the majority of cancer patients. Developing an effective antimetastatic agent is crucial for cancer therapy. Our recent research revealed that Cat L and S are expressed concurrently in metastatic pancreatic cancer cells. Asperphenamate analog ASPER-29, which exhibits dual Cat L and S inhibitory potency, showed a definite antimetastatic effect on pancreatic cancer BxPC-3 and PANC-1 cells. To further improve the antimetastatic ability of asperphenamate-type molecules, 24 derivatives were designed and synthesized by a scaffold-hopping strategy. The cathepsin inhibitory activity assay results showed that most of the derivatives exhibited dual inhibitory effects on Cat L and S. Among all derivatives, Compound B1a showed the strongest inhibitory activity, with IC50 values of 4.10 ± 0.14 µM and 1.79 ± 0.11 µM, which were 1.5-fold and 2.8-fold more potent than those of positive drugs against Cat L and S, respectively. Further wound-healing and transwell chamber assays demonstrated that B1a presented significant antimetastatic ability in vitro.

A novel cathepsin L inhibitor prevents the progression of idiopathic pulmonary fibrosis.

In previous work, the target of asperphenamate as a natural product was successfully determined as cathepsin by the natural product consensus pharmacophore strategy. In order to develop accurate SAR (structure-activity relationship) of asperphenamate-type cathepsin inhibitor, we chose several novel analogs with heterocyclic moiety to perform further study. The molecular simulation showed that 4-pyridyl derivative 3 with the greatest cathepsin inhibitory activity presented new interaction modes with protein utilizing its B-ring moiety. And then molecular dynamics (MD) simulation further revealed that 3 and cathepsin kept stable interaction in the binding site, which validated the molecular docking results. In view that cathepsins play an important role in fibrosis and some cathepsin inhibitors display the therapeutic ability for fibrosis, we investigated the anti-fibrotic effect of 3in vitro and in vivo. The results indicated that 3 displayed the strongest inhibitory effect on the formation of α-SMA and collagen I as the protein markers of fibrosis among all tested derivatives. Further in vivo assay confirmed that 3 indeed showed significant inhibitory ability against pulmonary fibrosis by the method of H&E and Masson staining as well as immunohistochemical staining for characteristic α-SMA proteins.

Discovery of a novel cathepsin inhibitor with dual autophagy-inducing and metastasis-inhibiting effects on breast cancer cells.

Drug resistance and cancer cells metastasis have been the leading causes of chemotherapy failure and cancer-associated death in breast cancer patients. In present, various active molecules either exhibiting novel mechanism of action such as inducing autophagy or inhibiting metastasis have been developed to address these problems. However, the compounds exhibiting such dual functions have rarely been described. Previous work in our group showed that TSA, as a synthetic analog of asperphenamate, induced autophagic cell death in breast cancer cells instead of apoptosis. Furthermore, the target enzyme of TSA was predicted to be cathepin L (Cat L) by natural product consensus pharmacophore strategy. Accumulated evidences have shown that cathepsins are closely associated with migration and invasion of breast cancer cells. It seemed likely that TSA-like molecules may possess the dual functions of inducing autophagy and inhibiting metastasis. Therefore, sixty optically active derivatives were firstly designed and synthesized by replacing the A-ring moiety of TSA with other substituted-phenyl sulfonyl groups. Further cathepsin inhibitory activity assay showed that (S, S) and (S, R) isomers displayed no activity against four kinds of cathepsins (L, S, K, B), while all derivatives tested were inactive toward K and B subtypes. Compound 6a with meta-bromo substituent displayed the greatest inhibitory activity, and its inhibitory capability against Cat L and S was 3.9 and 11.5-fold more potent than that of TSA, respectively. Molecular docking also exhibited that 6a formed more hydrogen bonds or π-π contacts with Cat L or S than TSA. In order to determine whether 6a could play dual roles, its anti-cancer mechanism was further investigated. On the one hand, MDC staining experiment and western blotting analysis validated that 6a can induce autophagy in MDA-MB-231 cells. On the other hand, its metastatic inhibitory ability was also confirmed by wound healing and transwell chamber experiment.

Discovery of novel cathepsin inhibitors with potent anti-metastatic effects in breast cancer cells.

It is still challenging to determine the potential targets of natural products, which is essential for further drug research and development. Due to its novel mechanism of action of inducing autophagy effects in breast cancer cells, asperphenamate has received our considerable attention. However, its unknown target inevitably impedes further study. In our previous work, the target enzyme of asperphenamate was predicted as cathepsin by the natural product consensus pharmacophore strategy. Then, asperphenamate and its three derivatives were chosen to study in detail by molecular docking calculations with AutoDock 4 suite. The docking results showed the three derivatives interacted more tightly with either cathepsin L or cathepsin S than with asperphenamate. The ortho-benzyloxyl phenylacetyl derivative 1 andp-toluenesulfonyl derivative 3 showed similar interactions with cathepsin L and adopted a better geometric shape within the binding pocket than did the N-CBZ-piperidyl analog 2. On the other hand, compound 2 formed more hydrogen bonds than 1 and 3 to make it tightly bind within cathepsin S. The cathepsin inhibitory activity assay verified the molecular simulation results. Compound 2 was remarkably less active than 1 and 3 against cathepsin L. However, compound 2 showed the strongest potency against cathepsin S with IC50 of 13.12 ±â€¯0.29 µM. Considering that cathepsin S plays a vital role in the process of metastasis in breast cancer cells, the inhibitory effect of 2 on migration and invasion was further studied in human breast cancer MDA-MB-231 cells by wound healing and transwell chamber assays. The results illustrated that 2 exhibited an apparent inhibitory ability to the metastasis of MDA-MB-231 cells. Next, 2 will be chosen as a lead compound to develop novel double functional chemotherapeutic agents with both novel mechanisms of action against apoptosis-resistant cancer cells, such as inducing autophagy and inhibiting cancer metastasis.

Comparison of the efficacy of three isolated gastrocnemius recession procedures in a cadaveric model of gastrocnemius tightness.

PURPOSE: Many surgical techniques for isolated gastrocnemius recession have been described. The purpose of the present study is to compare their therapeutic efficacy and intrinsic stability on a fresh cadaveric model of gastrocnemius tightness. METHODS: The cadaveric model was established by distracting the knee with spacers composed of low-temperature thermoplastic material, and was identified by the Silfverskiöld test. Procedures of gastrocnemius recession described by Barouk, Baumann and Strayer were performed. The lengthening distance and improvement of ankle dorsiflexion were measured. RESULTS: The mean ankle angle of the cadaveric models was -15.4° with the knee fully extended, and 8.4° with the knee flexed. The increased angle achieved by the Strayer procedure was significantly greater than that achieved by the Barouk procedure and the Baumann procedure with one cut (P < 0.05), but similar to that of the Baumann procedure with two cuts (P > 0.05). Compared with the intramuscular lengthening of the gastrocnemius (Baumann and Barouk procedures), the stability of the lengthening after the Strayer procedure was the lowest. CONCLUSION: The three techniques of isolated gastrocnemius recession have different efficacy and stability in cadaver trials, depending on their own anatomical characteristics. Our study supplies an anatomical guide for the selection of a proper procedure.

Bootstrap-adjusted quasi-likelihood information criteria for mixed model selection.

We propose two model selection criteria relying on the bootstrap approach, denoted by QAICb1 and QAICb2, in the framework of linear mixed models. Similar to the justification of Akaike Information Criterion (AIC), the proposed QAICb1 and QAICb2 are proved as asymptotically unbiased estimators of the Kullback-Leibler discrepancy between a candidate model and the true model. However, they are defined on the quasi-likelihood function instead of the likelihood and are proven to be asymptotically equivalent. The proposed selection criteria are constructed by the quasi-likelihood of a candidate model and a bias estimation term in which the bootstrap method is adopted to improve the estimation for the bias caused by using the candidate model to estimate the true model. The simulations across a variety of mixed model settings are conducted to demonstrate that the proposed selection criteria outperform some other existing model selection criteria in selecting the true model. Generalized estimating equations (GEE) are utilized to calculate QAICb1 and QAICb2 in the simulations. The effectiveness of the proposed selection criteria is also demonstrated in an application of Parkinson's Progression Markers Initiative (PPMI) data.

Mid-term Results of Intramuscular Lengthening of Gastrocnemius and/or Soleus to Correct Equinus Deformity in Flatfoot.

BACKGROUND: Intramuscular lengthening of the gastrocnemius and/or soleus (Baumann procedure) is widely used in patients who have cerebral palsy, with several advantages over other lengthening techniques. Tightness of the gastrocnemius or gastrocnemius-soleus complex has been confirmed to be related to flatfoot deformity. The purpose of this study was to evaluate the mid-term results of the Baumann procedure as a part of the treatment of flatfoot with equinus deformity. METHODS: We reviewed 35 pediatric and adult patients (43 feet) with flatfoot who underwent the Baumann procedure for the concomitant equinus deformity. The mean duration of follow-up was 39.4 months. Preoperative and follow-up evaluations included the maximal angle of dorsiflexion of the ankle with the knee fully extended and with the knee flexed to 90 degrees, the American Orthopaedic Foot & Ankle Society ankle-hindfoot (AOFAS-AH) scores, and postoperative complications. RESULTS: Preoperatively, the mean angle of passive ankle dorsiflexion with the knee extended was -4.7 ± 2.7 degrees and that with the knee flexed was 2.3 ± 2.5 degrees. At the final follow-up, both values improved significantly by a mean of 13.6 degrees (P < .001) and 9.7 degrees (P < .001), respectively. The average AOFAS-AH scores improved from 56.8 points preoperatively to 72.1 at the final follow-up. Recurrence of equinus was observed in 3 patients (4 feet). There were no cases of overcorrection, neurovascular injury, or healing problems. CONCLUSIONS: Our results indicate that the Baumann procedure can effectively and sequentially correct the tightness of the gastrocnemius or the gastrocnemius-soleus complex in patients with flatfoot deformity, without obvious postoperative complications. LEVEL OF EVIDENCE: Level IV, retrospective case series.