Enhanced antiproliferative activity of antibody-functionalized polymeric nanoparticles for targeted delivery of anti-miR-21 to HER2 positive gastric cancer.

MiR-21 is an oncogenic miR frequently elevated in gastric cancer. Overexpression of miR-21 decreases the sensitivity of gastric cancer cells to trastuzumab, which is a humanized monoclonal antibody targeting human epidermal growth factor receptor 2. However, optimization of miRNA or its anti-miRNA oligonucleotides (AMOs) for delivery is a challenge. Receptor-mediated endocytosis plays a crucial role in the delivery of biotherapeutics including AMOs. This study is a continuation of our earlier findings involving poly(ε-caprolactone) (PCL)-poly (ethylene glycol) (PEG) nanoparticles (PEG-PCL NPs), which were coated with trastuzumab to target gastric cancer cells with HER2 receptor over-expression using anti-miRNA-21 antisense oligonucleotides (AMO-21). The antibody conjugates (HER-PEG-PCL NPs) act against target cells via antibody-dependent mechanisms and also based on encapsutalated AMO-21. X-ray photoelectron spectroscopy validated the presence of trastuzumab on NP surface. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) revealed a stable antibody expression. The cell line specificity, cellular uptake, AMO-21 delivery, and cytotoxicity of the HER-PEG-PCL NPs were investigated. We found that the antibody conjugates significantly enhanced the cellular uptake of NPs. The HER-PEG-PCL NPs effectively suppressed the target miRNA expression in gastric cancer cells, which further up-regulated phosphatase and tensin homolog (PTEN). As a result, the sensitivity of HER2-expressing gastric cancer cells to trastuzumab was enhanced. The approach enhances the targeting by trastuzumab as well as antibody-dependent cellular cytotoxicity of immune effector cells. The antitumor effects of AMO-21-HER-PEG-PCL NPs were compared with trastuzumab in xenograft gastric cancer mice. The results provide insight into the biological and clinical potential of targeted AMO-21 delivery using modified trastuzumab for gastric cancer treatment.

Phase I clinical study of personalized peptide vaccination combined with radiotherapy for advanced hepatocellular carcinoma.

AIM: To assess the efficacy and safety of a new treatment modality, cellular immune therapy based on personalized peptide vaccination (PPV-DC-CTL) combined with radiotherapy, for treating advanced hepatocellular carcinoma (HCC). METHODS: A total of nine patients with advanced HCC were enrolled. Multidisciplinary consultation confirmed that all the patients definitely had no opportunity of surgery, because four patients had multiple liver metastases (the number of liver lesions > 3), one patient had liver metastases and portal vein tumor thrombosis, one patient had lung and bone metastases, two patients had liver and lung metastases and one patient had liver metastasis and peritoneal metastasis. Patients with metastasis were treated with precise radiotherapy combined with PPV-DC-CTL. RESULTS: Following radiotherapy and one to three cycles of PPV-DC-CTL treatment, AFP levels were significantly decreased in six patients and imaging assessment of the lesions showed a partial response (PR) in three patients and stable disease in the other three patients. The response rate was 33% and disease control rate was 66%. This regimen was found to be safe and well tolerated. None of the patients developed liver or kidney side effects. Only one patient developed grade II bone marrow suppression and the remaining patients had no significant hematological side effects. CONCLUSION: Radiotherapy combined with PPV-DC-CTL provides a new therapeutic strategy for patients with advanced HCC, which is well tolerated, safe, feasible and effective.

Efficacy of decitabine-loaded gelatinases-stimuli nanoparticles in overcoming cancer drug resistance is mediated via its enhanced demethylating activity to transcription factor AP-2 epsilon.

Hypermethylation of the transcription factor AP-2 epsilon (TFAP2E) gene affects 5-fluorouridine (5-FU) resistance in gastric cancer (GC) patients. The epigenetic inhibitor 5-Aza-2'-deoxycytidine (DAC), which reverses DNA methylation by targeting DNA methyltransferases (DNMTs), has potential to sensitize GC to 5-FU. Nevertheless, DNA demethylation only DAC transiently occurs since DAC is unstable in aqueous solutions, which limits its potential. Here we developed intelligent nanoparticles (NPs) comprising gelatinase with polyethylene glycol (PEG) and poly-ε-caprolactone) (PCL) to specifically deliver DAC (DAC-TNPs) to tumors. DAC-carrying PEG-PCL NPs (DAC-NPs) lacking gelatinase features served as controls. 72 hours after administration of DAC-TNPs or DAC-NPs, 5-FU was sequentially applied to GC cells and human GC xenografts in nude mice. Both in vitro and in vivo evaluations demonstrated that the combination treatment of DAC-TNPs and 5-FU greatly improved tumor suppression in GC cells and mouse xenograft models with hypermethylation TFAP2E (MKN45 cells). We thus propose that the sequential administration of DAC-TNPs and 5-FU could be significant in the development of novel targeted therapies.

Gelatinases-stimuli nanoparticles encapsulating 5-fluorouridine and 5-aza-2'-deoxycytidine enhance the sensitivity of gastric cancer cells to chemical therapeutics.

Aberrant methylation of the transcription factor AP-2 epsilon (TFAP2E) has been attributed to 5-fluorouridine (5-FU) sensitivity. 5-Aza-2'-deoxycytidine (DAC), an epigenetic drug that inhibits DNA methylation, is able to cause reactive expression of TFAP2E by demethylating activity. This property might be useful in enhancing the sensitivity of cancer cells to 5-FU. However, the effect of DAC is transient because of its instability. Here, we report the use of intelligent gelatinases-stimuli nanoparticles (NPs) to coencapsulate and deliver DAC and 5-FU to gastric cancer (GC) cells. The results showed that NPs encapsulating DAC, 5-FU, or both could be effectively internalized by GC cells. Furthermore, we found that the NPs enhanced the stability of DAC, resulting in improved re-expression of TFAP2E. Thus, the incorporation of DAC into NPs significantly enhanced the sensitivity of GC cells to 5-FU by inhibiting cell growth rate and inducing cell apoptosis. In conclusion, the results of this study clearly demonstrated that the gelatinases-stimuli NPs are an efficient means to simultaneously deliver epigenetic and chemotherapeutic drugs that may effectively inhibit cancer cell proliferation.

Enhancement of radiotherapy efficacy by miR-200c-loaded gelatinase-stimuli PEG-Pep-PCL nanoparticles in gastric cancer cells.

Radiotherapy is the main locoregional control modality for many types of unresectable tumors, including gastric cancer. However, many patients fail radiotherapy due to intrinsic radioresistance of cancer cells, which has been found to be strongly associated with cancer stem cell (CSC)-like properties. In this study, we developed a nanoparticle formulation to deliver miR-200c, which is reported to inhibit CSC-like properties, and then evaluated its potential activity as a radiosensitizer. miR-200c nanoparticles significantly augmented radiosensitivity in three gastric cancer cell lines (sensitization enhancement ratio 1.13-1.25), but only slightly in GES-1 cells (1.06). In addition to radioenhancement, miR-200c nanoparticles reduced the expression of CD44, a putative CSC marker, and the percentage of CD44(+) BGC823 cells. Meanwhile, other CSC-like properties, including invasiveness and resistance to apoptosis, could be suppressed by miR-200c nanoparticles. CSC-associated radioresistance mechanisms, involving reactive oxygen species levels and DNA repair capacity, were also attenuated. We have demonstrated that miR-200c nanoparticles are an effective radiosensitizer in gastric cancer cells and induce little radiosensitization in normal cells, which suggests that they are as a promising candidate for further preclinical and clinical evaluation.

Enhancement of radiotherapy efficacy by docetaxel-loaded gelatinase-stimuli PEG-Pep-PCL nanoparticles in gastric cancer.

Docetaxel (DOC) is widely used as radiosensitizer in various tumors, including gastric cancer (GC), but its therapeutic effect remains to be improved. In this study, using docetaxel-loaded nanoparticles (DOC-NPs) based on gelatinase-stimuli strategy, we compared their radioenhancement efficacy with docetaxel in GC. Compared with DOC, radiosensitization of DOC-NPs was improved significantly (sensitization enhancement ratio increased 1.09-fold to 1.24-fold, P<0.01) in all three gelatinase overexpressing GC cells, while increased slightly (1.02-fold, P=0.38) in gelatinase deficient normal gastric mucosa cells. The improved radiosensitization efficacy was associated with enhanced G2/M arrest, increased reactive oxygen species (ROS), more effective DSBs and promoted apoptosis. More importantly, the radiosensitization efficacy of DOC-NPs (estimated as ''very active'') was more prominent than DOC (estimated as ''moderately active'') by intravenous injection in xenograft. In conclusion, DOC-NPs are highly selective radiosensitizers in gelatinase over-expressing tumors, and more effective than DOC. By manipulating the common microenvironment difference between tumor and normal tissue, gelatinase-mediated nanoscale delivery system serves as a potential strategy possessing both universality and selectivity for radiosensitizers.

Targeted delivery of miR-200c/DOC to inhibit cancer stem cells and cancer cells by the gelatinases-stimuli nanoparticles.

Cancer stem cells (CSCs) are recently discovered as vital obstacles for the successful cancer therapy. Emerging evidences suggest that miR-200c functions as an effective CSCs inhibitor and can restore sensitivity to microtubule-targeting drugs. In the present work, the intelligent gelatinases-stimuli nanoparticles (NPs) was set up to co-deliver miR-200c and docetaxel (DOC) to verify their synergetic effects on inhibition of CSCs and non-CSC cancer cells. After tumor cells were treated with miR-200c NPs, miR-200c and its targeted gene class III beta-tubulin (TUBB3)TUBB3 expression were evaluated. The effects of miR-200c/DOC NPs on tumor cell viability, migration and invasion as well as the expression of E-cadherin and CD44 were studied. The antitumor effects of miR-200c/DOC NPs were compared with DOC NPs in xenograft gastric cancer mice. Moreover, the residual tumors after treatment were subcutaneously seeded into nude mice to further investigate the effective maintenance of NPs. We found that the gelatinases-stimuli NPs facilitated miR-200c into cells, achieving sustained miR-200c expression in tumor cells during 9 days. The miR-200c/DOC NPs significantly enhanced cytotoxicity of DOC, possibly by decreasing TUBB3 level, and reversing EMT. The miR-200c NPs achieved high levels of in vivo accumulation and long retention in gastric cancer xenografts after intravenous administration. The miR-200c/DOC NPs prominently suppressed in vivo tumor growth with elevated miR-200c and E-cadherin levels and down-regulated TUBB3 and CD44 expressions. When the residual tumors after miR-200c/DOC NPs treatment were re-transplanted into nude mice, the tumors demonstrated the slowest growth speed. The miR-200c/DOC NPs may provide a promising modality for co-delivery of nucleic acid and drugs to simultaneously inhibit CSCs and non-CSC cancer cells.

Gelatinase-stimuli strategy enhances the tumor delivery and therapeutic efficacy of docetaxel-loaded poly(ethylene glycol)-poly(ɛ-caprolactone) nanoparticles.

Nanoscale drug carriers have been extensively developed to improve drug therapeutic efficiency. However, delivery of chemotherapeutic agents to tumor tissues and cells has not been favorably managed. In this study, we developed a novel "intelligent" nanoparticle, consisting of a gelatinase-cleavage peptide with poly(ethylene glycol) (PEG) and poly(ɛ-caprolactone) (PCL)-based structure for tumor-targeted docetaxel delivery (DOC-TNPs). The docetaxel-loaded PEG-PCL nanoparticles (DOC-NPs) that did not display gelatinase-stimuli behaviors were used as a control. We found clear evidence that the DOC-TNPs were transformed by gelatinases, allowing drug release and enhancing the cellular uptake of DOC (P < 0.01). In vivo biodistribution study demonstrated that targeted DOC-TNPs could accumulate and remain in the tumor regions, whereas non-targeted DOC-NPs rapidly eliminated from the tumor tissues. DOC-TNPs exhibited higher tumor growth suppression than commercialized Taxotere(®) (docetaxel; Jiangsu Hengrui Medicine Company, Jiangsu, China) and DOC-NPs on hepatic H22 tumor model via intravenous administration (P < 0.01). Both in vitro and in vivo experiments suggest that the gelatinase-mediated nanoscale delivery system is promising for improvement of antitumor efficacy in various overexpressed gelatinase cancers.

The E-cadherin (CDH1)--160 C/A polymorphism and prostate cancer risk: a meta-analysis.

Published data on the association between E-cadherin (CDH1) -160 C/A polymorphism and prostate cancer (PCA) risk are inconclusive. To derive a more precise estimation of the relationship, a meta-analysis was performed. A logistic regression approach proposed for molecular association studies was used to estimate a biological model of the gene effect. A total of 11 studies including 2637 cases and 2673 controls were involved in this meta-analysis. Logistic regression analysis indicated that the CDH1 -160 C/A genotypes were associated with PCA risk. The genetic model test indicated that the genetic model was most likely to be dominant (CA+AA vs CC). Overall, meta-analysis indicated that the -160A allele carriers (CA+AA) had a 21% elevated risk of PCA, when compared with the homozygotes (CC) (odds ratio (OR)=1.21; 95% confidence interval (CI): 0.97-1.51; P=0.090, P(heterogeneity)=0.001). In the subgroup analyses by ethnicity, significantly elevated risks were associated with -160 variant genotypes (CA+AA) in both European and Asian populations (OR=1.24; 95% CI: 1.08-1.43; P=0.003, P(heterogeneity)=0.220 and OR=1.54; 95% CI: 1.23-1.93; P<0.001, P(heterogeneity)=0.200). However, no significant associations were found in Africans (OR=0.59; 95% CI: 0.32-1.09; P=0.090, P(heterogeneity)=0.070). Although some modest bias could not be eliminated, this meta-analysis suggests that the CDH1 -160A allele is a low-penetrant risk factor for developing PCA, especially in Europeans and Asians.

Predictive value of thymidylate synthase expression in advanced colorectal cancer patients receiving fluoropyrimidine-based chemotherapy: evidence from 24 studies.

The published data about thymidylate synthase (TS) expression and its predictive value in advanced colorectal cancer (CRC) patients receiving fluoropyrimidine-based chemotherapy seemed inconclusive. To derive a more precise estimation of the relationship, a metaanalysis was performed. Studies have been identified by searching PubMed and Embase. Inclusion criteria were advanced CRC patients, received fluoropyrimidine-based chemotherapy and evaluation of TS expression and overall response rate (ORR). The relative ratio (RR) for ORR in patients with low-TS expression over those with high-TS expression with 95% confidence interval (CI) was calculated for each study as an estimation of the predictive effect of TS. A total of 24 studies including 1,112 patients were involved in this metaanalysis. The overall RR was 2.20 (95% CI, 1.82-2.66; p = 0.000). For studies evaluating TS expression in metastatic lesions, the pooled RR was 3.23 (95% CI, 2.27-4.59; p = 0.000); for studies testing TS expression in primary lesions, a pooled RR of 1.89 (95% CI, 1.45-2.48; p = 0.000) was estimated. Focusing the analysis on immunohistochemistry (IHC)-based or RTPCR-based assessments, the pooled RR was 1.83 (95% CI, 1.44-2.34; p = 0.000) and 2.96 (95% CI, 2.07-4.22; p = 0.000), respectively. The results indicated that low-TS expression tumors in advanced CRC patients were more sensitive to fluoropyrimidine-based chemotherapy. Subgroup analyses indicated that the predictive value of TS expression evaluated in metastases was more prominent than that of primary lesions, and that TS expression tested by RTPCR was also of greater predictive value than by IHC.