Gene Therapy For Carcinoma Research Articles

SiRNA Knocking Down in HepG2 Cells Identifies PFKFB4 and HNF4α as Key Genes Important for Cancer Cell Survival.

Liposomes are versatile delivery systems for encapsulating small interfering RNAs (siRNAs) because they enhance cellular uptake and gene silencing. This study compares the new liposome formula to commercial lipofectamine in delivering siRNAs targeting hepatic carcinoma genes, focusing on HNF4-α and PFKFB4. Flow cytometry and confocal microscopy revealed efficient internalization of PE-Rhod- B labeled lipoplexes in HepG2 cells, while cytotoxicity assays demonstrated significant reductions in cell viability, particularly with siHNF4-α and siPFKFB4. The newly formulated liposomes showed superior efficacy, achieving nearly 93% cytotoxicity at 100 nM, compared to just 50% with lipofectamine at the same concentration. Furthermore, real-time PCR confirmed that the liposome-encapsulated siHNF4-α reduced HNF4-α mRNA expression by tenfold at 100 nM, compared to a twofold reduction with lipofectamine at 200 nM. Similarly, siPFKFB4 delivered via liposomes showed a dose-dependent 35-fold reduction in PFKFB4 mRNA expression at 100 nM, outperforming the maximum reduction achieved by lipofectamine. The IC50 values for all siRNA treatment groups were significantly lower when using the liposome formula, reflecting improved delivery efficiency. These results demonstrate the potential of liposome formulations for therapeutic siRNA delivery. The encapsulation enhances cellular uptake and gene silencing efficiency, making the liposome formula a promising candidate for targeted gene therapy in hepatic carcinoma. Further research should explore it's in vivo biodistribution and potential combination therapies.

ALPPL2‐Binding Peptide Facilitates Targeted mRNA Delivery for Efficient Hepatocellular Carcinoma Gene Therapy

AbstractmRNA‐based gene therapy has emerged as an advanced strategy for hepatocellular carcinoma (HCC) treatment. However, one of its main limitations is lacking delivery precision in vivo. Distinguishing HCC cells from other tissues is crucial for maintaining the high stability, positive therapeutic outcomes, and safety of mRNA therapeutics. Here, a novel HCC specific peptide HCC167 is developed by phage display. It is shown that the HCC167 peptide can specifically recognize variety of HCC substrates including patient samples with high affinity. Modifying nanoparticles with the HCC167 peptide facilitates an effective HCC active targeting ability. When loading with Bims‐encoding mRNA, the systemically administrated DMP‐HCC167/Bims complex efficiently suppresses HCC progression in multiple models including patient‐derived xenografts. Furthermore, the ALPPL2 protein is identified as a specific binding receptor of the HCC167 peptide on the HCC cell membrane, uncovering a new active targeting mechanism based on the HCC167‐ALPPL2 ligand–receptor complex. The binding structure is also investigated by computer‐aided modeling. The study hence exhibits a potent active targeting strategy for mRNA‐based HCC therapy.

Anti-Micro RNA-221 a Promising Genetic Therapy of Oral Squamous Cell Carcinoma (SCC-25).

Micro-RNA-221(miR-221) is one of oncogenic miRNAs that plays a vital role in the development and progression of oral cancers. The aim of this study is to introduce a new gene therapy for oral squamous cell carcinoma by blocking the expression of oncogenic miR-221 by its inhibitor. The present work was performed on squamous cell carcinoma cell line SCC-25 and anti-miR-221 was delivered to the cells using an ultrasound micro bubbles. Assessment of the effect of miR-221 inhibitor on SCC-25 cells was done using MTT assay, cell cycle analysis and apoptosis detection. In addition, reverse transcription-polymerase chain reaction was also used to detect the expression -miR-221 and its target genes. Using ANOVA, statistical analysis of the results showed significant inhibition of cell viability with and induction of cell apoptosis of SCC-25 cell line after transfection. Moreover, the expression of miR-221, Epidermal growth factor receptor (EGFR) and CDKNIB/p27 were downregulated without significant difference. Transfection of SCC-25 by inhibitor of miR-221 resulting in blockage of its expression leading to arresting of tumor growth. These results proved the effective role of micro-RNA inhibitors as novel therapeutic agent for oral cancers.

Efficient miRNA Inhibitor Delivery with Graphene Oxide-Polyethylenimine to Inhibit Oral Squamous Cell Carcinoma.

BackgroundMicroRNAs (miRNAs) are widely believed to be promising targets for oral squamous cell carcinoma (OSCC) gene therapy. miR-214 has been identified as a promoter of OSCC aggression and metastasis.MethodsGraphene oxide-polyethylenimine (GO-PEI) complexes were prepared and loaded with a miRNA inhibitor at different N/P ratios. The transfection efficiency of GO-PEI-inhibitor was tested in Cal27 and SCC9 cells. Moreover, the tumor inhibition ability of GO-PEI-inhibitor was measured in an OSCC xenograft mouse model by intratumoral injection.ResultsHere, we show that a GO-PEI complex efficiently delivers a miR-214 inhibitor into OSCC cells and controls the intracellular release of the miR-214 inhibitor. These results indicate that the GO-PEI-miR-214 inhibitor complex efficiently inhibited cellular miR-214, resulting in a decrease in OSCC cell invasion and migration and an increase in cell apoptosis by targeting PTEN and p53. In the xenograft mouse model, the GO-PEI-miR-214 inhibitor complex significantly prevented tumor volume growth.ConclusionThis study indicates that functionalized GO-PEI with low toxicity has promising potential for miRNA delivery for the treatment of OSCC.

δ-Catenin regulates proliferation and apoptosis in renal cell carcinoma via promoting β-catenin nuclear localization and activating its downstream target genes.

δ‐Catenin is a unique member of the catenin family and is proved to be overexpressed in diverse human cancer types. However, the clinical significance and underling mechanism of δ‐catenin expression in renal cell carcinoma (RCC) remain elusive. Herein, we detected the protein expression of δ‐catenin in 28 clinical specimens of paired renal cancer tissues and normal renal tissues by Western blot analysis. δ‐Catenin expression in 58 cases of renal cell carcinoma was also examined by immunohistochemistry, and its association with clinicopathological factors was analyzed by statistical analysis. In vitro and in vivo assays were employed to further explore the biological role of δ‐catenin in RCC. The results showed that δ‐catenin was highly expressed in both clinical samples and cell lines of RCC. RCC patients with higher δ‐catenin expression had a more advanced pTNM stage and tumor stage as well as lymph nodes metastasis than those with lower expression. By regulating the nuclear translocation of β‐catenin and β‐catenin‐mediated oncogenic signals, δ‐catenin promoted proliferation and inhibited apoptosis in RCC. In vivo assay indicated δ‐catenin facilitated tumor growth in ACHN cell xenograft mouse model. Taken together, our study suggests that δ‐catenin might be considered as a novel prognostic indicator and actionable target for gene therapy in renal cell carcinoma.

Human Bone Marrow Mesenchymal Stem Cells Functionalized by Hybrid Baculovirus-Adeno-Associated Viral Vectors for Targeting Hypopharyngeal Carcinoma.

Hypopharyngeal carcinoma is a common malignant tumor of the head and neck with a very poor prognosis; the median survival time for curatively treated patients was 17.2 months in India. However, cell-based gene therapy holds promise to improve patient outcomes. In this study, we investigated whether human bone marrow mesenchymal stem cells (BMSCs) possess potential homing capacity for hypopharyngeal carcinoma. To monitor the efficiency of BMSC transplantation therapy through reporter gene imaging, we employed a hybrid baculovirus vector containing the Luc-P2A-eGFP fusion or sodium iodide symporter (NIS) sequence under the control of the cytomegalovirus promoter. To enhance the transfection efficiency, baculovirus vectors (Bac-CMV-Luc-P2A-eGFP-ITR and Bac-CMV-NIS-ITR) were flanked by inverted terminal repeats (ITRs), which are key elements of adeno-associated viruses. The infection efficiency of Bac-CMV-Luc-P2A-eGFP-ITR in BMSCs was as high as 92.84 ± 1.14% with no obvious toxic effects at a multiplicity of infection of 400. Moreover, Bac-CMV-NIS-ITR-infected BMSCs showed highly efficient radioactive iodide (125I) uptake; these high uptake levels were maintained for at least 2 h. Transwell migration assays further demonstrated the chemotaxis of BMSCs to hypopharyngeal carcinoma cells (FaDu cells) in vitro. BMSCs modified by firefly luciferase report gene or NIS were injected into nude mice with hypopharyngeal carcinoma, and changes in the localization of the BMSCs were successfully tracked with bioluminescent imaging and micro-single-photon emission computed tomography imaging. These data indicate the potential utility of BMSCs as a promising targeted-delivery vehicle for hypopharyngeal carcinoma gene therapy. Importantly, BMSCs may represent a promising targeting vector for general tumor radionuclide therapy.

Insulin-like Growth Factor I Receptor: A Novel Target for Hepatocellular Carcinoma Gene Therapy.

Human insulin-like growth factor (IGF) axis affects the molecular pathogenesis of hepatocellular carcinoma (HCC), especially in the abnormality of hepatic IGF-I receptor (IGF-IR) or IGF-II expression as a key molecule in hepatocarcinogenesis. However, the over-expression of hepatic IGFIR is associated with HCC progression with largely unknown mechanisms. The IGF-IR as one key molecule of the IGF signal pathway plays an important role in the hepatocyte malignant transformation. Attaching importance to IGF-IR might improve the prognostic or the therapeutic technique of HCC. This article reviews IGF-IR alteration during HCC development, and the effects of silencing IGF-IR gene by specific short hairpin RNA on the inhibition of cell proliferation in vitro or HCC xenograft growth in vivo to elucidate it as a novel molecular-targeted therapy for HCC.

Suicide Gene Therapy for Oral Squamous Cell Carcinoma.

Suicide gene therapy has been tested for the treatment of a variety of cancers, including oral cancer. Among the various suicide gene therapy approaches that have been reported, the Herpes Simplex Virus thymidine kinase (HSV-tk)/ganciclovir (GCV) system is one of the most extensively studied systems, holding great promise in cancer therapy. In this chapter, we describe methods to use the HSV-tk/GCV system to achieve antitumor activity, both in cultured oral cancer cells and in orthotopic and subcutaneous murine models of oral squamous cell carcinoma, using ligand-associated lipoplexes for enhancing therapeutic delivery.

Transferrin-functionalized chitosan-graft-poly(l-lysine) dendrons as a high-efficiency gene delivery carrier for nasopharyngeal carcinoma therapy

Transferrin-functionalized chitosan-graft-poly(l-lysine) dendrons (CS-PLLD-Tf) were synthesized in this work and then used to deliver the MMP-9 shRNA plasmid to HNE-1 cells for gene therapy. The obtained CS-PLLD-Tf displayed excellent gene transfection in vitro, verifying the strategy of constructing a high-efficiency gene carrier by conjugating the multiple low generation PLLDs to one molecule of CS to form a copolymer. Particularly, CS-PLLD-Tf showed much higher gene transfection efficiency than PEI-25k and CS-PLLD in the presence of serum, and more than 40% HNE-1 cells were transfected. This result led to an obvious decrease in MMP-9 protein expression and the apoptosis of HNE-1 cells as well as inhibition of the growth and invasion of HNE-1 tumors. The in vivo anti-tumor assay showed that CS-PLLD-Tf delivered the MMP-9 plasmid effectively and the tumor volume decreased more than 77% compared with the PBS control group. Moreover, CS-PLLD-Tf displayed higher bioavailability and good biocompatibility such as lower cytotoxicity, excellent blood compatibility and non-toxicity to organs, suggesting a potential application in gene therapy of tumors.

Bifidobacterium breve as a delivery vector of IL-24 gene therapy for head and neck squamous cell carcinoma in vivo.

Beneficial bacteria are becoming ever more popular gene delivery method for hypoxia-tumor targeting in vivo. In this study we investigated the therapeutic effect of new recombinant Bifidobacterium breve strain expressing interleukin (IL)-24 gene (B. breve-IL24) on head and neck tumor xenograft in mice. Briefly, B. breve transformants were obtained through electro-transformation. Bacteria-tumor-targeting ability were analyzed in vivo over different time points (1, 3 and 7 days post-bacteria injection). Furthermore, the therapeutic effect of bacteria on tumor cells in vivo were analyzed as follows: 30 Balb/c nude mice bearing subcutaneous tumor were randomly divided in three groups (Drug group, green fluorescent protein (GFP) group and Saline group). The therapy lasted for 2 weeks and included B. breve-IL24 administration via tail vein for Drug group, B. breve-GFP for GFP group and phosphate buffered saline for Saline group. The tumor growth was monitored using standard caliper technique, while the apoptosis induction in vivo was analyzed by Real-time Positron Emission Tomography/Computed Tomography (PET/CT) imaging ([18F]-ML-10 tracer). At the end of the experiment, tumor tissues were collected and analyzed by western blotting. Briefly, our results suggested that our new recombinant bacterium has the capability of targeting tumor tissue in vivo. As for the therapeutic effect, our new strain has revealed to be a promising therapeutic approach against tumor growth in vivo. Briefly, higher tumor growth inhibition and higher tumor cell apoptosis induction were observed in Drug group compared with the GFP and Saline groups. To conclude, a new recombinant strain B. breve-IL24 offers a novel, safe and clinically acceptable therapeutic approach for tumor therapy in vivo.

Gene Therapy for Pancreatic Cancer: Specificity, Issues and Hopes.

A recent death projection has placed pancreatic ductal adenocarcinoma as the second cause of death by cancer in 2030. The prognosis for pancreatic cancer is very poor and there is a great need for new treatments that can change this poor outcome. Developments of therapeutic innovations in combination with conventional chemotherapy are needed urgently. Among innovative treatments the gene therapy offers a promising avenue. The present review gives an overview of the general strategy of gene therapy as well as the limitations and stakes of the different experimental in vivo models, expression vectors (synthetic and viral), molecular tools (interference RNA, genome editing) and therapeutic genes (tumor suppressor genes, antiangiogenic and pro-apoptotic genes, suicide genes). The latest developments in pancreatic carcinoma gene therapy are described including gene-based tumor cell sensitization to chemotherapy, vaccination and adoptive immunotherapy (chimeric antigen receptor T-cells strategy). Nowadays, there is a specific development of oncolytic virus therapies including oncolytic adenoviruses, herpes virus, parvovirus or reovirus. A summary of all published and on-going phase-1 trials is given. Most of them associate gene therapy and chemotherapy or radiochemotherapy. The first results are encouraging for most of the trials but remain to be confirmed in phase 2 trials.

Downregulation of miRNA-26b inhibits cancer proliferation of laryngeal carcinoma through autophagy by targeting ULK2 and inactivation of the PTEN/AKT pathway.

Laryngeal carcinoma is one of the most common tumors of the head and neck cancers, the pathogenesis of which remains yet unclear. It has been discovered through research that microRNAs (miRNAs) play an important role during the genesis of laryngeal carcinoma. In the present study we investigated the effect of miRNA-26b on the proliferation of laryngeal carcinoma and elucidated the potential underlying mechanisms in order to provide new targets for laryngeal carcinoma. Firstly, we found that miRNA-26b expression was significantly increased in patients with laryngeal carcinoma, compared with normal volunteers. The downregulation of miRNA-26b inhibited cell proliferation and induced apoptosis of Hep-2 cells. Furthermore, downregulation of the expression of miRNA‑26b promoted Bax, LC3 and p62 protein expression, decreased ULK2 mRNA and protein expression, as well as PTEN protein expression and increased phosphorylated‑AKT protein expression in Hep-2 cells as determined using quantification by real-time PCR and western blotting. The concomitant downregulation of ULK2 and miRNA-26b futher enhanced the miRNA‑26b-induced autophagy and apoptosis in addition to the miRNA-26b-inhibited cell proliferation of Hep-2 cells by targeting ULK2 and inactivating the PTEN/AKT pathway as determined by immunocytofluorescence. These findings revealed that miRNA-26b may play a key role in cell growth and death of laryngeal carcinoma through ULK2 and the PTEN/AKT pathway, and thus may be a new target for gene therapy in laryngeal carcinoma.

170 - Antitumor effect for combined therapy of SOCS-1 gene therapy and radiotherapy in esophageal squamous cell carcinoma

170 - Antitumor effect for combined therapy of SOCS-1 gene therapy and radiotherapy in esophageal squamous cell carcinoma

Autoregulated expression of p53 from an adenoviral vector confers superior tumor inhibition in a model of prostate carcinoma gene therapy

ABSTRACTAlternative treatments for cancer using gene therapy approaches have shown promising results and some have even reached the marketplace. Even so, additional improvements are needed, such as employing a strategically chosen promoter to drive expression of the transgene in the target cell. Previously, we described viral vectors where high-level transgene expression was achieved using a p53-responsive promoter. Here we present an adenoviral vector (AdPGp53) where p53 is employed to regulate its own expression and which outperforms a traditional vector when tested in a model of gene therapy for prostate cancer. The functionality of AdPGp53 and AdCMVp53 were compared in human prostate carcinoma cell lines. AdPGp53 conferred greatly enhanced levels of p53 protein and induction of the p53 target gene, p21, as well as superior cell killing by a mechanism consistent with apoptosis. DU145 cells were susceptible to induction of death with AdPGp53, yet PC3 cells were quite resistant. Though AdCMVp53 was shown to be reliable, extremely high-level expression of p53 offered by AdPGp53 was necessary for tumor suppressor activity in PC3 and DU145. In situ gene therapy experiments revealed tumor inhibition and increased overall survival in response to AdPGp53, but not AdCMVp53. Upon histologic examination, only AdPGp53 treatment was correlated with the detection of both p53 and TUNEL-positive cells. This study points to the importance of improved vector performance for gene therapy of prostate cancer.

Expression and clinical significance of moesin and E-cadherin in invasive carcinoma of breast, no specific type

To investigate the correlation of moesin and E-cadherin with biological behavior of breast cancer and its mechanism by comparing expression of moesin and E-cadherin in breast invasive carcinoma of no specific type(BIC-NST), breast ductal carcinoma in situ(BDCIS) and normal breast tissues adjacent to carcinoma. Breast cancer cases of the Huizhou Municipal Center People Hospital were collected between Jan 2008 and Dec 2010, expression of moesin and E-cadherin in 104 cases of BIC-NST, 84 cases of BDCIS and 53 cases of normal breast tissues adjacent to carcinoma were detected by tissue-microarray and SP immunohistochemical staining. Western blot was used to detect moesin expression of 16 BIC-NST fresh tissues. Expression rate of moesin in BIC-NST and BDCIS were significantly higher than normal tissues(P<0.01), but the expression rate of E-cadherin in BIC-NST and BDCIS were significantly lower than those of normal tissues(P<0.01). Expression rate of moesin in BIC-NST grade Ⅲ group was significantly higher than that of the grade Ⅰ group.There was a significantly positive correlation between histological grade and moesin expression(P<0.05). However, E-cadherin expression rate in BIC-NST grade Ⅲ group was significantly lower than that in grade Ⅰ group , and there was a significantly negative correlation between histological grade and E-cadherin expression(P<0.05). Moreover, no significant correlation was observed between moesin and E-cadherin expression in BDCIS tissues. Expression of moesin in clinical stage Ⅱ + Ⅲ BIC-NST was significantly higher than that in stage Ⅰ(P<0.01) . Expression of moesin was significantly associated with lymph node metastasis (P<0.01). But no significant correlation was observed between moesin expression and age, tumor size and vascular invasion . However, expression of E-cadherin in clinical stage Ⅱ+ Ⅲ BIC-NST was significantly lower than that in stage Ⅰ(P<0.01). Expression of E-cadherin was significantly associated with lymph node metastasis and vascular invasion (P<0.01). But no significant correlation was observed between E-cadherin expression, age and tumor size. There was a negative correlation between expression of moesin and E-cadherin in BIC-NST(P=0.021)and BDCIS(P=0.032). Higher moesin and lower E-cadherin signal transduction is closely related to the recurrence and development of breast carcinoma, therefore moesin and E-cadherin might provide new targets for gene therapy in breast carcinoma.

680. TransfeX-Mediated HSV-tk/Ganciclovir Suicide Gene Therapy in HeLa Cervical Carcinoma and HSC-3, FaDu, and H357 Oral Cancer Cells

INTRODUCTION: Herpes Simplex Virus thymidine kinase/ganciclovir (HSV-tk/GCV)-induced suicide gene therapy has been used to successfully treat various cancers. TransfeX-mediated transfection of pCMV.Luc into HeLa cervical carcinoma and HSC-3, FaDu, and H357 oral squamous cell carcinoma (OSCC) cell lines in the presence of 10% serum has proved to be highly efficient, with low non-specific cytotoxicity. The plasmid pNGVL1-tk encoding HSV-tk under the control of the CMV promoter was delivered to the cells in vitro via the novel cationic liposomal reagent, TransfeX, followed by treatment with ganciclovir. METHODS: HeLa, HSC-3, FaDu, and H357 cells were seeded in 48-well culture plates one day prior to transfection, at a density of 1.5 × 105 cells per well in 1 ml of complete DMEM medium containing 10% serum (DME/10). Lipoplexes for transfection were prepared by mixing 1 µg pCMV.Luc or pNGVL1-tk per 100 µl of serum-free DMEM followed by the addition of 2 µl of TransfeX. Lipoplexes were added directly to cells grown in the presence of 10% serum. Twenty-four hours after transfection, 20 µg of ganciclovir was added per well and cells were incubated an additional 24 h. The Alamar blue assay was used to determine cell viability after 24, 48, and 120 h. Fresh medium and ganciclovir were added after each of these readings. RESULTS: After 24 h, 65% and 50% cytotoxicity were seen in HeLa cells and HSC-3 cells, respectively. FaDu and H357 cells were more resistant to treatment, showing 15% and 5% cytotoxicity, respectively, after 24 h. After 120 h, cytotoxicity remained the same in HeLa, and increased to 90% in HSC-3. In FaDu and H357, 65% and 30% cytotoxicity, respectively, were observed after 120 h. CONCLUSIONS: Suicide gene therapy is most effective against HSC-3 OSCC among the cells examined. Longer periods of incubation with ganciclovir or prolonged gene expression via enhanced episome vectors may be necessary to increase cytotoxicity in FaDu and H357 cells.View Large Image | Download PowerPoint SlideView Large Image | Download PowerPoint Slide

Image-aided Suicide Gene Therapy Utilizing Multifunctional hTERT-targeting Adenovirus for Clinical Translation in Hepatocellular Carcinoma.

Trans-splicing ribozyme enables to sense and reprogram target RNA into therapeutic transgene and thereby becomes a good sensing device for detection of cancer cells, judging from transgene expression. Previously we proposed PEPCK-Rz-HSVtk (PRT), hTERT targeting trans-splicing ribozyme (Rz) driven by liver-specific promoter phosphoenolpyruvate carboxykinase (PEPCK) with downstream suicide gene, herpes simplex virus thymidine kinase (HSVtk) for hepatocellular carcinoma (HCC) gene therapy. Here, we describe success of a re-engineered adenoviral vector harboring PRT in obtaining greater antitumor activity with less off-target effect for clinical application as a theranostics. We introduced liver-selective apolipoprotein E (ApoE) enhancer to the distal region of PRT unit to augment activity and liver selectivity of PEPCK promoter, and achieved better transduction into liver cancer cells by replacement of serotype 35 fiber knob on additional E4orf1-4 deletion of E1&E3-deleted serotype 5 back bone. We demonstrated that our refined adenovirus harboring PEPCK/ApoE-Rz-HSVtk (Ad-PRT-E) achieved great anti-tumor efficacy and improved ability to specifically target HCC without damaging normal hepatocytes. We also showed noninvasive imaging modalities were successfully employed to monitor both how well a therapeutic gene (HSVtk) was expressed inside tumor and how effectively a gene therapy took an action in terms of tumor growth. Collectively, this study suggests that the advanced therapeutic adenoviruses Ad-PRT-E and its image-aided evaluation system may lead to the powerful strategy for successful clinical translation and the development of clinical protocols for HCC therapy.

Abstract 4213: Gene therapy for esophageal squamous cell carcinoma using SOCS-1 expressing adenoviral vector

Abstract Objective Esophageal squamous cell carcinoma (ESCC) is the major histological form of oesophageal cancer in East Asian countries. Despite recent improvements in multimodality treatment, including surgery, radiotherapy and chemotherapy, the prognosis for patients with ESCC remains unsatisfactory. Therefore, development of novel therapy is urgently required. Suppressor of cytokine signaling-1 (SOCS-1) has been cloned as a negative regulator of various cytokine signaling. Recently, lines of evidences suggest that overexpression of SOCS-1 has been considered as a promising therapeutic approach for treatment of cancer. This study was aimed to evaluate the therapeutic effect of the overexpression of SOCS-1 by adenoviral vector (AdSOCS-1) against ESCC. METHODS By WST-8 assay, cell growth inhibition via infection with various dose of AdSOCS-1 was evaluated in 11 ESCC cell lines in vitro. We also evaluated the induction of apoptosis by overexpression of SOCS-1 in vitro. To evaluate the therapeutic efficacy of AdSOCS-1, ESCC cell line (TE-8) was subcutaneously implanted into the ICR nu/nu mice. In addition, therapeutic efficacy was also assessed using human xenografts subcutaneously transplanted in imuune-deficient mice. Mice were intra-tumorally injection with AdSOCS-1 or control adenovirus vector (AdLacZ) twice a week for 4weeks. RESULTS Among 11 ESCC cell lines, AdSOCS-1, markedly suppressed proliferation of 8 ESCC cell lines in vitro. AdSOCS-1 strongly induced apoptosis in vitro, via inhibiting JAK/STAT3 pathway. In the TE8 xenograft model, the volume of tumors after therapy in AdSOCS-1 group (1,580 ± 1,005mm3) were lower than that in control group (2,690 ± 1,309mm3). Furthermore, compared with AdLacZ, injection with AdSOCS-1 also inhibited the tumor growth of patient derived ESCC xenografts. CONCLUSIONS Our results indicated that overexpression of SOCS-1 inhibited the progression of ESCC both in ESCC cell lines and ESCC xenograft model derived from patient. SOCS-1 can be a promising novel therapeutic approach for intra-lesional therapy in ESCC. Citation Format: Hisashi Hara, Rie Nakatsuka, Tsuyoshi Takahashi, Satoshi Serada, Minoru Fujimoto, Yasuhiro Miyazaki, Tomoki Makino, Yukinori Kurokawa, Makoto Yamasaki, Hiroshi Miyata, Kiyokazu Nakajima, Shuji Takiguchi, Masaki Mori, Yuichiro Doki, Tetsuji Naka. Gene therapy for esophageal squamous cell carcinoma using SOCS-1 expressing adenoviral vector. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4213. doi:10.1158/1538-7445.AM2015-4213

Adeno-associated virus mediated gene transfer of Shepherdin inhibits gallbladder carcinoma growth in vitro and in vivo

Gene therapy, a significantly crucial strategy for treatment of malignancies, has been gradually accepted in recent years. However, this therapeutic approach has being facing great challenges concerning problems which include complicated development of cancer with multiple gene control, effective target shortage, low efficiency of gene transferring and safety of the vector delivery system. Shepherdin, a novel peptidomimetic molecule designed from Lys-79 to Leu-87 of survivin, has been identified as a tumor suppressor with the function that can not only competitively interfere with the interaction between survivin and Hsp90 (heat shock protein-90) leading to the degradation of survivin to anti-tumor, but also competitively target the ATP-dependent binding pocket of Hsp90 resulting in the dysfunction of Hsp90 chaperone to cell apoptosis via a mitochondrial dependent or independent pathway. In the present study, we designed and constructed a recombinant Adeno-associated virus (rAAV) loading fusion gene NT4–TAT–6His–Shepherdin. The expression of Shepherdin in gallbladder carcinoma (GBC) cells was detected and its strong inhibitory effects against GBC growth were evaluated after AAV mediated gene transfer of Shepherdin into GBC cells and xenograft tumors. MTT assay and flow cytometric analysis demonstrated that rAAV containing Shepherdin gene could significantly inhibit the growth of GBC and this effect was closely associated with apoptosis. These results indicated that rAAV–NT4–TAT–6His–Shepherdin may be considered a novel therapeutic strategy in the gene therapy for gallbladder carcinoma.

Nanoformulation of D-α-tocopheryl polyethylene glycol 1000 succinate-b-poly(ε-caprolactone-ran-glycolide) diblock copolymer for siRNA targeting HIF-1α for nasopharyngeal carcinoma therapy.

Hypoxia-inducible factor-1α (HIF-1α) is a crucial transcription factor that plays an important role in the carcinogenesis and development of nasopharyngeal carcinoma. In this research, a novel biodegradable D-α-tocopheryl polyethylene glycol 1000 succinate-b-poly(ε-caprolactone-ran-glycolide) (TPGS-b-(PCL-ran-PGA)) nanoparticle (NP) was prepared as a delivery system for small interfering ribonucleic acid (siRNA) molecules targeting HIF-1α in nasopharyngeal carcinoma gene therapy. The results showed that the NPs could efficiently deliver siRNA into CNE-2 cells. CNE-2 cells treated with the HIF-1α siRNA-loaded TPGS-b-(PCL-ran-PGA) NPs showed reduction of HIF-1α expression after 48 hours of incubation via real-time reverse transcriptase-polymerase chain reaction and Western blot analysis. The cytotoxic effect on CNE-2 cells was significantly increased by HIF-1α siRNA-loaded NPs when compared with control groups. In a mouse tumor xenograft model, the HIF-1α siRNA-loaded NPs efficiently suppressed tumor growth, and the levels of HIF-1α mRNA and protein were significantly decreased. These results suggest that TPGS-b-(PCL-ran-PGA) NPs could function as a promising genetic material carrier in antitumor therapy, including therapy for nasopharyngeal carcinoma.