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MTS assay of cell viability/proliferation-indicated by mitochondrial activity-after treatment with DOX. The mitochondrial activity in untreated cells was assigned a value of 100. Experiments were performed in quadruplicate. Bars indicate standard deviation.
Source publication
p53 gene transfer has been proposed as a potential therapeutic option for treatment of hepatocellular carcinoma (HCC). Compared to other commonly used gene transfer vectors such as adenovirus and retrovirus, recombinant adeno-associated virus serotype 2 (rAAV2) has shown promising results in human clinical trials. Significant enhancement in the gen...
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Context 1
... DOX is capable of inducing rAAV2 transduction, at DOX = 1 μM, Figure 2D shows a significant cell loss due to DOX's cytotoxicity. We used the cell viability/proliferation MTS assay to quantify the cytotoxicity of DOX, and the results show that the increased cytotoxicity at high DOX doses (Fig. 3) correlated well with the decrease in transgene expression ( Fig. 2A). Specifically, when cells were treated with 5-μM DOX for 12 hours, the viability of HepG2 and Hep3B dropped to 36% and 8% of that of untreated cells, respectively. Meanwhile, the luciferase transgene expression in HepG2 and Hep3B decreased 64% and 99% in comparison ...
Context 2
... the ubiquitous membrane-associated heparan sulfate proteoglycans as primary receptors, rAAV2 is capable of transgene Figure 5. Hep3B viability after treatment with DOX alone or a combined DOX and rAAV2-p53 vectors. MTS assay. The data of Hep3B treated with DOX alone (empty bars) are the same as those shown in (Fig. 3). The mito- chondrial activity in untreated cells was assigned a value of 100. *, #, •, , p, and ◊ indicate P values <0.05. There were no statistical differences between the rAAV2-p53 transduced and untransduced cells and between the rAAV2-p53 transduced and reporter vector-transduced cells at DOX = 0, 0.1, 0.2, 5 and 10 μM. P values ...
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Citations
... Gene therapy has become a promising new treatment strategy for unresectable HCC as hepatocellular carcinogenesis correlates with the activation of oncogenes and dysfunction of anti-oncogenes [108]. Designing a safe and effective transfection vector is the key to gene therapy. ...
Hepatocellular carcinoma (HCC) has the sixth-highest new incidence and fourth-highest mortality worldwide. Transarterial chemoembolization (TACE) is one of the primary treatment strategies for unresectable HCC. However, the therapeutic effect is still unsatisfactory due to the insufficient distribution of antineoplastic drugs in tumor tissues and the worsened post-embolization tumor microenvironment (TME, e.g., hypoxia and reduced pH). Recently, using nanomaterials as a drug delivery platform for TACE therapy of HCC has been a research hotspot. With the development of nanotechnology, multifunctional nanoplatforms have been developed to embolize the tumor vasculature, creating conditions for improving the distribution and bioavailability of drugs in tumor tissues. Currently, the researchers are focusing on functionalizing nanomaterials to achieve high drug loading efficacy, thorough vascular embolization, tumor targeting, controlled sustained release of drugs, and real-time imaging in the TACE process to facilitate precise embolization and enable therapeutic procedures follow-up imaging of tumor lesions. Herein, we summarized the recent advances and applications of functionalized nanomaterials based on TACE against HCC, believing that developing these functionalized nanoplatforms may be a promising approach for improving the TACE therapeutic effect of HCC.
... HCC often develops a high resistance to conventional antineoplastic agents, a non-selective cytotoxic molecule that could result in the systemic adverse effects. The recent advances in gene therapy, i.e., RNA interference (RNAi)-based gene therapy, have been utilized in the current HCC treatment [3,4]. The efficacy of RNAi requires the vector to be delivered to the interior of the target cell [5]. ...
... Viruses offer greater efficiency of gene delivery, but non-viral vectors are preferred due to safety concerns with the viral vectors. Nanoparticles (NP) as non-viral vectors for targeted gene delivery or drug delivery system have gained great attention for improving therapeutic efficiency and lowering toxicity on the systemic and/or cellular levels in HCC treatment [4,6]. Thus, it becomes very important to identify the molecular mechanism and biological pathway underlying cellular disturbance and toxicity of NP in target cells and tissues. ...
Interactions of the 40 and 80 nm gold nanoparticles (AuNP) functionalized with cationic branched polyethylenimine (BPEI), anionic lipoic acid (LA), or neutral polyethylene glycol (PEG) with human hepatocellular carcinoma (HCC) cell line C3A have been investigated in the absence and presence of human plasma protein corona (PC). All bare (no PC) AuNP besides 80 nm LA-AuNP were cytotoxic to C3A but PC attenuated their cytotoxicities. Time-dependent cellular uptake of AuNP increased besides 40 nm BPEI-AuNP but PC suppressed their uptakes besides 80 nm PEG-AuNP. Biphasic responses of oxidative/nitrosative stress by BPEI-AuNP occurred in C3A cells, whereas PEG-AuNP was a potent antioxidant. All bare AuNP inhibited cytochrome P450 (CYP) 3A4 activity irrespective of size and surface charge but PC recuperated its activity besides PEG-AuNP. The 40 nm PEG-AuNP-modulated gene expression was mainly involved in mitochondrial fatty acid β-oxidation and to a less degree hepatic efflux/uptake transporters. These studies contribute to a better understanding of AuNP interaction with key biological processes and their underlying molecular mechanisms in HCC, which may be further implicated in the development of more effective therapeutic target in HCC treatment.
Electronic supplementary material
The online version of this article (10.1186/s11671-018-2684-1) contains supplementary material, which is available to authorized users.
... The broad field of gene delivery holds great potential for the treatment of severe human diseases such as cancers [1,2], cardiovascular diseases [3,4], neurodegenerative disorders [5][6][7], and infectious diseases [8,9] in either preclinical or clinical trials [10][11][12][13][14]. Extensive research have be conducted for the design and synthesis of safe and effective non-viral vectors that could not only strive against viral counterparts in the level of gene expression and specificity, but also provide greater gene size flexibility and reduce or entirely eradicate acquired immune responses. ...
... The broad field of gene delivery holds great potential for the treatment of severe human diseases such as cancers [1,2], cardiovascular diseases [3,4], neurodegenerative disorders [5][6][7], and infectious diseases [8,9] in either preclinical or clinical trials [10][11][12][13][14]. Extensive research have be conducted for the design and synthesis of safe and effective non-viral vectors that could not only strive against viral counterparts in the level of gene expression and specificity, but also provide greater gene size flexibility and reduce or entirely eradicate acquired immune responses. Recently, the co-delivery of anti-cancer agents has been recognized as a promising strategy for the treatment of cancers with remarkable resistance to conventional therapies [12]. Such co-delivery can enhance treatment outcomes and mitigate the adverse side effects of cytotoxic drugs at high doses. ...
... Such co-delivery can enhance treatment outcomes and mitigate the adverse side effects of cytotoxic drugs at high doses. Anticancer drugs such as paclitaxel (PTX) and doxorubicin (Dox) have demonstrated great promise to achieve greater cytotoxic effect via co-delivering with bio-active agents such as therapeutic plasmid DNA (pDNA) [12][13][14] and siRNA [15,16]. However, despite the successful fabrication of core-shell nanoparticles for drug and gene co-delivery [17,18], simultaneous and on-demand release of agents in cancer cells is still one of the main challenges of systemic cancer therapy. ...
Statement of significance:
We employed cystamine dihydrochloride as a disulfide linkage for the conjugation of PCLdiol and low molecular weight PEI segments through a straightforward and efficient reaction pathway at mild conditions. The new copolymer was essentially non-toxic in different carcinoma cell lines and showed great tendency to form positively charged nanoparticles. Therefore, it can be utilized as a promising platform for simultaneous drug and gene delivery to aggressive cancers. The results of drug and gene co-delivery experiments confirmed the pivotal role of disulfide linkage on the controlled release of both drug and gene molecules in response to glutathione concentration gradient between extracellular and intracellular microenvironments. In addition, the co-delivery of doxorubicin and p53 plasmid DNA using the new copolymer displayed greater cytotoxic effect compared with single agent (i.e. Dox) loaded counterpart, which indicated the significance of rapid dissociation of therapeutic agents from the carrier for synergistic cytotoxic effects on cancer cells.
... The p53 protein is an important mediator of various cellular processes, such as modulation of senescence, apoptosis, and cell cycle genes [6,7]. Recent clinical trials have demonstrated the role of p53 siRNA and the efficacy of RNA interference-based drugs in general in anticancer therapy [22,23], although the precise transcriptional mechanisms by which p53 siRNA initiates and supports apoptosis still need to be clarified. The extrinsic signaling pathway of apoptosis is based on the activation of so-called 'death receptors,' whereas the intrinsic mechanism is activated by modifications in DNA structure. ...
Triple-negative breast cancer (TNBC) is a highly aggressive phenotype that is resistant to standard therapy. Thus, the development of alternative therapeutic strategies for TNBC is essential. The purpose of our in vitro study was to evaluate the impact of p53 gene silencing in conjunction with the administration of a natural compound, epigallocatechingallate (EGCG). RT2Profiler PCR Array technology was used to evaluate the impact of dual treatment on the main genes involved in apoptosis in the Hs578T cell culture model of TNBC. Gene expression analysis revealed 28 genes were significantly altered (16 upregulated and 12 downregulated) in response to combined p53 siRNA and EGCG treatment. Further analysis revealed that p53 siRNA and EGCG dual therapy leads to the activation of pro-apoptotic genes and the inhibition of pro-survival genes, autophagy, and cell network formation. These results indicate that this dual therapy targets both the apoptotic and angiogenic pathways, which may improve treatment effectiveness for tumors resistant to conventional treatment.
... In the last 10 years, adenoviral-mediated P53 gene agent has been used in numerous clinical trials for advanced HNSCC worldwide, either alone or in combination with conventional treatments, such as chemotherapy or radiotherapy, and rAd-p53 seems to act synergistically with conventional treatments such as chemotherapy and/ or radiotherapy [12,[14][15][16][17][18][19][20][21][22][77][78][79][80][81][82]87,94,95]. In addition, this apparent synergy still exists in patients who were resistant to chemotherapy and radiotherapy. ...
Introduction:
Wild-type p53 gene is an essential cancer suppressor gene which plays an important role in carcinogenesis and malignant progressions. The p53 gene family participates in almost all the key procedures of cancer biology, such as programmed cell death, angiogenesis, metabolism and epithelial-mesenchymal transition. The mutation or functional defects of the p53 gene family are detected in most of the solid malignant tumors, and the restoration of the p53 gene by adenovirus-mediated gene therapy becomes a promising treatment for cancer patients now.
Areas covered:
In the present review, the potential therapeutic effects of recombinant adenovirus p53 rAd-p53 ( Gendicine ™) were reviewed to explore the biological mechanism underlying the adenovirus-mediated p53 gene therapy. Then, the key points of the drug administration were discussed, including the routes of administration, dosage calculation and treatment cycles, based on findings of the preclinical and clinical trials in order to establish a standard treatment for the p53 gene therapy.
Expert opinion:
As an important part of the combined therapy for the cancer patients, the adenovirus-mediated p53 gene therapy was blossomed to be a promising treatment strategy. A new evaluation criteria and guideline for the gene therapy is urgently needed for the further clinical practice.
... Both gamma ray and UV light can enhance transgene expression and transduction efficiency [11,12]. Moreover, tritiated thymidine [13], some antitumor drugs [14][15][16][17] as well as some DNA-damaging agents have also been considered to increase the efficiency of rAAV transduction in various cells [18,19]. Mitomycin C (MMC) is a potent DNA crosslinker that alkylates double strand DNA [20] and has been widely used as a chemotherapeutic agent [21]. ...
The effect of chemotherapy drug Mitomycin C (MMC) in combination with recombinant adeno-associated virus II (rAAV2) in cancer therapy was investigated, and the mechanism of MMC affecting rAAV2's bioactivity was also studied. The combination effect was evaluated by the level of GFP and TNF expression in a human glioma cell line, and the mechanism of MMC effects on rAAV mediated gene expression was investigated by AAV transduction related signal molecules. C57 and BALB/c nude mice were injected with rAAV-EGFP or rAAV-TNF alone, or mixed with MMC, to evaluate the effect of MMC on AAV-mediated gene expression and tumor suppression. MMC was shown to improve the infection activity of rAAV2 both in vitro and in vivo. Enhancement was found to be independent of initial rAAV2 receptor binding stage or subsequent second-strand synthesis of target DNA, but was related to cell cycle retardation followed by blocked genome degradation. In vivo injection of MMC combined with rAAV2 into the tumors of the animals resulted in significant suppression of tumor growth. It was thus demonstrated for the first time that MMC could enhance the expression level of the target gene mediated by rAAV2. The combination of rAAV2 and MMC may be a promising strategy in cancer therapy.
... 6,7 Of these, combined therapy has proven the best solution to date. [8][9][10] A previous study 11 successfully combined p53 gene therapy, embolic therapy, and nanotherapy at the liver tumor site by exploiting poly-L-lysine (PLL)-modified hydroxyapatite nanoparticles (nHAPs) to serve as embolic material and gene vector at the same time. However, complete tumor elimination was not observed in any of the animals and the survival prolongation was limited, thus further improvements to the therapy are necessary before clinical application. ...
Background/aim
A local nanotherapy (LNT) combining the therapeutic efficacy of trans-arterial embolization, nanoparticles, and p53 gene therapy has been previously presented. The study presented here aimed to further improve the incomplete tumor eradication and limited survival enhancement and to elucidate the molecular mechanism of the LNT.
Methods
In a tumor-targeting manner, recombinant expressing plasmids harboring wild-type p53 and Rb were either co-transferred or transferred separately to rabbit hepatic VX2 tumors in a poly-L-lysine-modified hydroxyapatite nanoparticle nanoplex and Lipiodol® (Guerbet, Villepinte, France) emulsion via the hepatic artery. Subsequent co-expression of p53 and Rb proteins within the treated tumors was investigated by Western blotting and in situ analysis by laser-scanning confocal microscopy. The therapeutic effect was evaluated by the tumor growth velocity, apoptosis and necrosis rates, their sensitivity to Adriamycin® (ADM), mitomycin C, and fluorouracil, the microvessel density of tumor tissue, and the survival time of animals. Eventually, real-time polymerase chain reaction and enhanced chemiluminescence Western blotting were used to investigate the expressive changes of important genes related to the therapy.
Results
The administration procedure proved safe for the rabbits’ liver function, the p53 plus Rb LNT showed significantly better antitumoral effect and lower expression of malignant genes than the p53 or Rb LNT, although no significant difference was observed in animal survival when the p53 plus Rb LNT was compared with the p53 LNT.
Conclusion
Rb works synergistically with p53 in combined therapy mediated by a poly-L-lysine-modified hydroxyapatite nanoparticle nanoplex to augment the antitumoral effect through the downregulated expression of important genes related to apoptosis, necrosis, growth, differentiation and multidrug resistance of tumor cells. LNT with p53 and Rb is potentially an effective antitumor therapy for hepatocellular carcinoma.
... Por otra parte, en un modelo de retinoblastoma en ratas, la expresión de IFN-β (interferón beta) utilizando VAA2 a una dosis de 1 x 10 9 pv/mL administrados por vía intravitreal, demostró efectos anti-tumorales. 37,38 ...
Viral vectors based on adeno-associated virus (AAV) are widely used in gene therapy protocols, because they have characteristics that make them valuable for the treatment of genetic and chronic degenerative diseases. AAV2 serotype had been the best characterized to date. However, the AAV vectors developed from other serotypes is of special interest, since they have organ-specific tropism which increases their potential for transgene delivery to target cells for performing their therapeutic effects. This article summarizes AAV generalities, methods for their production and purification. It also discusses the use of these vectors in vitro, in vivo and their application in gene therapy clinical trials.
... However, an obstacle to these applications is low transgene expression efficiency, mainly due to a limited second strand synthesis [13,14]. Previous studies reported that DNAdamaging agents, such as UV light, gamma irradiation, cis-platinum, and tritiated thymidine can significantly increase the efficiency of AAV transduction in various cells, including nondividing cells [15], airway cells [16], neuronal cells [17] and cancer cells [12,18,19]. Cisplatin is an alkylating agent that targets DNA and results in bulky adducts as well as intra-and inter-strand crosslink [20,21]. ...
Adeno-associated virus-2 (AAV-2)-mediated gene therapy is quite suitable for local or regional application in head and neck cancer squamous cell carcinoma (HNSCC). However, its low transduction efficiency has limited its further development as a therapeutic agent. DNA damaging agents have been shown to enhance AAV-mediated transgene expression. Cisplatin, one of the most effective chemotherapeutic agents, has been recognized to cause cancer cell death by apoptosis with a severe toxicity. This study aims to evaluate the role of cisplatin in AAV-mediated tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) expression and the effect on HNSCC both in vitro and in vivo.
Five human HNSCC cell lines were treated with recombinant soluble TRAIL (rsTRAIL) and infected with AAV/TRAIL to estimate the sensitivity of the cancer cells to TRAIL-induced cytotoxicity. KB cells were infected with AAV/EGFP with or without cisplatin pretreatment to evaluate the effect of cisplatin on AAV-mediated gene expression. TRAIL expression was detected by ELISA and Western blot. Cytotoxicity was measured by MTT assay and Western blot analysis for caspase-3 and -8 activations. Following the in vitro experiments, TRAIL expression and its tumoricidal activity were analyzed in nude mice with subcutaneous xenografts of HNSCC.
HNSCC cell lines showed different sensitivities to rsTRAIL, and KB cells possessed both highest transduction efficacy of AAV and sensitivity to TRAIL among five cell lines. Preincubation of KB cells with subtherapeutic dosage of cisplatin significantly augmented AAV-mediated transgene expression in a heparin sulfate proteoglycan (HSPG)-dependent manner. Furthermore, cisplatin enhanced the killing efficacy of AAV/TRAIL by 3-fold on KB cell line. The AAV mediated TRAIL expression was observed in the xenografted tumors and significantly enhanced by cisplatin. AAV/TRAIL suppressed the tumors growth and cisplatin augmented the tumoricidal activity by two-fold. Furthermore, Combination treatment reduced cisplatin-caused body weight loss in nude mice.
The combination of AAV-mediated TRAIL gene expression and cisplatin had synergistic therapeutic effects on head and neck cancers and reduced the potential toxicity of cisplatin. These findings suggest that the combination of AAV/TRAIL and cisplatin may be a promising strategy for HNSCC therapy.
Hepatocellular carcinoma (HCC) is the most common form of primary liver cancer with high incidence globally. Increasing mortality and morbidity rates combined with limited treatment options available for advanced HCC press for novel and effective treatment modalities. Gene therapy represents one of the most promising therapeutic options. With the recent approval of herpes simplex virus for advanced melanoma, the field of gene therapy has received a major boost. Adeno-associated virus (AAV) is among the most widely used and effective viral vectors today with safety and efficacy demonstrated in a number of human clinical trials. This review identifies the obstacles for effective AAV based gene delivery to HCC which primarily include host immune responses and off-target effects. These drawbacks could be more pronounced for HCC because of the underlying liver dysfunction in most of the patients. We discuss approaches that could be adopted to tackle these shortcomings and manufacture HCC-targeted vectors. The combination of transductional targeting by modifying the vector capsid and transcriptional targeting using HCC-specific promoters has the potential to produce vectors which can specifically seek HCC and deliver therapeutic gene without significant side effects. Finally, the identification of novel HCC-specific ligands and promoters should facilitate and expedite this process.
