melitten and Liver-Neoplasms

Apitherapy (using bee products) has gained broad recognition in cancer therapeutics globally. Honeybee venom has a broad range of biological potential, and its utilization is rapidly emerging in apitherapy. Bee products have significant potential to strengthen the immune system and improve human health. Thus, this review is targeted toward recapitulating the chemo-preventive potential of melittin (MEL), which constitutes a substantial portion of honeybee venom. Honeybee venom (apitoxin) is produced in the venom gland of the honeybee abdomen, and adult bees utilize it as a primary colony defense mechanism. Apitoxin comprises numerous biologically active compounds, including peptides, enzymes, amines, amino acids, phospholipids, minerals, carbohydrates, and volatile components. We are mainly focused on exploring the potential of melittin (a peptide component) of bee venom that has shown promising potential in the treatment of several human cancers, including breast, stomach, lung, prostate, ovary, kidney, colon, gastric, esophageal, cervical cancers, melanoma, osteosarcoma, and hepatocellular carcinoma. This review has summarized all potential studies related to the anticancerous efficacy of melittin (apitoxin), its formulations, conjugates, and nano-formulations against several human carcinomas, which would further pave the way for future researchers in developing potent drugs for cancer management.

Topics: Animals; Bee Venoms; Bees; Bone Neoplasms; Carcinoma, Hepatocellular; Humans; Liver Neoplasms; Male; Melitten; Peptides

Melittin (MEL) has been reported to exhibit anti-cancer effects in vitro against several types of cancer. Long non-coding RNA (lncRNA) ADAMTS9-AS2 can be used as a tumor suppressor. However, there is insufficient data on the potential link between MEL and ADAMTS9-AS2 in hepatocellular carcinoma (HCC).. RT-qPCR, CCK-8, colony formation, scratch wound healing and transwell assays were used to detect the function of MEL or ADAMTS9-AS2 on HCC cells. Furthermore, Western blot analysis was applied to determine that whether an association existed in MEL or ADAMTS9-AS2 with the PI3K/AKT/mTOR signal pathway. In addition, RT-qPCR and Western blot analysis validated that whether MEL has a demethylation effect.. All the experimental data showed that MEL or ADAMTS9-AS2 inhibited the proliferation, migration and invasion of MHCC97-H and HepG2 cells, which may relate to PI3K/AKT/mTOR signal pathway. Moreover, the result showed that MEL treatment inhibited the expression of DNA methyltransferase protein-1 (DNMT1), which acted as the role of demethylation, and then up-regulated the expression of ADAMTS9-AS2, affecting the development of HCC.. ADAMTS9-AS2 played a role in MEL-induced HCC inhibition. This study provided an interesting theoretical basis and further evidence for the potential application of MEL in the treatment of HCC.

Topics: ADAMTS9 Protein; Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Movement; Cell Proliferation; Demethylation; Humans; Liver Neoplasms; Melitten; MicroRNAs; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; TOR Serine-Threonine Kinases

Chemoimmunotherapy has shown great potential to activate an immune response, but the immunosuppressive microenvironment associated with T cell exhaustion remains a challenge in cancer therapy. The proper immune-modulatory strategy to provoke a robust immune response is to simultaneously regulate T-cell exhaustion and infiltration. Here, a new kind of carrier-free nanoparticle is developed to simultaneously deliver chemotherapeutic drug (doxorubicin, DOX), cytolytic peptide (melittin, MPI), and anti-TOX small interfering RNA (thymocyte selection-associated high mobility group box protein, TOX) using a fluorinated prodrug strategy. In this way, the enhanced immunogenic cell death (ICD) induced by the combination of DOX and MPI can act as "offense" signaling to increase CD8

Topics: Cell Line, Tumor; Doxorubicin; Humans; Immunotherapy; Liver Neoplasms; Melitten; Neoplasms; Prodrugs; RNA, Small Interfering; Tumor Microenvironment

Liver sinusoidal endothelial cells (LSECs) are responsible for the immunologic tolerance of liver which is a common site for visceral metastases, suggesting its potential role as an target for cancer immunotherapy. However, targeted modulation of LSECs is still not achieved thus far. Here, we report LSECs are specifically targeted and modulated by melittin nanoparticles (α-melittin-NPs). Intravital imaging shows that LSECs fluoresce within 20 s after intravenous injection of α-melittin-NPs. α-melittin-NPs trigger the activation of LSECs and lead to dramatic changes of cytokine/chemokine milieu in the liver, which switches the hepatic immunologic environment to the activated state. As a result, α-melittin-NPs resist the formation of metastatic lesions with high efficiency. More strikingly, the survival rate reaches 80% in the spontaneous liver metastatic tumor model. Our research provides support for the use of α-melittin-NPs to break LSEC-mediated immunologic tolerance, which opens an avenue to control liver metastasis through the immunomodulation of LSECs.

Topics: Animals; Computational Biology; Female; Flow Cytometry; Hepatocytes; Immune Tolerance; Liver; Liver Neoplasms; Melitten; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Nanoparticles; Reverse Transcriptase Polymerase Chain Reaction

Chemotherapy resistance represents a major obstacle in the treatment of patients with hepatocellular carcinoma (HCC). The purpose of this study was to investigate the anti-cancer effect of MEL-pep, a novel analog of the natural antibacterial peptide melittin (MEL), on human 5-fluorouracil-resistant HCC cells (BEL-7402/5-FU) and to clarify the molecular mechanisms involved in these effects. We found that MEL-pep inhibited the proliferation of BEL-7402/5-FU cells and reversed 5-FU resistance in vitro. MEL-pep directly bound to BEL-7402/5-FU cells and disrupted the cell membrane. P-glycoprotein (P-gp) plays an important role in the development of resistance to anticancer drugs. We found that MEL-pep inhibited P-gp expression and increased the intracellular accumulation of the P-gp substrate rhodamine-123 in BEL-7402/5-FU cells. Additionally, the phosphorylation of Akt and NF-κB/p65 nuclear translocation was all inhibited by MEL-pep. Insulin - like growth factor I, a phosphatidylinositol 3 kinase(PI3K) /protein kinase B(AKT) agonist, reversed MEL-pep induced P-gp suppression. Therefore, MEL-pep inhibited P-gp expression by deactivating the PI3K/Akt signaling pathway. Finally, in a BEL-7402/5-FU cell-derived xenograft tumor model in mice, we found that the intratumoral administration of MEL-pep inhibited tumor growth in a dose-dependent manner. Thus, MEL-pep could be a promising candidate in the treatment of chemotherapy resistant HCC.

Topics: Animals; Antimetabolites, Antineoplastic; ATP Binding Cassette Transporter, Subfamily B; Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Membrane; Cell Membrane Permeability; Cell Proliferation; Cell Survival; Drug Resistance, Neoplasm; Fluorouracil; Gene Expression Regulation, Neoplastic; Humans; Inhibitory Concentration 50; Insulin-Like Growth Factor I; Liver Neoplasms; Melitten; Mesenchymal Stem Cells; Mice; Mice, Nude; Phosphatidylinositol 3-Kinases; Primary Cell Culture; Proto-Oncogene Proteins c-akt; Rats; Rats, Sprague-Dawley; Transcription Factor RelA; Tumor Burden; Xenograft Model Antitumor Assays

Melittin is the main effective component of bee venom and has extensive biological functions; however, serious side effects have restricted its clinical application. Preclinical and clinical studies showed that the main adverse events were allergic reaction and pain at the administration site. To decrease the toxicity, we prepared melittin nano-liposomes by encapsulating melittin with poloxamer 188 and explored the inhibitory activities on liver cancer together with biological safety. Here, we showed that melittin nano-liposomes significantly inhibited the survival of hepatocellular carcinoma (HCC) cells in vitro and prominently suppressed the growth of subcutaneous and orthotopic HCC transplantation tumors in vivo. It was important that it induced less inflammation and allergy in mice compared with melittin. Overall, melittin nano-liposomes would have a better application in HCC therapy due to its significant anti-tumor activity and better biological safety.

Topics: Animals; Bees; Capsules; Carcinoma, Hepatocellular; Heterografts; Humans; Hypersensitivity; Inflammation; Liposomes; Liver Neoplasms; Melitten; Mice; Nanoparticles; Poloxamer

Hepatocarcinoma is one of the malignant cancers with significant morbidity and mortality. Immunotherapy has emerged in clinical treatment, owing to the limitation and severe side effects of chemotherapy. In the immune system, natural killer (NK) cells are important effectors required to eliminate malignant tumor cells without the limitation of major histocompatibility complex (MHC) molecule issues. Hence, treatment which could stimulate NK cells is of great interest. Here, we investigated the efficacy of the combined therapy of TT-1 (a mutant of melittin) and interferon-α (IFN-α) on NK cells and human liver cancer HepG-2/Huh7 cells in vitro and in vivo, as well as the mechanism involved. The combination therapy significantly inhibited the growth of HepG-2/Huh7 cells in vivo, but this effect was impaired after depleting NK cells. TT-1 not only up-regulated MHC class I-related chain molecules A (MICA) expression, but also prevented the secretion of soluble MICA (sMICA). Both the mRNA and protein of a disintegrin and metallopeptidase 10 (ADAM 10) in HepG-2/Huh7 cells were decreased after TT-1 treatment. The combined therapy of TT-1 and IFN-α could suppress the growth of HepG-2/Huh7 xenografted tumor effectively via promoting the interaction of NK group 2, member D (NKG2D) and MICA, indicating that TT-1+IFN-α would be a potential approach in treating liver cancer.

Topics: Adjuvants, Immunologic; Amino Acid Sequence; Animals; Carcinoma, Hepatocellular; Cell Proliferation; Hep G2 Cells; Histocompatibility Antigens Class I; Humans; Interferon-alpha; Killer Cells, Natural; Liver Neoplasms; Liver Neoplasms, Experimental; Melitten; Mice; Mice, Nude; NK Cell Lectin-Like Receptor Subfamily K; Peptide Fragments; Xenograft Model Antitumor Assays

Vascular endothelial growth factor and its tyrosine kinase receptors have been identified as key mediators of the regulation of pathologic blood vessel growth and maintenance in the promotion of angiogenesis and tumor growth. Therefore, an alternative approach to destroying tumor endothelium would be to make this tissue particularly sensitive to VEGF-mediated drug delivery. To verify this hypothesis, we generated a protein containing VEGF165 fused to melittin. Melittin is a small linear peptide composed of 26 amino acid residues that can exert toxic or inhibitory effects on many types of tumor cells. This protein is a cytolytic peptide that attacks lipid membranes, leading to significant toxicity. In the present study, the Pichia pastoris expression system was used to express the fusion protein. Under optimal conditions, stable VEGF165-melittin production was achieved using a series of purification steps. The activity of VEGF165-melittin fusion protein was compared with melittin for its ability to suppress the growth of tumor cell line in vitro. The fusion toxin selectively inhibited growth of human hepatocellular carcinoma HepG-2 cell line with high expression of VEGFR-2. We found that sensitivity of VEGFR-2 transfected 293 cells to VEGF165-melittin enhanced as the cellular VEGFR-2 density increased. In an in vivo initial experiment, the fusion protein inhibited tumor growth in xenografts assays. Furthermore, successful expression and characterization of the fusion protein demonstrated its efficacy for use as a novel treatment strategy for cancer.

Topics: Animals; Antineoplastic Agents; Cell Line, Tumor; Cell Proliferation; Cell Survival; Hep G2 Cells; Humans; In Vitro Techniques; Liver Neoplasms; Melitten; Mice; Recombinant Fusion Proteins; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factor Receptor-2; Xenograft Model Antitumor Assays

Hepatocellular carcinoma (HCC) has a high mortality rate worldwide and still remains to be a noticeable public health problem. Therefore, new remedies are urgently needed. Melittin, a major component of bee venom, is known to suppress cell growth in various cancers including HCC. However, the mechanism of the anticancer effect of melittin on HCC has not been fully elucidated. It has been reported that Methyl-CpG binding protein 2 (MeCP2) plays a key role in tumor proliferation, apoptosis, migration and invasion. In the present study, we found the high expression of MeCP2 in human HCC tissues and in the SMMC-7721 cell line. MeCP2 silencing inhibited cell proliferation, while over-expression of MeCP2 promoted cell growth in SMMC-7721 cells. It indicates that MeCP2 may be an attractive target for human HCC. We further found that melittin could inhibit cell proliferation by reducing MeCP2 expression in vitro. Interestingly, the inhibitory effect of melittin on cell proliferation was due to a delay in G0/G1 cell cycle progression, without influencing cell apoptosis. Next, we investigated the potential molecular mechanisms and found that MeCP2 could modulate Shh signaling in SMMC-7721 cells. Further study indicates that melittin may induce the demethylation of PTCH1 promoter, resulting in the increased expression of PTCH1. Furthermore, the expression of Shh and GLI1 was significantly lowered upon treatment of melittin. These results suggest that melittin can block Shh signaling in vitro. In short, these results indicate that melittin inhibits cell proliferation by down-regulating MeCP2 through Shh signaling in SMMC-7721 cells.

Topics: Antineoplastic Agents; Carcinoma, Hepatocellular; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Proliferation; DNA Methylation; Dose-Response Relationship, Drug; Gene Expression Regulation, Neoplastic; Hedgehog Proteins; Humans; Liver Neoplasms; Melitten; Methyl-CpG-Binding Protein 2; Patched Receptors; Patched-1 Receptor; Promoter Regions, Genetic; Receptors, Cell Surface; Resting Phase, Cell Cycle; RNA Interference; Signal Transduction; Time Factors; Transcription Factors; Transfection; Zinc Finger Protein GLI1

Hepatocellular carcinoma (HCC) is a highly malignant disease, and its outcome of routine therapies is poor. Comprehensive treatment including gene therapy is an important way to improve patients' prognosis and survival. In this study, we successfully constructed a triple-controlled cancer-selective oncolytic adenovirus, QG511-HA-Melittin, carrying melittin gene, in which the hybrid promoter, hypoxia-response element (HRE)-AFP promoter, was used to control viral E1a expression targeting AFP-positive cancer cells in hypoxia microenviroment, and the E1b-55 kDa gene was deleted in cancer cells with p53-deficiency. The cytological experiments found that the viral replication of QG511-HA-Melittin was increased to 12800-folds in Hep3B cells within 48 h, and 130-folds in SMMC-7721, but the virus did not replicate in L-02 cells. QG511-HA-Melittin had a strong inhibition effect on AFP-positive HCC cell proliferation, such as Hep3B and HepG2, whereas, there was low or no inhibition effect of QG511-HA-Melittin on AFP-negative cancer cells SMMC-7721 and normal cells L-02. In the in vivo experiment, compared with the blank control group, QG511-HA-Melittin can significantly inhibit the growth of HCC xenografts (P<0.05). The survival of mice in QG511-HA-Melittin group was much longer than that of the blank control group. Both in vitro and in vivo experiments manifested that QG511-HA-Melittin exerts an inhibitory effect on HCC cells, which may provide a new strategy for HCC biotherapy.

Topics: Adenoviridae; Animals; Antineoplastic Agents; Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Proliferation; Genetic Therapy; Humans; Liver Neoplasms; Male; Melitten; Mice, Inbred BALB C; Mice, Nude; Oncolytic Virotherapy; Xenograft Model Antitumor Assays

Hepatic resection and orthotopic liver transplantation are the only potentially curative treatments for hepatocellular carcinoma (HCC) but are indicated only in a minority of patients. Biosynthetic nanoscale peptide Melittin (Mel) is postulated to disrupt microbial phospholipid membranes by formation of stable or transient pores. Survivin, a member of the inhibitor of apoptosis family, is transcriptionally upregulated in most malignant tissues but not in normal tissues. It has been reported that the survivin promoter activity is tumor-specific and makes it a good candidate for construction of gene therapy vectors. In the present study, a non-viral vector (pSURV-Mel), encoding Mel gene, was developed to evaluate its anti-tumor effect in HCC cell lines and in vivo in a mouse model of human HCC xenograft tumor. Our results showed that the survivin promoter is specifically activated in tumor cells, and the pSURV-Mel plasmid expressed Mel selectively in tumor cells and also induced cytotoxicity. Moreover, intratumoral Injection of pSURV-Mel significantly suppressed the growth of xenograft tumors. Mechanistically, pSURV-Mel induced cell death by an apoptosis-dependent pathway. All taken together, this study elucidates a relatively safe, highly effective and cancer specific gene therapy strategy for HCC. The mechanisms of non-viral vector-induced cell death which were revealed by this work will shed light on the construction of more powerful vectors for cancer therapy.

Topics: Animals; Biomimetic Materials; Carcinoma, Hepatocellular; Cell Line, Tumor; DNA; Female; Genetic Therapy; Humans; Inhibitor of Apoptosis Proteins; Liver Neoplasms; Melitten; Mice; Mice, Inbred BALB C; Nanoparticles; Promoter Regions, Genetic; Repressor Proteins; Survivin; Treatment Outcome

Melittin is a water-soluble toxic peptide derived from the venom of the bee. Although many studies show the anti-tumor activity of melittin in human cancer including glioma cells, the underlying mechanisms remain elusive. Here the effect of melittin on human hepatocelluar carcinoma HepG2 cell proliferation in vitro and further mechanisms was investigated. We found melittin could inhibit cell proliferation in vitro using Flow cytometry and MTT method. Besides, we discovered that melittin significantly downregulated the expressions of CyclinD1 and CDK4. Results of western Blot and Real-time PCR analysis indicated that melittin was capable to upregulate the expression of PTEN and attenuate histone deacetylase 2 (HDAC2) expression. Further studies demonstrated that knockdown of HDAC2 completely mimicked the effects of melittin on PTEN gene expression. Conversely, it was that the potential utility of melittin on PTEN expression was reversed in cells treated with a recombinant pEGFP-C2-HDAC2 plasmid. In addition, treatment with melittin caused a downregulation of Akt phosphorylation, while overexpression of HDAC2 promoted Akt phosphorylation. These findings suggested that the inhibitory of cell growth by melittin might be led by HDAC2-mediated PTEN upregulation, Akt inactivation, and inhibition of the PI3K/Akt signaling pathways.

Topics: Carcinoma, Hepatocellular; Cell Cycle Checkpoints; Cell Proliferation; Down-Regulation; Hep G2 Cells; Histone Deacetylase 2; Humans; Liver Neoplasms; Melitten; Proto-Oncogene Proteins c-akt; PTEN Phosphohydrolase; Signal Transduction; Tumor Suppressor Proteins

A random phage 12-mer peptide library and a whole-cell subtractive biopanning protocol against HepG2 cells were used to select a novel peptide-specific binding to hepatocellular carcinoma cells. As a result, peptide SLSLITMLKISR (AM-2) was screened as a novel homing peptide to hepatocellular carcinoma cells, tested by immunofluorescence and immunochemistry assays. Subsequently, peptide AM-2 was linked to melittin by A(EAAAK)2A, and the antitumor effect of this ligation product was detected by MTT assay, fluorescence-activated cell sorting, and scanning electron microscopy methods. Results of cell growth inhibition tests confirmed that the affinity of melittin was increased after being incorporated into AM-2, and AM-2-melittin specifically targeted and killed HepG2 cells in vitro. Thus, AM-2 is a valuable ligand for tumor targeting, which leads to increased binding and killing effect of hepatocellular carcinoma cells in vitro when ligated to melittin, and AM-2-melittin has a clinical potential application as target agents for the treatment of human hepatocellular carcinoma.

Topics: Amino Acid Sequence; Carcinoma, Hepatocellular; Cell Line, Tumor; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Humans; Liver Neoplasms; Melitten; Peptide Library; Peptides; Protein Binding

Melittin, a water-soluble toxic peptide derived from bee venom of Apis mellifera was reported to have inhibitory effects on hepatocellular carcinoma (HCC). However, its role in antimetastasis and the underlying mechanism remains elusive. By utilizing both HCC cell lines and an animal model based assay system, we found that Rac1, which has been shown to be involved in cancer cell metastasis, is highly expressed in aggressive HCC cell lines and its activity correlated with cell motility and cytoskeleton polymerization. In addition, Rac1-dependent activity and metastatic potential of aggressive HCC cells are remarkably high in both cellular and nude mouse models. We provide evidence here that melittin inhibits the viability and motility of HCC cells in vitro, which correlates with its suppression of Rac1-dependent activity, cell motility, and microfilament depolymerization. Furthermore, melittin suppresses both HCC metastasis and Rac1-dependent activity in nude mouse models. The specificity of the effect of melittin on Rac1 was confirmed in HCC cells both in vitro and in vivo.. Melittin inhibits tumor cell metastasis by reducing cell motility and migration via the suppression of Rac1-dependent pathway, suggesting that melittin is a potential therapeutic agent for HCC.

Topics: Actin Cytoskeleton; Animals; Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Movement; Cell Survival; Disease Models, Animal; Gene Expression Regulation, Neoplastic; Humans; Liver Neoplasms; MAP Kinase Kinase 4; Melitten; Mice; Mice, Nude; Neoplasm Metastasis; rac1 GTP-Binding Protein; Signal Transduction; Xenograft Model Antitumor Assays

Melittin has antitumor effects on osteosarcoma, leukemia, and cervical cancer in vitro. Our previous experiments showed that melittin could inhibit proliferation and induce apoptosis of human hepatocellular carcinoma BEL-7402 cells. This study was to examine the effects of melittin on the growth and angiogenesis of BEL-7402 cell xenografts in nude mice.. The xenografts derived from BEL-7402 cells were established in BALB/C nude mice. Inoculated mice were randomly divided into normal saline (NS, 10 ml/kg) group, positive control (thalidomide, TLD, 200 mg/kg) group, low dose melittin (40 microg/kg) group, moderate dose melittin (60 microg/kg) group and high dose melittin (80 microg/kg) group. Tumor volume was measured. Tumor tissue was observed under microscope. Microvessel density (MVD) and the expression of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) and nuclear factor kappaB (NF-kappaB) were detected by SABC immunohistochemistry. The mRNA levels of VEGF and bFGF were analyzed by real-time fluorescent quantitative polymerase chain reaction.. The relative tumor volume (V/V0) and MVD were significantly lower in low, moderate and high dose melittin groups than in NS group (4.42+/-0.58, 3.47+/-0.97, and 3.06+/-1.23 vs. 9.06+/-1.45, P<0.01; 11.33+/-1.86, 9.17+/-1.17, and 6.67+/-1.21 vs. 16.50+/-2.35, P<0.01). Tumor tissue necrosis was observed in melittin-treated groups and tumor vessels were destroyed by melittin. The positive expression indexes of VEGF (2.59+/-0.27, 2.61+/-0.17, 1.55+/-0.22 vs. 3.80+/-0.60, P<0.01), bFGF (2.45+/-0.78, 2.27+/-0.36, 2.10+/-0.27 vs. 4.43+/-0.34, P<0.01) and NF-kappaB (2.79+/-0.29, 2.71+/-0.66, 2.26+/-0.56 vs. 4.98+/-0.63, P<0.01) were significantly lower in low, moderate and high dose melittin groups than in NS group. The mRNA levels of VEGF and bFGF were also significantly lower in melittin-treated groups than in NS group.. Melittin could inhibit the growth of BEL-7402 cell xenografts in nude mice. The down-regulation of VEGF, b-FGF and NF-kappaB expression and the inhibition of angiogenesis might play key roles in the antitumor effect of melittin.

Topics: Animals; Antineoplastic Agents; Carcinoma, Hepatocellular; Cell Line, Tumor; Down-Regulation; Fibroblast Growth Factor 2; Humans; Liver Neoplasms; Male; Melitten; Mice; Mice, Inbred BALB C; Mice, Nude; Microvessels; Neoplasm Transplantation; Neovascularization, Pathologic; NF-kappa B; Random Allocation; RNA, Messenger; Tumor Burden; Vascular Endothelial Growth Factor A

Recently the use of peptides in bee venom (PBV) for cancer therapy has attracted considerable attention. In this study, the sterically stabilized liposomal PBV (PBV-SL) was prepared using soybean phosphatidylcholine, cholesterol, and cholesterol-PEG-COOH. The humanized antihepatoma disulfide-stabilized Fv (hdscFv25) was coupled to sterically stabilized liposomes using the N-hydroxysuccinimide ester method. The hdscFv25-immunoliposomes (SIL[hdscFv25]) were immunoreactive as determined by ELISA assay. SIL[hdscFv25] showed higher tumor cells selectivity. PBV-SIL[hdscFv25] can kill SMMC-7721 cells in vitro with higher efficiency than non-targeted liposomes. Whereas cytotoxicties were compared for Hela cells, no significant differences was observed between PBV-SIL[hdscFv25] and PBV-SL. Sterically stabilized immunoliposomal peptides in bee venom could be one drug targeting delivery system.

Topics: Antineoplastic Agents; Bee Venoms; Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Survival; Cholesterol; Drug Delivery Systems; HeLa Cells; Humans; Immunoconjugates; Liposomes; Liver Neoplasms; Melitten; Peptides; Recombinant Proteins

Combination of the degradable polymeric gene carriers OEI-HD-1 and LT- OEI-HD-1 with an EGF targeting conjugate resulted in strongly (up to 900-fold) enhanced polyplex activity in EGF-receptor rich HUH7 hepatocellular carcinoma cells. The targeting ligand effect was DNA dose dependent, could be blocked by competitive receptor binding with unbound EGF ligand, and was not observed in receptor-negative control cells. Measures which enhance intracellular endosomal escape, either photochemically enhanced intracellular release (PCI) or the incorporation of a novel membrane-active melittin analog NMA-3, further enhanced gene transfer activity of EGF/OEI-HD-1 polyplexes.

Topics: Animals; Binding, Competitive; Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Membrane; DNA; Endosomes; Epidermal Growth Factor; ErbB Receptors; Genes, Reporter; Genetic Therapy; Humans; Liver Neoplasms; Luciferases; Melitten; Mice; Photosensitizing Agents; Plasmids; Polyethyleneimine; Porphyrins; Transfection

The aim of this study was to investigate the cellular effects of melittin on the growth and apoptosis of human hepatocellular carcinoma (HCC) cells and to provide the molecular mechanism for potential application of a recombinant adenovirus carrying the melittin gene (Ad-rAFP-Mel) in the treatment of liver cancer.. Human HCC cells (BEL-7402) were infected with Ad-rAFPMel at different times. In vitro cell growth was determined by MTT assay. Cellular apoptosis was evaluated quantitatively and qualitatively by phase-contrast microscopy, transmission electron microscopy, DNA ladder electrophoresis, TUNEL staining and flow cytometry.. Ad-rAFP-Mel infection had an inhibitory effect on the proliferation of BEL-7402 cells. The morphological changes of apoptosis were confirmed by microscopy and DNA electrophoresis. The ultrastructural characteristics of apoptotic cells, such as chromatin condensation and nuclear fragmentation, were also observed by electron microscopy in the Ad-rAFP-Mel-infected cells. Ad-rAFPMel infection markedly induced cellular apoptosis, and Fas expression on Bel-7402 cells infected by Ad-rAFPMel was up-regulated.. The fact that melittin can induce apoptosis of the HCC cell line BEL-7402 leads us to consider adenovirus-mediated delivery of melittin as a promising approach for the treatment of HCC. However, the underlying mechanism needs to be further investigated.

Topics: Apoptosis; Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Proliferation; Cell Survival; Humans; Liver Neoplasms; Melitten; Recombinant Proteins

To search for a new clinical application of melittin (Mel): treating hepatocellular carcinoma with Mel gene.. Recombinant adenoviruses carrying the Mel gene and alpha-fetoprotein (AFP) promoter (Ad-rAFP-Mel) were constructed through a bacterial homologous recombinant system. The efficiency of adenovirus-mediated gene transfer and the inhibitory effect of Ad-rAFP-Mel on the proliferation of hepatocarcinoma cells were determined by X-gal stain and MTT assay, respectively. The tumorigenicity of hepatocarcinoma cells transfected by Ad-rAFP-Mel and the antitumor effect of Ad-rAFP-Mel on transplanted tumor in nude mice were detected in vivo.. The Mel mRNA was transcribed in BEL-7402 hepatocellular carcinoma cells transducted by Ad-rAFP-Mel. The efficiency of adenovirus-mediated gene transferred to BEL-7402 cells was 100% when the multiplicity of infection of Ad-rAFP-Mel was 10 in vitro, and was also high in vivo. The inhibitive rates of Ad-rAFP-Mel and Ad-rAFP for BEL7402 cells were 66.2 +/- 2.7% and 2.9 +/- 2.3% (t=30.83, P=6.6 x 10(-6)) by MTT assay. The inhibitive rates of Ad-CMV-Mel for BEL7402, SMMC7721 and L02 cells were 58.9 +/- 9.6%, 65.9 +/- 3.8% and 31.7 +/- 1.2%, respectively, and of Ad-rAFP-Mel were 66.2 +/- 2.7%, 16.1 +/- 6.6% and 7.5 +/- 3.3%, respectively (t=1.27, P=0.27; t=11.31, P=3.5 x 10(-4); and t=12.12, P=2.7 x 10(-4) versus the Ad-CMV-Mel group in the same cells). The tumorigenicity rates of hepatocarcinoma cells transfected by Ad-rAFP-Mel were decreased. A significant antineoplastic effect was detected on transplanted tumor in nude mice by intratumoral injection of Ad-rAFP-Mel.. Ad-rAFP-Mel can inhibit specifically proliferation of AFP-producing human hepatocarcinoma cells in vitro and in vivo. This suggests that animal toxin gene can be used as an antitumor gene.

Topics: Adenoviridae; Animals; Carcinoma, Hepatocellular; Cell Proliferation; Gene Transfer Techniques; Genetic Therapy; Humans; Liver Neoplasms; Male; Melitten; Mice; Mice, Inbred BALB C; Mice, Nude; Neoplasms, Experimental; Promoter Regions, Genetic; Transfection; Transplantation, Heterologous; Tumor Cells, Cultured

To observe the induced apoptosis of recombinant adenovirus carrying melittin gene (Ad-rAFP-Mel) for hepatocellular carcinoma cell line (BEL-7402).. The morphological observe, DNA electrophoresis, TUNEL and Flow cytometry assay were used to study the apoptosis of BEL-7042 cell line transfected by Ad-rAFP-Mel.. The morphological changes and apoptosis of BEL-7402 transfected by Ad-rAFP-Mel were confirmed with microscopy and DNA electrophoresis, TUNEL, Flow cytometry assay. The DNA ladder could be demonstrated on DNA electrophoresis in Ad-rAFP-Mel group. The apoptosis rates of BEL-7402 cells in Ad-rAFP-Mel, Ad-rAFP, and control groups were (21.5+/-2.4)%, (10.5+/-4.4)% and (3.0+/-1.4)% respectively by TUNEL assay (F = 38.0, P < 0.05) and were (7.3+/-0.5)%, (3.9+/-0.1)% and (0.8+/-0.1)% respectively by flow cytometry assay (F = 415.1, P < 0.05).. It seems that melittin inducing apoptosis might be one of the antitumor mechanisms.

Topics: Adenoviridae; Apoptosis; Carcinoma, Hepatocellular; Cell Line, Tumor; Gene Expression; Gene Silencing; Genetic Therapy; Genetic Vectors; Humans; Liver Neoplasms; Melitten; Recombinant Fusion Proteins; Transcription, Genetic; Transfection