Development and application of a fully automatic multi-function cassette module Mortenon M1 for radiopharmaceutical synthesis.

INTRODUCTION: Functions of existing automatic module systems for synthesis of radiopharmaceuticals mainly focus on the radiolabeling of small molecules. There are few modules which have achieved full-automatic radiolabeling of non-metallic and metallic nuclides on small molecules, peptides, and antibody drugs. This study aimed to develop and test a full-automatic multifunctional module system for the safe, stable, and efficient production of radiopharmaceuticals. METHODS: According to characteristics of labeling process of radioactive drugs, using UG and Solidworks softwares, full-automatic cassette-based synthesis module system Mortenon M1 for synthesis of radiopharmaceuticals with various radionuclides, was designed and tested. Mortenon M1 has at least three significant highlights: the cassettes are disposable, and there is no need of manual cleaning; the synthesis method program is flexible and can be edited freely by users according to special needs; this module system is suitable for radiolabeling of both small-molecule and macromolecular drugs, with potentially various radionuclides including 18F, 64Cu, 68Ga, 89Zr, 177Lu, etc. By program control methods for certain drugs, Mortenon M1 was used for radiolabeling of both small-molecule drugs such as [68Ga]-FAPI-46 and macromolecular drugs such as [89Zr]-TROP2 antibody. Quality control assays for product purity were performed with radio-iTLC and radio-HPLC, and the radiotracers were confirmed for application in microPET imaging in xenograft tumor-bearing mouse models. RESULTS: Functional tests for Mortenon M1 module system were conducted, with [68Ga]-FAPI-46 and [89Zr]-TROP2 antibody as goal synthetic products, and it displayed that with the cassette modules, the preset goals could be achieved successfully. The radiolabeling synthesis yield was good ([68Ga]-FAPI-46, 70.63% ± 2.85%, n = 10; [89Zr]-TROP2, 82.31% ± 3.92%, n = 10), and the radiochemical purity via radio-iTLC assay of the radiolabeled products was above 99% after purification. MicroPET imaging results showed that the radiolabeled tracers had reasonable radioactive distribution in MDA-MB-231 and SNU-620 xenograft tumor-bearing mice, and the tumor targeted radiouptake was satisfactory for diagnosis. CONCLUSION: This study demonstrated that the full-automatic module system Mortenon M1 is efficient for radiolabeling synthesis of both small-molecule and macromolecular substrates. It may be helpful to reduce radiation exposure for safety, provide qualified radiolabeled products and reliable PET diagnosis, and ensure stable production and supply of radiopharmaceuticals.

PD-L1 monoclonal antibody-conjugated nanoparticles enhance drug delivery level and chemotherapy efficacy in gastric cancer cells.

BACKGROUND: Docetaxel (DOC) is widely used as a chemotherapy drug for various tumors, including gastric cancer (GC), but the clinical application of DOC has been limited due to the hydrophobicity of the drug. We aimed to formulate a multifunctional nanoparticle (NP) system to reduce the side effects of the chemotherapy agent, to promote synergistic therapeutic effects, and to achieve targeted delivery of the therapy. METHODS: The polyethylene glycol-poly(ε-caprolactone) NPs (PEG-PCL NPs) were prepared by a ring opening copolymerization technique and were then conjugated with a programmed death-ligand 1 (PD-L1) monoclonal antibody (mAb). The effects of the surface coating on particle size, size distribution, zeta potential, drug encapsulation efficiency, loading capacity, and the drug release kinetics were investigated. By using a panel of PD-L1-expressing human GC cell lines and PD-L1-overexpressing cells, we studied cellular uptake, cytotoxic effects, and cellular apoptosis in the presence of PD-L1 mAb-conjugated NPs. RESULTS: The characterization of the structure and biological functions of DOC-PEG-PCL-mAb NPs was investigated in vitro. X-ray photoelectron spectroscopy validated the presence of the PD-L1 mAbs on the NP surface. The cellular uptake analysis showed that the antibody-conjugated NPs achieved significantly higher cellular uptake. The results of an in vitro cytotoxicity experiment on three GC lines further proved the targeting effects of the antibody conjugation. In addition, we found that the DOC-PEG-PCL-mAb NPs induced cell apoptosis and enhanced G2-M arrest in cancer cells, indicating the inhibition of microtubule synthesis. When compared with the control groups, DOC-PEG-PCL-mAb NPs are more effective in inhibiting PD-L1 expression in GC cells. CONCLUSION: Our results reported here highlight the biological and clinical potential of DOC-PEG-PCL-mAb NPs using PD-L1 mAbs in GC treatment.

Association between EGFR mutation and expression of BRCA1 and RAP80 in non-small cell lung cancer.

Association between the epithelial growth factor receptor (EGFR) mutation and the expression of breast cancer 1 (BRCA1) and receptor-associated protein 80 (RAP80) in non-small cell lung cancer (NSCLC) was studied. From September 2014 to September 2016, 51 patients with NSCLC who were hospitalized in Department of Thoracic Surgery in The Affiliated Jiangyin Hospital of Southeast University Medical College and underwent biopsy or operation were selected. The pathological changes of lung tissue were detected by hematoxylin and eosin histopathological staining. The fluorescent expression of BRCA1 and RAP80 protein in the two groups was determined by immunofluorescence staining. Reverse transcriptase polymerase chain reaction method and western blot analysis were used to detect BRCA1 and RAP80 mRNA and protein expression. Then the EGFR gene mutation was detected and analyzed. The results show that non-small cell lung cancer has an association with smoking. Compared with the control, the lung tissue structure of the NSCLC group was damaged. The protein fluorescence expression of BRCA1 and RAP80 in non-small cell lung cancer group was significantly increased. The expression of BRCA1 and RAP80 mRNA and protein in NSCLC group was significantly increased. The difference in expression of BRCA1 and RAP80 in the control and the non-small cell lung cancer group was statistically significant (P<0.05). EGFR gene mutations detected 14 of the 51 patients with genetic mutations. Non-small cell lung cancer and smoking have certain relevance, and BRCA1 and RAP80 expression in the development and progression of NSCLC has a close relationship. EGFR mutation in non-small cell lung cancer significantly related to the mutation of EGFR and BRCA1 and RAP80 gene expression plays an important role in the diagnosis and treatment of NSCLC.

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.

Hearing loss due to metastasis of gastric cancer to temporal bone: A case report.

Metastatic temporal bone tumors are rare, and tend to be asymptomatic. The clinical symptoms consist of aural discharge, bleeding, hearing loss and facial nerve paresis. The most common origin of the metastasis is breast cancer, and other sites of the primary tumor include the thyroid gland, brain, lungs, prostate and blood. Clinical reports of hearing loss due to gastric cancer metastatic to temporal bone are rare. In the present study, a case of gastric cancer metastasis to temporal bone without other organ involvement is described. The patient presented with the symptom of hearing loss, and the metastatic tumor was diagnosed by radiological imaging, including magnetic resonance imaging, computed tomography and bone scan.

Human Recombinant Endostatin Combined with Cisplatin Based Doublets in Treating Patients with Advanced NSCLC and Evaluation by CT Perfusion Imaging.

AIMS: To study the effectiveness of human recombinant endostatin injection (Endostar®) combined with cisplatin doublets in treating advanced non-small cell lung cancer (NSCLC), and to evaluate outcome by CT perfusion imaging. METHODS: From April 2011 to September 2014, 76 patients with advanced NSCLC who were treated with platinum-based doublets were divided into group A (36 patients) and group B (40 patients). Endostar® 15 mg/day was administered 4 days before chemotherapy and combined with chemotherapy from day 5 in group A, and combined with chemotherapy from the first day in Group B. Endostar® in the two groups was injected intravenously for 14 days. RESULTS: Treatment effectiveness in the two groups differed with statistical significance (p<0.05). Effectiveness evaluated by CT perfusion imaging, BF, BV, MTT and PS also demonstrated significant differences (all p<0.05). Adverse reactions in the two groups did not significantly vary (p>0.05). CONCLUSIONS: The response rate with Endostar® administered 4 days before chemotherapy and combined with chemotherapy from day 5 in group A was better than Endostar® combined with chemotherapy from the first day, and CT perfusion imaging could be a reasonable method for evaluation of patient outcomes.

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.

Multi-responsive and logic controlled release of DNA-gated mesoporous silica vehicles functionalized with intercalators for multiple delivery.

Novel DNA-gated mesoporous silica nanoparticle (MSN) vehicles functionalized with disulfide-linked acridinamine intercalators are constructed for multi-responsive controlled release. The DNA-gated MSN vehicles release cargo encapsulated in the MSN pores under different stimuli, including disulfide reducing agents, elevated temperature, and deoxyribonuclease I (DNase I), for codelivery of drugs and DNA/genes in different forms. Furthermore, the cascade release of encapsulated and intercalative drugs is controlled by AND logic gates in combination of dual stimuli. The ingeniously designed DNA-gated MSN vehicles integrates multiple responses and AND logic gate operations into a single smart nanodevice not only for codelivery of drugs and DNA/genes but also for cascade release of two drugs and has promising biological applications to meet diverse requirements of controlled release.

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.

Facile preparation of paclitaxel loaded silk fibroin nanoparticles for enhanced antitumor efficacy by locoregional drug delivery.

Non-toxic, safe materials and preparation methods are among the most important factors when designing nanoparticles (NPs) for future clinical application. Here we report a novel and facile method encapsulating anticancer drug paclitaxel (PTX) into silk fibroin (SF), a biocompatible and biodegradable natural polymer, without adding any toxic organic solvents, surfactants or other toxic agents. The paclitaxel loaded silk fibroin nanoparticles (PTX-SF-NPs) with a diameter of 130 nm were formed in an aqueous solution at room temperature by self-assembling of SF protein, which demonstrated mainly silk I conformation in the NPs. In cellular uptake experiments, coumarin-6 loaded SF NPs were taken up efficiently by two human gastric cancer cell lines BGC-823 and SGC-7901. In vitro cytotoxicity studies demonstrated that PTX kept its pharmacological activity when incorporating into PTX-SF-NPs, while SF showed no cytotoxicity to cells. The in vivo antitumor effects of PTX-SF-NPs were evaluated on gastric cancer nude mice exnograft model. We found that locoregional delivery of PTX-SF-NPs demonstrated superior antitumor efficacy by delaying tumor growth and reducing tumor weights compared with systemic administration. Furthermore, the organs of mice in NP treated groups didn't show obvious toxicity, indicating the in vivo safety of SF NPs. These results suggest that SF NPs are promising drug delivery carriers, and locoregional delivery of SF NPs could be a potential future clinical cancer treatment regimen.