Chemoimmunotherapy by combining oxaliplatin with immune checkpoint blockades reduced tumor burden in colorectal cancer animal model.

BACKGROUND: Colorectal cancer (CRC) is among one of the top common cancers worldwide. Developing novel comprehensive treatment strategies is critical for improving survival of late stage CRC patients. Recent advances in immune checkpoint blockades provided a novel strategy for treating cancers via stimulating the antitumor immune response. However, the effects of immune checkpoint blockades were limited in CRC due to intrinsic resistance. Oxaliplatin (OXA) based chemotherapy was the foundation of CRC adjuvant chemotherapy. Here, we investigated the potential roles of OXA in inducing immunogenicity and synergizing with immune checkpoints in CRC. METHOD: Immunogenicity of OXA was tested in CRC cell lines. Immune checkpoint blockades sensitive and resistant CRC models were used to study the potential synergistic roles of OXA with immune checkpoint blockades. RESULTS: We found CT26 mouse model was sensitive to immune checkpoint blockades, while MC38 mouse model was resistant. OXA could induce immunogenic cell death in several human and mouse CRC cell lines. Short term OXA treatment increased immune cell infiltration in MC38 mouse model and therefore enhanced the efficacy of immune checkpoint in MC38 mouse model. As a response to the OXA and immune checkpoint blockades combination, inhibitory immune checkpoints were down-regulated in MC38 tumors, while immune enhancing cytokines were up-regulated. Short term OXA treatment induced antitumor immune response in an immune checkpoint blockades resistant mouse model, therefore synergized with immune checkpoint blockades.

DNA-PKcs interference sensitizes colorectal cancer cells to a mTOR kinase inhibitor WAY-600.

Colorectal cancer (CRC) is one leading contributor of cancer-related mortalities. Mammalian target of rapamycin (mTOR), existing in two complexes (mTORC1/2), is a valuable target for possible CRC interference. In the current study, we showed that WAY-600, a potent mTOR inhibitor, only exerted moderate activity against primary and HT-29 CRC cells. We proposed that DNA-dependent protein kinase catalytic subunit (DNA-PKcs) could be the major resistance factor of WAY-600 in CRC cells. DNA-PKcs inhibitors, including NU7026 and NU7441, dramatically enhanced WAY-600-induced cytotoxic and pro-apoptotic effect against the CRC cells. Further, WAY-600-exerted cytotoxicity was significantly increased in DNA-PKcs-silenced (by targeted siRNA/shRNA) CRC cells, but was attenuated with DNA-PKcs overexpression. Our evidence suggested that DNA-PKcs Thr-2609 phosphorylation might be critical for WAY-600's resistance. Mutation of this site through introducing a dominant negative DNA-PKcs (T2609A) dramatically potentiated WAY-600's sensitivity in HT-29 cells. Meanwhile, overexpression of protein phosphatase 5 (PP5) dephosphorylated DNA-PKcs at Thr-2609, and significantly increased WAY-600's sensitivity in HT-29 cells. In vivo, WAY-600-induced anti-HT-29 xenograft growth activity was significantly potentiated with NU7026 co-administration. These results suggest that DNA-PKcs could be the major resistance factor of WAY-600 in CRC cells.

CircCCT3 Acts as a Sponge of miR-613 to Promote Tumor Growth of Pancreatic Cancer Through Regulating VEGFA/VEGFR2 Signaling.

BACKGROUND: Circular RNAs (CircRNAs) have been recently implicated in the progression of pancreatic cancer (PC). AIMS: To investigate the involvement of CircCCT3 in PC and studying its interactions and functioning during the progression of PC in vitro and in vivo, using methods of molecular biology and bioinformatics. STUDY DESIGN: Experimental study. METHODS: The expressions of CircCCT3 and miR-613 in pancreatic carcinoma tissues and cell lines were evaluated by quantitative real-time polymerase chain reaction (PCR). The relationship between clinical pathologic features as well as the survival rate and CircCCT3 expression was analyzed with chi-square test and the Kaplan-Meier method. CCK-8, wound healing, transwell assays, and the fluorescein isothiocyanate- AnnexinV/propidium iodide (FITC-AnnexinV/PI) assay were used to assess cell proliferation, migration, invasion, and apoptosis after CircCCT3 overexpression or downregulation. The Dual- Luciferase reporter assay, RNA immunoprecipitation (RIP), RNA pull-down and fluorescence in situ hybridization (FISH) assays were performed to validate the potential interaction of CircCCT3, miR-613, and vascular endothelial growth factor (VEGFA). The nude mouse xenograft tumor assay was used to detect CircCCT3 effects on pancreatic tumorigenesis in vivo. Western blotting analysis was performed to examine the VEGFA and the vascular endothelial growth factor receptor 2 (VEGFR2) protein expressions following. RESULTS: CircCCT3 expression was significantly increased in PC tissues (3.41 ± 0.57 vs. 1.00 ± 0.10, P < .01) and cell lines (Patu8988 2.57 ± 0.20; SW1990 2.88 ± 0.10; BxPC-3 2.45 ± 0.20; Panc02 2.99 ± 0.10 vs. H6c7 1.00 ± 0.10; all P < .001). CircCCT3 expression was negatively correlated with miR-613 expression. PC patients with high CircCCT3 expression exhibited significantly poorer overall survival rate than those patients with low CircCCT3 expression (P = .013). Moreover, it was found that CircCCT3 promoted cell proliferation, migration, and invasion, and inhibited cell apoptosis in PC cells. The CircCCT3 acted as a sponge for the miR-613 to facilitate VEGFA and VEGFR2 expression. si-CirCCT3 also inhibited tumor growth of PC in nude mice. si-CircCCT3 reduced VEGFA and VEGFR2 expression, whereas overexpression of CircCCT3 increased VEGFA and VEGFR2 expression. CONCLUSION: Increased CircCCT3 suggests a poor prognosis for PC patients and promotes the migration and invasion through targeting VEGFA/VEGFR2 signaling. CircCCT3 may serve as a potential and promising therapeutic target for PC treatment.

Serum Fractalkine Levels Were Positively Associated with Disease Severity of Liver Cirrhosis.

OBJECTIVE: The chemokine receptor CX3CR1 and its specific ligand fractalkine (CX3CL1, FKN) has been implicated in modulating inflammatory and fibroproliferative diseases. The current study was performed to investigate the correlation of serum fractalkine levels with disease severity of liver fibrosis/cirrhosis (LC). METHODS: 162 LC patients and 140 healthy controls well enrolled in our study. Serum fractalkine levels were detected using commercial ELISA kit. Liver biopsy specimens were obtained using 16 G disposable needle in LC patients. The Child-Pugh grade was recorded to assess liver function. ROC curve analysis was performed to assess the potential diagnostic power of serum fractalkine with regard to the disease severity of Child-Pugh grade system. Pathological assessment of cirrhotic severity was performed by Laennec staging system. The L3 skeletal muscle index (L3SMI) was applicated to assess the nutrition status. RESULTS: Serum fractalkine levels were significantly higher in LC patients compared with healthy controls. The case group included 50 Child-Pugh A patients, 59 Child-Pugh B patients, and 53 Child-Pugh C patients. Cirrhosis patients with Child-Pugh C had drastically higher serum fractalkine levels compared with those with Child-Pugh B and A. Child-Pugh B patients showed significantly higher serum PACAP concentrations compared with those with Child-Pugh A. ROC curve analysis demonstrated that serum fractalkine may act as a potential indicator for disease progression of LC determined by Child-Pugh classification. Besides, serum fractalkine levels were positively related to ALT and AST concentrations and negatively related to L3SMI. CONCLUSION: Serum fractalkine levels were positively associated with disease severity of LC.

Hsa_circ_0038646 promotes cell proliferation and migration in colorectal cancer via miR-331-3p/GRIK3.

Increasing evidence supports the essential roles of circular RNAs (circRNAs) and microRNAs (miRNAs/miRs) in different types of human cancer. For example, hsa_circ_0137008 functions as a sponge for mi-338-5p and inhibits the malignant phenotype in colorectal cancer. Furthermore, hsa_circ_RNA_0011780 downregulates FBXW7 by targeting miR-554a and suppressing the progression of non-small cell lung cancer. Thus far, only a single report has identified that the miRNA miR-331-3p exerts a pivotal effect on human colorectal cancer (CRC) evolution. However, both the up- and downstream regulatory mechanisms of miR-331-3p are unclear. In the present study, it was predicted via bioinformatics analysis that the circRNA, hsa_circ_0038646, and the glutamate receptor ionotropic kainate 3 (GRIK3) gene contain binding sites that can interact with miR-331-3p. Thus, hsa_circ_0038646/miR-331-3p/GRIK3 may be a novel therapeutic pathway for CRC. Reverse transcription-quantitative PCR and western blotting analyses were performed, as well as cell proliferation, luciferase reporter and Transwell migration assays. Hsa_circ_0038646 was overexpressed in both CRC cells and tissues, and this aberrant expression was positively related with increasing tumor grade. Knockdown of hsa_circ_0038646 significantly weakened human CRC cell proliferation and migration. It was shown that hsa_circ_0038646 can sponge miR-331-3p to suppress its expression, and that suppression of miR-331-3p can reverse the effects of hsa_circ_0038646 inhibition in CRC cells. It was determined that GRIK3 is a downstream target of miR-331-3p, and that hsa_circ_0038646 could increase the levels of GRIK3 by suppressing miR-331-3p in CRC cells. Restoring GRIK3 expression rescued the weakened CRC cell proliferation and migration following hsa_circ_0038646 knockdown. The present study indicated that hsa_circ_0038646 functions as a tumor promoter in CRC by increasing GRIK3 expression via sponging of miR-331-3p. The hsa_circ_0038646/miR-331-3p/GRIK3 axis may be a novel therapeutic and diagnostic target of CRC.

Protective effect of YHI and HHI-I against experimental acute pancreatitis in rabbits.

AIM:To observe the protective effect of combined i.v. administraction of Yuanhu injection (YHI) and Huoxuehuayu injection-I (HHI-I) against acute pancreatitis (AP) in rabbits.METHODS:Sever acute pancreatitis (SAP) was induced by retrograde infusion of artificial bile juice into biliary-pancreatic duct, and treated with YHI and HHI-I intravenously. The protective effect was judged by the survival time and rate, serum amylase, serum interleukin-6, pancreatic microcirculation and pathological alteration.RESULTS:Combined use of YHI and HHI-I could markedly increase the rabbits' 5-day survival rate after AP (83.3% in the treatment group and 33.3% in control). The serum amylase value (x-± s) decreased to 1596.6U/L± 760.50U/L in the 5th day from the high level (6320.83U/L± 2614.12U/L) in the 1st day after AP in the treatment group, while in the control group the amylase activity in the 5th day was 2095.0U/L± 1081.87U/L, being significantly different from that before AP (837.17U/L± 189.12U/L). YHI and HHI-I also obviously improved the pancreatic microcirculation and lowered the serum interleukin-6 level, one of the indices of severe pancreatitis. Pathological examination indicated all the changes typical for AP in YHI and HHI-I treatment group were milder than those in the control.CONCLUSION:YHI and HHI-I used in combination might have protective effect against acute pancreatitis in rabbits.