Circulating Tumor Cells as an Indicator of Treatment Options for Hepatocellular Carcinoma Less Than or Equal to 3†cm in Size: A Multi-Center, Retrospective Study.

Background: The status of circulating tumor cells (CTCs) is related to the recurrence of hepatocellular carcinoma (HCC), which is also one of the reasons for the poor prognosis of HCC. The purpose of this study was to explore whether CTCs can help guide the choice of treatment methods for HCC. Methods: This study is a multicenter retrospective study, including 602 patients with HCC. CTCs were detected in the overall cohort before operation. There were 361 patients in the training cohort and 241 patients in the validation cohort. Patients were divided into CTC-negative group (CTCs = 0/5 mL) and the CTC-positive group (CTCs ≥ 1/5 mL) according to CTCs status. Subgroup analysis was performed according to CTCs status. We compared overall survival, and recurrence outcomes for HCC patients with different CTC statuses after undergoing radiofrequency ablation (RFA) or surgical resection (SR). Results: There was no significant difference in overall survival (OS) and recurrence-free survival (RFS) between the RFA group and SR group for CTC-negative patients in both the training cohort and the validation cohort (P > 0.05). However, among CTC-positive patients, the clinical outcome of patients in the SR group was significantly better than those in the RFA group. CTC-positive patients who underwent RFA had increased early recurrence compared to those who underwent SR. RFA is an independent risk factor for survival and recurrence in CTC-positive HCC patients. Conclusions: The CTC status could serve as an indicator to guide the choice between surgical resection or radiofrequency ablation for early hepatocellular carcinoma. Surgical resection is recommended for CTC-positive patients.

Application of Transparent Cap-assisted Choledochoscopy in Endoscopic Gallbladder-preserving Surgery.

BACKGROUND: The gold standard treatment for cholecystolithiasis is laparoscopic cholecystectomy. However, the complications of cholecystectomy have led to adoption of gallbladder-preserving surgery. The study was to investigate significance of transparent cap-assisted choledochoscopy in gallbladder-preserving surgery. MATERIALS AND METHODS: This is a retrospective study of patients who underwent gallbladder-preserving surgery by laparoscopic choledochoscopy along with choledochoscopy with or without a transparent cap from January 2018 to September 2018 in our hospital. The differences in the duration of gallbladder exploration, surgical complications, adverse events, and the recurrence of stones within 6 months after surgery were compared between 2 groups. RESULTS: Fifty patients underwent laparoscopic choledochoscopy along with choledochoscopy without transparent cap (Group A), while 50 patients underwent laparoscopic along with transparent cap-assisted choledochoscopy (Group B). Gallbladder exploration time was 27.96±12.24 minutes in Group A, and 12.04±6.01 minutes in Group B. One case had stone recurrence within 6 months in Group B, while 8 cases had stone recurrence in group A. CONCLUSIONS: Comparing with laparoscope combined with choledochoscopy, transparent cap-assisted choledochoscopy has advantages in gallbladder-preserving surgery.

Activation of MET promotes resistance to sorafenib in hepatocellular carcinoma cells via the AKT/ERK1/2-EGR1 pathway.

Sorafenib is an oral multikinase inhibitor that has become an established therapeutic approach in advanced hepatocellular carcinoma (HCC). However, the benefit of sorafenib in clinical therapy is often affected by drug resistance. Therefore, it is important to explore the mechanisms underlying sorafenib resistance and to develop individualized therapeutic strategies for coping with this problem. In this study, we found that addition of HGF to sorafenib-treated HCC cells activated MET and re-stimulated the downstream AKT and ERK1/2 pathways. Thereby, restored sorafenib-treated HCC cells proliferation, migration and invasion ability, and rescued cells from apoptosis. In addition, we found that HGF treatment of HCC cells induced early growth response protein (EGR1) expression, which is involved in sorafenib resistance. Importantly, the HGF rescued effect in sorafenib-treated HCC cells could be abrogated by inhibiting MET activation with PHA-665752 or by downregulating EGR1 expression with small interfering RNA (siRNA). Therefore, inhibition of the HGF/MET pathway may improve response to sorafenib in HCC, and combination therapy should be further investigated.

Magnetic resonance imaging tracking and assessing repair function of the bone marrow mesenchymal stem cells transplantation in a rat model of spinal cord injury.

The transplantation of bone marrow mesenchymal stem cells (BMSCs) to repair spinal cord injury (SCI) has become a promising therapy. However, there is still a lack of visual evidence directly implicating the transplanted cells as the source of the improvement of spinal cord function. In this study, BMSCs were labeled with NF-200 promoter and lipase-activated gadolinium-containing nanoparticles (Gd-DTPA-FA). Double labeled BMSCs were implanted into spinal cord transaction injury in rat models in situ, the function recovery was evaluated on 1st, 7th, 14th, 28 th days by MRI, Diffusion Tensor Imaing, CT imaging and post-processing, and histological observations. BBB scores were used for assessing function recovery. After transplantation of BMSCs, the hypersignal emerged in spinal cord in T1WI starting at day 7 that was focused at the injection site, which then increased and extended until day 14. Subsequently, the increased signal intensity area rapidly spread from the injection site to entire injured segment lasting four weeks. The diffusion tensor tractography and histological analysis both showed the nerve fibre from dividing to connecting partly. Immunofluorescence showed higher expression of NF-200 in Repaired group than Injury group. Electron microscopy showed detachment and loose of myelin lamellar getting better in Repaired group compared with the Injury group. BBB scores in Repaired group were significantly higher than those of injury animals. Our study suggests that the migration and distribution of Gd-DTPA-FA labeled BMSCs can be tracked using MRI. Transplantation of BMSCs represents a promising potential strategy for the repair of SCI.

Targeted Inhibition of Leucine-Rich Repeat and Immunoglobulin Domain-Containing Protein 1 in Transplanted Neural Stem Cells Promotes Neuronal Differentiation and Functional Recovery in Rats Subjected to Spinal Cord Injury.

OBJECTIVE: Leucine-rich repeat and immunoglobulin domain-containing protein (LINGO)-1 is expressed in neural stem cells, and its neutralization results in sustained neuronal immaturity. Thus, targeted inhibition of LINGO-1 via RNA interference may enhance transplanted neural stem cell survival and neuronal differentiation in vivo. Furthermore, LINGO-1 RNA interference in neural stem cells represents a potential therapeutic strategy for spinal cord injury. DESIGN: Department of Spine Surgery, First Affiliated Hospital of Sun Yat-sen University. SETTING: Translational Medicine Center Research Laboratory, First Affiliated Hospital of Sun Yat-sen University. SUBJECTS: Female Sprague-Dawley rats. INTERVENTIONS: The animals were divided into three groups that underwent laminectomy and complete spinal cord transection accompanied by transplantation of control-RNA interference-treated or LINGO-1-RNA interference-treated neural stem cells at the injured site in vivo. In vitro, neural stem cells were divided into four groups for the following treatments: control, control RNA interference lentivirus, LINGO-1 RNA interference lentivirus and LINGO-1 complementary DNA lentivirusand the Key Projects of the Natural Science Foundation of Guangdong Province (No. S2013020012818). MEASUREMENTS AND MAIN RESULTS: Neural stem cells in each treatment group were examined for cell survival and neuronal differentiation in vitro and in vivo via immunofluorescence and Western blot analysis. Axonal regeneration and tissue repair were assessed via retrograde tracing using Fluorogold, electron microscopy, hematoxylin-eosin staining and MRI. Rats were also examined for functional recovery based on the measurement of spinal cord-evoked potentials and the Basso-Beattie-Bresnahan score. LINGO-1-RNA interference-treated neural stem cell transplantation increased tissue repair and functional recovery of the injured spinal cord in rats. Similarly, LINGO-1 RNA interference increased neural stem cell survival and neuronal differentiation in vitro. The mechanism underlying the effect of LINGO-1 RNA interference on the injured rat spinal cord may be that the significant inhibition of LINGO-1 expression in neural stem cells inactivated the RhoA and Notch signaling pathways, which act downstream of LINGO-1. CONCLUSIONS: Our findings indicate that transplantation of LINGO-1-RNA interference-treated neural stem cells facilitates functional recovery after spinal cord injury and represents a promising potential strategy for the repair of spinal cord injury.

Tivantinib induces G2/M arrest and apoptosis by disrupting tubulin polymerization in hepatocellular carcinoma.

BACKGROUND: Tivantinib has been described as a highly selective inhibitor of MET and is currently in a phase III clinical trial for the treatment of hepatocellular carcinoma (HCC). However, the mechanism of tivantinib anti-tumor effect has been questioned by recent studies. RESULTS: We show that tivantinib indiscriminately inhibited MET dependent and independent HCC cells proliferation. In contrast, other MET inhibitors, JNJ-38877605 and PHA-665752, just specifically inhibited the growth of MET dependent HCC cells. Tivantinib neither inhibit constitutive MET phosphorylation nor HGF-induced MET phosphorylation in HCC cells. In the microtubule polymerization analysis, tivantinib affected microtubule dynamics by a mechanism as a microtubule depolymerizer. Interesting, unlike other microtubule-targeting agents, paclitaxel and vincristine, tivantinib showed similar anti-proliferative activity in parental and multidrug-resistant cells. Further studies demonstrated that tivantinib induced a G2/M arrest and promoted apoptosis by both intrinsic and extrinsic pathway. The in vivo efficacy evaluation showed that tivantinib exhibited a good anti-tumor growth activity with anti-proliferative and pro-apoptotic effects. CONCLUSIONS: The potent anti-tumor activity of tivantinib in HCC was achieved by targeting microtubule. Tivantinib treatment for patients with HCC should not be selected based on MET status.

Cabozantinib reverses multidrug resistance of human hepatoma HepG2/adr cells by modulating the function of P-glycoprotein.

BACKGROUND & AIMS: Cabozantinib, a small-molecule multitargeted tyrosine kinase inhibitor, has entered into a phase III clinical trial for the treatment of hepatocellular carcinoma (HCC). This study assessed the mechanistic effect of cabozantinib on the reversal of P-glycoprotein (P-gp)-mediated multidrug resistance (MDR). METHODS: CCK-8 assays and tumour xenografts were used to investigate the reversal of MDR in vitro and in vivo respectively. Substrate retention assays were evaluated by fluorescence microscope and flow cytometry. Western blotting was used to detect protein expression levels. mRNA expression was determined by qPCR. The ATPase activity of P-gp was investigated using Pgp-Glo(™) assay systems. The binding mechanism of cabozantinib to P-gp at the molecular level was evaluated using docking analysis. RESULTS: Cabozantinib enhanced the cytotoxicity of P-gp substrate drugs in HepG2/adr and HEK293-MDR1 cells but had no effect on non-P-gp substrates. In addition, cabozantinib increased the accumulation of P-gp substrates in HepG2/adr cells but had no effect in HepG2 cells. Furthermore, cabozantinib did not alter the expression of P-gp mRNA or protein but did stimulate the activity of P-gp ATPase. The docking study indicated that cabozantinib and verapamil may partially share a binding site on P-gp. The reversal concentrations of cabozantinib did not affect the expression of MET, AKT and ERK1/2. Significantly, cabozantinib increased the inhibitory efficacy of doxorubicin in P-gp-overexpressing HepG2/adr cell xenografts in nude mice. CONCLUSION: Cabozantinib reverses P-gp-mediated MDR by directly inhibiting the efflux function of P-gp, indicating that cabozantinib may help to reverse P-gp-mediated MDR in HCC and other cancer chemotherapy.

[Laparoscopic hepatectomy for recurrent hepatocellular carcinoma after previous open hepatectomy].

OBJECTIVE: To evaluate the feasibility, safety and long-term outcomes of laparoscopic repeat hepatectomy for recurrent hepatocellular carcinoma (HCC) following previous resection. METHODS: Between January 2003 and January 2011, 14 patients with recurrent HCC were carefully selected to undergo repeat laparoscopic hepatectomy, among which 9 patients were male, 5 patients were female, and the average age was 54 years. Prior to re-resection, all patients had undergone at least one open hepatectomy for HCC. The perioperative and long-term outcomes of these patients were retrospectively analyzed. RESULTS: Repeat laparoscopic hepatectomy for these 14 patients were successfully performed without major perioperative complications. The mean operative time, intraoperative blood loss and hospital stay were (124 ± 82) minutes, (112 ± 43) ml and (7 ± 4) days, respectively. The mean follow-up period was 23 months (range 14 to 42 months). At the time of follow-up, 11 patients were still alive, among which 3 patients developed recurrent disease and 8 patients remained disease free. One patient died of liver dysfunction at 21 months, and another 2 patients died of tumor recurrence at 17, 31 months, respectively. CONCLUSION: Laparoscopic surgery for recurrent HCC remains a viable option for repeat hepatectomy in selected patients who have undergone open hepatectomy.

Cabozantinib suppresses tumor growth and metastasis in hepatocellular carcinoma by a dual blockade of VEGFR2 and MET.

PURPOSE: MET signaling has been suggested a potential role in hepatocellular carcinoma (HCC) and associated with prometastasis during antiangiogenesis therapy. We investigated the potential association between MET expression and therapeutic response to sorafenib in patients with HCC. Antitumor effects of cabozantinib, a dual inhibitor of MET and VEGFR2, were examined in cultured HCC cells as well as in vivo models. EXPERIMENTAL DESIGN: Total MET and phosphorylated MET (p-MET) were measured in 29 resected HCC specimens, and correlated with response to sorafenib as postoperative adjuvant therapy. In the second set of experiments using cultured HCC cells, and mouse xenograft and metastatic models, effects of cabozantinib were examined. RESULTS: High level of p-MET in resected HCC specimens was associated with resistance to adjuvant sorafenib therapy. In cultured HCC cells that expressed p-MET, cabozantinib inhibited the activity of MET and its downstream effectors, leading to G1-phase arrest. Cabozantinib inhibited tumor growth in p-MET-positive and p-MET-negative HCC by decreasing angiogenesis, inhibiting proliferation, and promoting apoptosis, but it exhibited more profound efficacy in p-MET-positive HCC xenografts. Cabozantinib blocked the hepatocyte growth factor (HGF)-stimulated MET pathway and inhibited the migration and invasion of the HCC cells. Notably, cabozantinib reduced the number of metastatic lesions in the lung and liver in the experimental metastatic mouse model. CONCLUSIONS: Patients with HCC with high level of p-MET are associated with resistance to adjuvant sorafenib treatment. The dual blockade of VEGFR2 and MET by cabozantinib has significant antitumor activities in HCC, and the activation of MET in HCC may be a promising efficacy-predicting biomarker. Clin Cancer Res; 20(11); 2959-70. ©2014 AACR.

Specific lipase-responsive polymer-coated gadolinium nanoparticles for MR imaging of early acute pancreatitis.

Currently, available methods for diagnosis of acute pancreatitis (AP) are mainly dependent on serum enzyme analysis and imaging techniques that are too low in sensitivity and specificity to accurately and promptly diagnose AP. The lack of early diagnostic tools highlights the need to search for a highly effective and specific diagnostic method. In this study, we synthesized a conditionally activated, gadolinium-containing, nanoparticle-based MRI nanoprobe as a diagnostic tool for the early identification of AP. Gadolinium diethylenetriaminepentaacetic fatty acid (Gd-DTPA-FA) nanoparticles were synthesized by conjugation of DTPA-FA ligand and gadolinium acetate. Gd-DTPA-FA exhibited low cytotoxicity and excellent biocompatibility when characterized in vitro and in vivo studies. L-arginine induced a gradual increase in the intensity of the T1-weighted MRI signal from 1 h to 36 h in AP rat models. The increase in signal intensity was most significant at 1 h, 6 h and 12 h. These results suggest that the Gd-DTPA-FA as an MRI contrast agent is highly efficient and specific to detect early AP.

Effect of BIBF 1120 on reversal of ABCB1-mediated multidrug resistance.

BACKGROUND: The overexpression of ATP-binding cassette (ABC) transporters is one of the main causes of multi-drug resistance (MDR) which represents a major obstacle to the success of cancer chemotherapy. In this study, we examined the effect of BIBF 1120, an inhibitor of vascular endothelial growth factor receptors (VEGFRs), platelet-derived growth factor receptors (PDGFRs) and fibroblast growth factor receptors (FGFRs) tyrosine kinases, on the reversal of multidrug resistance in vitro. METHODS: The doxorubicin and rhodamine 123 retention assay was performed by flowcytometry. Western blot were employed to identify ABCB1 expression level and the effect of BIBF 1120 on the blockade of Akt and ERK1/2 phosphorylation. The expression of mdr1 mRNA was determined by RT-PCR analysis. The ATPase activity of ABCB1 was investigated using Pgp-Glo™ assay systems. RESULTS: BIBF 1120 significantly enhanced the cytotoxicity of doxorubicin and paclitaxel and increased the accumulation of ABCB1 substrates in ABCB1-overexpressing cancer cells, whereas it had no effect on the parental cells. On the other hand, BIBF 1120 did not alter the cytotoxicity of non-ABCB1 substrates and was unable to reverse ABCC1 or ABCG2-mediated MDR. Furthermore, BIBF 1120 inhibited the ATPase activity of ABCB1 in a concentration-dependent manner. However, no detectable alteration on the expression level of mdr1 mRNA or ABCB1 protein was identified in ABCB1-overexpressing cancer cells by different treatments of BIBF 1120. Interestly, total and phosphorylated forms of AKT and ERK1/2 were not inhibited by BIBF 1120 at the reversal concentrations. CONCLUSION: Our results suggest that BIBF 1120 is capable of overcoming ABCB1-mediated drug resistance by inhibiting ABCB1 function, which may have clinical significance for BIBF 1120 combinational treatment of certain resistant cancers.