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Adenovirus vectors have a number of advantages for gene therapy. However, because of their lack of tumor tropism and their preference for liver infection following systemic administration, they cannot be used for systemic attack on metastatic disease. Many epithelial tumors (e.g., colon, lung, and breast) express carcinoembryonic antigen (CEA). To block the natural hepatic tropism of adenovirus and to “retarget” the virus to CEA-expressing tumors, we used a bispecific adapter protein (sCAR-MFE), which fuses the ectodomain of the coxsackie/adenovirus receptor (sCAR) with a single-chain anti-CEA antibody (MFE-23). sCAR-MFE untargets adenovirus-directed luciferase transgene expression in the liver by >90% following systemic vector administration. Moreover, sCAR-MFE can “retarget” adenovirus to CEA-positive epithelial tumor cells in cell culture, in s.c. tumor grafts, and in hepatic tumor grafts. The sCAR-MFE bispecific adapter should, therefore, be a powerful agent to retarget adenovirus vectors to epithelial tumor metastases. [Cancer Res 2007;67(11):5354–61]

e13551 Background: To evaluate the feasibility of adding p53 gene agent to the current standard treatment of NPC cases. Methods: Gendicine was given intravenously at a dose of 2×1012VP/day on 5 con...

Although prostate-restricted replicative adenovirus has exhibited significant antitumor efficacy in preclinical studies, it is necessary to develop more potent adenoviruses for prostate cancer gene therapy. We evaluated the synergistic killing effect of prostate-restricted replicative adenovirus and AdEndoAngio, a replication-defective adenovirus expressing the endostatin-angiostatin fusion protein (EndoAngio). When coadministered with AdEndoAngio, prostate-restricted replicative adenovirus significantly elevated EndoAngio expression, suggesting that AdEndoAngio coreplicates with prostate-restricted replicative adenovirus. Conditioned medium from prostate cancer cells infected by prostate-restricted replicative adenovirus plus AdEndoAngio inhibited the growth, tubular network formation, and migration of human umbilical vein endothelial cells better than conditioned medium from prostate cancer cells infected by AdEndoAngio alone. Furthermore, in vivo animal studies showed that the coadministration of prostate-restricted replicative adenovirus plus AdEndoAngio resulted in the complete regression of seven out of eight treated androgen-independent CWR22rv tumors, with a tumor nodule maintaining a small size for 14 weeks. The residual single tumor exhibited extreme pathologic features together with more endostatin-reactive antibody-labeled tumor cells and fewer CD31-reactive antibody-labeled capillaries than the AdEndoAngio-treated tumors. These results show that combination therapy using prostate-restricted replicative adenovirus together with antiangiogenic therapy has more potent antitumor effects and advantages than single prostate-restricted replicative adenovirus and deserves more extensive investigation. [Mol Cancer Ther 2006;5(3):676–84]

Transcatheter arterial chemoembolisation (TACE) has become the standard treatment for unresectable hepatocellular carcinoma (HCC). However, this method is often unsuccessful. The p53 gene, which is present as a mutant form in many human tumours, is known to have broad spectrum antitumour effects when expressed normally. In this study, we report a 23 year old patient with recurrent HCC who was treated with the p53 gene (Gendicine) combining TACE, which resulted in a good clinical prognosis.

Replication-competent adenovirus-mediated suicide gene therapy has proven to be safe in humans when delivered intraprostatically. Although signs of efficacy are emerging, it is likely that further improvements will be needed before this technology will have widespread applicability in the clinic. Toward this end, we have developed a second-generation, replication-competent adenovirus (Ad5-yCD/mutTK(SR39)rep-ADP) containing an improved yeast cytosine deaminase (yCD)/mutant(SR39) herpes simplex virus thymidine kinase fusion (yCD/mutTK(SR39)) gene and the adenovirus death protein (ADP) gene. Relative to the first-generation Ad5-CD/TKrep adenovirus, Ad5-yCD/mutTK(SR39)rep-ADP demonstrated greater tumor cell kill in vitro and significantly greater tumor control in preclinical models of human cancer. Quantification of transgene volume following direct injection of fadenovirus into human tumor xenografts and the naïve canine prostate demonstrated that ADP enhanced adenoviral spread in vivo. Toxicology studies were performed to determine whether the improved yCD/mutTK(SR39) fusion and ADP genes increased toxicity. Intraprostatic injection of Ad5-yCD/mutTK(SR39)rep-ADP did not result in significantly increased toxicity relative to the parental Ad5-CD/TKrep adenovirus, the latter of which has proven to be safe in two Phase I prostate cancer clinical trials. Together, these results provide the scientific basis for evaluating the safety and efficacy of the second-generation Ad5-yCD/mutTK(SR39)rep-ADP adenovirus in humans.

Objective To test the oncolytic activity of cyclo-oxygenase 2 (COX-2) promoter–based conditional replication-selective adenovirus vector for squamous cell carcinoma cells of the head and neck. Design In vitro study. Subjects None. Interventions A conditional replication-selective adenovirus vector in which the expression of E1a, required for viral replication, is controlled by the COX-2 promoter, Ad-COX2-E1a, was generated. Its oncolytic activity according to the levels of COX-2 and of Coxsackie and adenovirus receptor expression was tested in a series of human head and neck squamous cell carcinoma cell lines. Results The respective COX-2 messenger RNA expression ratios of KB, H891, T891, T892, and L871 were 1.5, 60.0, 1.0, 14.6, and 1.3. The corresponding Coxsackie and adenovirus receptor messenger RNA expression ratios were 1, 1, 5, 3, and 1. In vitro assays showed significant growth suppression of cancer cell lines with strong expressions of COX-2. Conclusion This study demonstrated the possibility of oncolytic therapy using the COX-2 promoter–based conditional replication-selective adenovirus for head and neck squamous cell carcinoma expressing COX-2.

The ability to repair or enhance an individual's genetic make-up provides the sublime opportunity to ameliorate or eliminate many clinical disorders that affect mankind. Gene therapy is thus a revolutionary clinical strategy that may potentially treat an array of genetic and non-genetic diseases, as well as a novel method for drug delivery and vaccination. To these ends, adenovirus vectors are currently the most promising means to deliver specific genes of interest into target cells of the patient. A major limitation of the use of adenovirus vectors in clinical trials, however, is the rapidly induced inflammatory response against these infectious particles. This review aims to describe the cellular and molecular mechanisms involved in the adenovirus-mediated inflammatory response.

For cancer gene therapy, replicative adenovirus is a promising vector to overcome low infectivity and poor gene delivery of nonreplicative adenovirus in vivo, but its therapeutic efficacy is still unsatisfactory because of the limited spread of replicative virus in a solid tumor. Therefore, the combined therapy with other antitumor agents may be necessary. Nonreplicative adenovirus expressing a therapeutic gene may be a promising candidate because E1 proteins expressed by replicative adenovirus would render nonreplicative adenovirus replicative, augmenting a transgene expression. In this study, we first found that mouse hepatoma Hepa 1-6 cells were permissive for the replication and cytopathic effect of human adenovirus, which enabled us to examine the potential of combined replicative adenovirus and nonreplicative adenovirus expressing an immunostimulator in an immunocompetent mouse-syngeneic Hepa 1-6 tumor model. Nonreplicative adenovirus expressing interleukin-12 (AdIL-12) was used as a model. In vitro coinfection of two adenoviruses produced higher concentrations of IL-12 than infection of AdIL-12 alone in this cell line. In vivo experiments with Hepa 1-6 tumors in syngeneic immunocompetent C57BL/6 mice showed higher concentrations of serum IL-12 and greater therapeutic efficacy in the combination therapy than infection of either adenovirus. These data indicate that the combination of replicative adenovirus and nonreplicative adenovirus expressing an immunostimulator appears to be very efficacious for cancer gene therapy.

e21514 Background: To evaluate the efficacy and safety for the treatment of advanced soft tissue sarcoma with recombinant human adenovirus p53 (rAd-p53, Gendicine) injection and radiotherapy. Metho...

Gene therapy using replication-competent adenovirus that selectively propagates in tumor cells may be an effective treatment for cancer. We developed an adenovirus that would be replication specific for hepatocellular carcinoma (HCC). Based on our finding that the E1B55k-deficient adenovirus was able to replicate in human primary hepatocytes, we therefore designed an adenovirus carrying E1A and attenuated E1B gene driven by the alpha-fetoprotein promoter (Adv-AFP-E1AdB), thus restricting the replication specificity in AFP-producing HCC. Replication of Adv-AFP-E1AdB in primary hepatocytes was practically negligible 4 days after infection. Although Adv-AFP-E1AdB replicated slowly in AFP-producing HCC, it efficiently destroyed HCC cells independent of their p53 status. Experiments were conducted in vivo using systemic administration of Adv-AFP-E1AdB and we observed tumor size reduction in nude mice having liver cancer. The use of replication-competent adenovirus deficient of the E1B gene coupled to an AFP-targeting strategy may be a safe and efficacious treatment for HCC.

Genes are the basis of much human disease, whether they are the defective genes involved in a hereditary disease, the genes of a pathogen dedicated to taking advantage of a host’s physiology for their own ends, or the wayward genes of a cancer promoting uncontrolled cell division. The rapid advances in genetics since the early 1970s, notably the development of recombinant DNA techniques, have allowed the isolation and manipulation of genetic material and permitted the identification of some of the various genes involved in human disease.

Recombinant adenovirus vectors are efficient at transferring genes into somatic tissues but are limited for use in clinical gene therapy by immunologic factors that result in the rapid loss of gene expression and inhibit secondary gene transfer. This study demonstrates that systemic coadministration of recombinant adenovirus with soluble CTLA4Ig, which is known to block co-stimulatory signals between T cells and antigen presenting cells, leads to persistent adenoviral gene expression in mice without long-term immunosuppression. This form of immunotherapy greatly enhances the likelihood that recombinant adenovirus vectors will be useful for human gene therapy.

Remarkable progress has been made in the field of gene therapy during the last few years. Genetic as well as acquired diseases such as cancer could potentially be treated by this approach. Among the available vectors for gene transfer, replication-deficient recombinant adenoviruses (Ad) appear attractive. They are easy to manipulate, infect many different types of cells (even nondividing ones), can be prepared at high titers, and, of particular importance, they are capable of direct in vivo introduction of foreign genetic material into the body.

Publisher Summary This chapter reviews adenovirus-based expression vectors and recombinant vaccines. A powerful new approach to the development of vaccines is the use of microorganisms as engineered vectors for expression of genes from heterologous species. Such vectors are used in two major ways, that is, as high-level expression systems for production of antigens, or as live recombinant vaccines. It seems likely that different vectors have relative advantages and disadvantages for different applications and the development of several systems in parallel seems worthwhile. The chapter describes some of the merits and potential problems associated with the use of adenoviruses as expression vectors for production of proteins in mammalian cells, and as recombinant vaccines for immunization of humans and animals against disease. From all of the studies performed to date, human adenoviruses have considerable potential as expression vectors and live-virus vaccines for a wide variety of human and animal diseases. It has been suggested that a well-characterized adenovirus serotype, such as human adenovirus type 5, is suited to the development of oral vaccines for use in children.

CHINESE GENE THERAPYAn English-language review of Chinese gene-therapy clinical trial results has been cited at least a dozen times by experts as definitive, but some say the findings are hard to evaluate, and others question the quality of the data. ([Read more][1].) [1]: http://www.sciencemag.org/cgi/content/full/314/5803/1233

The study was to evaluate the safety and the efficacy of recombinant adenovirus-p53 (Adp53, trademarked as Gendicine) combined with radiotherapy in nasopharyngeal carcinoma (NPC) patients. A controlled clinical trial on Gendicine combined with radiotherapy, that is, gene therapy + radiotherapy (GTRT), was conducted in 24 patients, and 25 patients treated with radiotherapy alone (RT) formed the control group. In the GTRT group, Gendicine 1|[times]|1012 virus particle (vp) was intratumorally injected once a week over to eight weeks, and concurrent irradiation was given. For both groups, the conventional fractionation 2Gy/day, five fractions a week, with a total dose of 70Gy in 35 fractions were given to either primary tumor or neck lymph node. Patients were monitored for adverse event and serum anti-adenoviral antibody, and tumors were monitored for response. A comparative study was performed between both groups on the immediate response rate by CT at the end of 4th week, 7th week and 2 months after the treatment. The median follow-up interval of surviving patients was 34.5 months (24|[sim]|45 months). Wild-type p53 gene therapy enhanced significantly radiotherapy efficacy by 1.59 times in patients with NPC at 4- week time point. Two months after the treatment complete response rate of GTRT group was 2.1 times that of RT group (62.5% v.s. 29.6%). The 3-year overall survival rate of GTRT group was 14.4% higher than that of GT group. No dose-limiting toxicity and adverse events were noted, except for transient fever after Gendicine administration. Intratumoral injection with Gendicine was safe and effective for patients with NPC.

Top of pageAbstract Purpose: To evaluate the safety and curative effect of selective intraarterial rAd-p53 infusion combined with chemotherapy for locally advanced HNC, 23 patients were enrolled into a prospective clinical phase II study. Materials and Method: Twenty-three patients with locoregionally advanced SCC or ACC were subdivided into three groups. All patients underwent unilateral or bilateral retrograde selective catheterization of ECA. Through the arterial pumps, rAd-p53 combined with chemotherapeutic agents was administrated to group I (7 patients), rAd-p53 was administrated to group II (6 patients), and chemotherapeutic agents were administrated to group III Results: The complete response (CR) rate of group I (28.57%) was higher than that of group II (16.67%) and group III (10%). No replication-deficient virus was detected in patients' serum, urine or sputum at any assay. Comparing with group III, the clinical toxic syndromes and complications in the combination therapy group were significantly decreased vs. that of chemotherapy group for both the number of cases and the extent (P

Transcatheter arterial chemoembolization (TACE) has become the standard treatment for unresectable hepatocellular carcinoma (HCC). But this method has some shortages. p53 gene, which was found to be mutant in many human tumors, has been proved with broad-spectrum anti-tumor effects. We reported a 23-year-old patient with recurrent HCC after irregular hepatectomy. The p53 gene was applied to this patient. We injected percutaneously and infused transcatheterally p53 gene (Gendicine, Shenzhen Sibiono Bentech, China) into his recurrent nodules in liver respectively and 4 d later, the patient received TACE therapy. In the 2 mo follow-up, the patient was in good clinical condition with normal liver function and no recurrence was identified. The case report proposed that recurrent HCC could be successfully treated with p53 gene therapy combining TACE.

The use of adenovirus vector for cancer gene therapy is limited by their low transduction efficiency for lymphoma, leukemia and myeloma. We previously reported that highly efficient gene delivery of CD40-ligand by a modified adenovirus vector with the integrin-binding motif, RGD, in the H1 loop of the fiber knob (AxCAhCD40L-F/RGD) could induce phenotypic alteration followed by T cell immune response to autologous leukemia cells. But the utility of adenovirus with RGD-motif is still limited by their lack of specificity on tumor cells. Recent studies revealed a novel strategy of targeting adenovirus using a bispecific single-chain antibody (scFv) specific for adenovirus and target molecules on tumor cell surface. However, this approach should permit the production of high quantities of active bispecific scFv for in vivo use. To target adenovirus to hematopoietic tumor cells efficiently, we herein constructed a modified adenovirus vector that contained a synthetic immunoglobulin G-binding domain (termed Z33) in H1 loop of the fiber knob. A recombinant adenovirus encoding EGFP, lacZ (as reporter gene; Ax3CAZ3-F/Z33 or Ax3EGFP-F/Z33) or CD40L (as a therapeutic gene; Ax3CD40L-F/Z33) with Z33-modified fiber were tested for gene transfer efficiency into human tumors such as lymphoma, leukemia and myeloma cells. By the treatment with various antibodies specific for CD20 (Rituximab), CD40, CD38, CCR2 or CXCR4 that are expressed on leukemic cells, we achieved 3 to 10-fold enhancement of gene expression in lymphoma/leukemia (Ramos, Daudi or THP-1) and myeloma cells (MM1S, IM-9 or KMS5) compared with control IgG-treated tumors. We also examined specific gene delivery to freshly isolated leukemia B cells from patients that also contains normal lymphocytes. By using antibody to CD20 or CD40, we could selectively deliver CD40L gene with Ax3CD40L-F/Z33 into leukemia B cells (>50% at 300 pu/cell), but not in T and monocytes, followed by the induction of immune costimulatory molecules that are important in anti-leukemia immune response. Overall, our results indicated that combination of Z33-modified adenovirus vector and tumor specific antibody can be used as a modality for the gene therapy of leukemia and myeloma.

The present invention relates to novel adenovirus vectors for use in gene therapy which are designed to prevent the generation of replication-competent adenovirus (RCA) during in vitro propagation and clinical use. The invention also provides methods for the production of the novel virus vectors. These vectors maximize safety for clinical applications in which adenovirus vectors are used to transfer genes into recipient cells for gene therapy.

Viable recombinant adenoviruses that carry a portion of the type 12 E1A and E1B transcription units in a type 5 background were used to identify genes controlling expression of the adenovirus tumor-specific transplantation antigen (TSTA). The TSTA immunity is not crossreacting between the group A and group C adenovirus serotypes. Viruses carrying the E1A region (sub370-12E1A), or both E1A and E1B (sub370-12E1AB) regions of Ad12, induce a strong transplantation immunity against tumors induced by syngeneic cells transformed with adenovirus 12, but fail to induce any protection against syngeneic cells transformed with adenovirus 2. Immunization with the virus carrying only the E1B region (sub370-12E1B) of adenovirus 12 induces no immunity to adenovirus 12 transformed cell line, but confers a strong protection against cells transformed with adenovirus 2. These results provide strong evidence that the adenovirus tumor-specific transplantation antigen is a function of the E1A early region.