Monogene and Polygene Therapy for the Treatment of Experimental Prostate Cancers by Use of Apoptotic Genes bax and bad Driven by the Prostate-Specific Promoter ARR2PB (original) (raw)
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JNCI Journal of the National Cancer Institute, 2001
Background: Adenovirus-mediated overexpression of the apoptosis-inducing protein Bax can induce apoptosis in prostate cancer cell lines. Constitutive overexpression of Bax could result in unwanted apoptosis in every site of accidental Bax accumulation in vivo. Therefore, we developed an adenoviral construct (Av-ARR 2 PB-Bax) in which the probasin promoter, modified to contain two androgen response elements, drives Bax expression. This promoter would be expected to limit expression of Bax to cells expressing the androgen receptor. Methods: A variety of androgen receptor (AR)-positive and-negative cell lines of prostatic or nonprostatic origin were infected with Av-ARR 2 PB-Bax or a control virus, Av-ARR 2 PB-CAT, in which the same promoter drives expression of the chloramphenicol acetyl transferasereporter gene. Bax expression and apoptosis in vitro were assessed by western blot analysis. Tumor size and apoptosis in vivo were assessed after four weekly injections of Av-ARR 2 PB-Bax or Av-ARR 2 PB-CAT into subcutaneous LNCaP xenografts growing in uncastrated male mice. All statistical tests were two-sided. Results: Bax was overexpressed in an androgen-dependent way in AR-positive cell lines of prostatic origin but not in AR-positive cells of nonprostatic origin or in AR-negative cell lines of either prostatic or nonprostatic origin. The androgen dihydrotestosterone activated apoptosis in LNCaP cells infected with Av-ARR 2 PB-Bax but not in those infected with Av-ARR 2 PB-CAT. Av-ARR 2 PB-Bax-injected LNCaP xenograft tumors decreased in tumor size from 34.1 mm 3 (95% confidence interval [CI] = 25.1 mm 3 to 43.1 mm 3) to 24.6 mm 3 (95% CI =-2.5 mm 3 to 51.7 mm 3), but the difference was not statistically significant (P = .5). Tumors injected with Av-ARR 2 PB-CAT increased in size, from 28.9 mm 3 (95% CI = 12.7 mm 3 to 45.1 mm 3) to 206 mm 3 (95% CI = 122 mm 3 to 290 mm 3) (P = .002) and contained statistically significant more apoptotic cells (23.3% [95% CI = 21.1% to 25.6%] versus 9.5% [95% CI = 8.0% to 11.1]) (P<.001). Conclusions: Av-ARR 2 PB-Bax induces androgen-dependent therapeutic apoptosis in vitro and in vivo by activating apoptosis in AR-positive cells derived specifically from prostatic epithelium and does not affect nonprostatic cells. [
Current progress in adenovirus mediated gene therapy for patients with prostate carcinoma
Gene Ther Mol Biol, 2003
Prostate cancer is the most frequently diagnosed male cancer in the world. Like all cancers, prostate cancer is a disease of uncontrolled cell growth. In some cases tumors are slow growing and remain local, but in others they may spread rapidly to the lymph nodes, other organs and especially bone. Although surgery and radiation can cure early stages of organ confined prostate carcinoma (stages I and II), there is no curative therapy at this time for locally advanced or metastatic disease (stages III and IV). The likelihood of postsurgical local recurrence increases with capsular penetration as detected in 30 % of the patients at the time of radical prostatectomy. Moreover, 10-15 % of patients have metastatic cancer at the time of diagnosis. Considering the fact that 60 % local recurrence is observed in patients receiving radiation therapy with or without adjuvant hormonal ablation therapy, it is generally believed that androgen ablation therapy simply delays the progression of prostate carcinoma to a more advanced stage. In addition, the overall ten-year survival rate of patients with locally recurrent prostate cancer is only around 35 %; thus; the ultimate progression into androgen independent prostate carcinoma appears to be inevitable. Gene therapy arose as a novel treatment modality with the potential to decrease the morbidity associated with conventional therapies. Therefore, gene therapy is expected to lower the incidence of tumor recurrence and finally improve the outcome of patients with recurrent and androgen independent prostate carcinoma. Viral vectors are most commonly used for the purpose of gene therapy. Currently, there are a total of 40 clinical trials being conducted using viral vectors for the treatment of prostate carcinoma. 22 out of 40 clinical protocols (55 %) approved for the treatment of prostate cancer utilize adenovirus vectors. Most of these adenovirus mediated therapeutic approaches employ either selectively replicating adenoviruses or suicide gene therapy approaches. In this review, we mainly concentrated on the progress in adenovirus mediated gene therapy approaches for prostate cancer. Analysis of the death ligand mediated gene therapy approach was also discussed in detail, while our novel findings were incorporated as an example for up-to-date approaches used for adenovirus mediated gene therapy against prostate carcinoma.
Oncology Reports, 2012
The biodistribution and safety of adenoviral vectors encoding the human REIC/Dkk-3 tumor suppressor gene (Ad-REIC) were examined in this preclinical study for in situ prostate cancer gene therapy. First, the in vitro apoptotic effects of Ad-REIC in normal and cancer cells derived from the prostate and liver were examined. Significant apoptotic effects were observed at 100 MOI (multiplicity of infection) in prostate cancer cells (LNCaP, PC3) and hepatoma cells (HEP3B and HEPG2); however, no effects were seen in normal cells. To analyze the safety of intraprostatic Ad-REIC administration, the biodistribution and histology after Ad-REIC injection were evaluated in various organs of normal male C57BL6 mice. In a supporting study, vector dissemination following intravenous injection of Ad-REIC into tail veins was determined. To evaluate whether Ad-REIC was present in the collected tissue specimens, human REIC gene detection was performed using DNA-PCR. Intraprostatic treatment administered at lower doses showed vector biodistribution into the colon, urinary bladder and prostate. At higher doses, vector dissemination was observed in tissues more distant from the prostate, including the lung, thymus, heart, liver and adrenal gland. After intravenous injection of Ad-REIC, dissemination was observed in the liver and spleen. These results indicate that the biodistribution of Ad-REIC is determined by the dose and route of administration. Although acute inflammatory effects were observed in the prostate after intraprostatic administration at higher doses, no abnormal histological findings were noted in the other tissues, including those of intravenously treated mice. Regarding the safety of Ad-REIC administration, no deaths and no signs of toxicity or unusual behavior were observed in the mice in any treatment group. Based on these preclinical experiments, adenovirus-mediated in situ REIC/Dkk-3 gene therapy is considered to be safe for use as a treatment for human prostate cancer.
Molecular …, 2006
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 AdEndo-Angio, 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 AdEndo-Angio 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 androgenindependent 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]
Molecular Therapy, 2004
TARP (T cell receptor ;-chain alternate reading frame protein) is a protein that in males is uniquely expressed in prostate epithelial cells and prostate cancer cells. We have previously shown that the transcriptional activity of a chimeric sequence comprising the TARP promoter (TARPp) and the PSA enhancer (PSAe) is strictly controlled by testosterone and highly restricted to cells of prostate origin. Here we report that a chimeric sequence comprising TARPp and the PSMA enhancer (PSMAe) is highly active in testosterone-deprived prostate cancer cells, while a regulatory sequence comprising PSAe, PSMAe, and TARPp (PPT) has high prostate-specific activity both in the presence and in the absence of testosterone. Therefore, the PPT sequence may, in a gene therapy setting, be beneficial to prostate cancer patients that have been treated with androgen withdrawal. A recombinant adenovirus vector with the PPT sequence, shielded from interfering adenoviral sequences by the mouse H19 insulator, yields high and prostatespecific transgene expression both in cell cultures and when prostate cancer, PC-346C, tumors were grown orthotopically in nude mice. Intravenous virus administration reveals both higher activity and higher selectivity for the insulator-shielded PPT sequence than for the immediateearly CMV promoter. Therefore, we believe that an adenovirus with therapeutic gene expression controlled by an insulator-shielded PPT sequence is a promising candidate for gene therapy of prostate cancer.
Molecular Cancer Therapeutics, 2006
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 AdEndo-Angio, 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 AdEndo-Angio 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 androgenindependent 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]
Molecular Therapy, 2005
PSES is a chimeric enhancer containing enhancer elements from prostate-specific antigen (PSA) and prostate-specific membrane antigen (PSMA) genes that are prevalently expressed in androgen-independent prostate cancers. PSES shows strong activity equivalent to cytomegalovirus (CMV) promoter, specifically in PSA/PSMA-positive prostate cancer cells, the major cell types in prostate cancer in the absence of androgen. We developed a recombinant adenovirus (AdE4P-SESE1a) by placing adenoviral E1a and E4 genes under the control of the bidirectional enhancer PSES and enhanced green fluorescent protein gene for the purpose of intratumoral virus tracking under the control of CMV promoter. Because of PSES being very weak in nonprostatic cells, including HEK293 and HER911 that are frequently used to produce recombinant adenovirus, AdE4PSESE1a can only be produced in the HER911E4 cell line which expresses both E1 and E4 genes. AdE4PSESE1a showed similar viral replication and tumor cell killing activities to wild-type adenovirus in PSA/PSMApositive prostate cancer cells. The viral replication and tumor cell killing activities were dramatically attenuated in PSA/ PSMA-negative cells. To test whether AdE4PSESE1a could be used to target prostate tumors in vivo, CWR22rv s.c. tumors were induced in nude mice and treated with AdE4PSESE1a via intratumoral and tail vein injection. Compared to tumors treated with control virus, the growth of CWR22rv tumors was dramatically inhibited by AdE4PSESE1a via tail vein injection or intratumoral injection. These data show that adenoviral replication can be tightly controlled in a novel fashion by controlling adenoviral E1a and E4 genes simultaneously with a single enhancer. (Cancer Res 2005; 65(5): 1941-51) Requests for reprints:
CANCER RESEARCH, 2002
Adenovirus-mediated suicide gene therapy may hold promise in the treatment of human cancer. We have developed a novel approach that utilizes a lytic, replication-competent adenovirus (Ad5-CD/TKrep) to deliver a cytosine deaminase/herpes simplex virus-1 thymidine kinase fusion gene to tumors. The cytosine deaminase and herpes simplex virus-1 thymidine kinase suicide genes render malignant cells sensitive to specific pharmacological agents and, importantly, sensitize them to radiation. The Phase I study described here represents the first gene therapy trial in which a replication-competent virus was used to deliver a therapeutic gene to humans. The indication is local recurrence of prostate cancer after definitive radiation therapy. An escalating dose (10 10 , 10 11 , and 10 12 viral particles) of the Ad5-CD/TKrep virus was injected intraprostatically under transrectal ultrasound guidance into 16 patients in four cohorts. Two days later, patients were given 5-fluorocytosine and ganciclovir prodrug therapy for 1 (cohorts 1-3) or 2 (cohort 4) weeks. There were no doselimiting toxicities, and the maximum tolerated dose of the Ad5-CD/TKrep vector was not defined. Ninety-four percent of the adverse events observed were mild or moderate (grade 1/2) in nature. Seven of 16 (44%) patients demonstrated a >25% decrease in serum prostate-specific antigen, and 3 of 16 (19%) patients demonstrated a >50% decrease in serum prostatespecific antigen. Transgene expression and tumor destruction at the injection site were confirmed by sextant needle biopsy of the prostate at 2 weeks. Two patients were negative for adenocarcinoma at 1 year followup. Although Ad5-CD/TKrep viral DNA could be detected in blood as far out as day 76, no infectious adenovirus was detected in patient serum or urine. Together, the results demonstrate that intraprostatic administration of the replication-competent Ad5-CD/TKrep virus followed by 2 weeks of 5-fluorocytosine and ganciclovir prodrug therapy can be safely applied to humans and is showing signs of biological activity.
Cancer Gene Therapy, 2006
The use of conditionally replicating adenoviruses offers an attractive complementary treatment strategy for localized prostate cancer. We have produced a replicating adenovirus, Ad[I/PPT-E1A], where E1A gene expression is controlled by a recombinant regulatory sequence designated PPT. The PPT sequence comprises a PSA enhancer, a PSMA enhancer and a T-cell receptor g-chain alternate reading frame protein promoter, and it is shielded from transcriptional interference from adenoviral backbone sequences by an H19 insulator. Ad[I/PPT-E1A] yields prostate-specific E1A protein expression, viral replication and cytolysis in vitro. Furthermore, Ad[I/PPT-E1A] considerably regresses the growth of subcutaneous LNCaP prostate cancer tumors in nude mice. Importantly, the viral replication and cytolytic effect of Ad[I/PPT-E1A] are independent of the testosterone levels in the prostate cancer cells. This may be beneficial in a clinical setting since many prostate cancer patients are treated with androgen withdrawal.
Cancer Gene Therapy, 2010
In this study, we investigated the preclinical utility and antitumor efficacy of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) delivered by Ad-IU2, a prostate-specific replication-competent adenovirus (PSRCA), against androgen-independent prostate cancer. Through transcriptional control of adenoviral early genes E1a, E1b and E4, as well as TRAIL by two bidirectional prostate-specific enhancing sequences (PSES), expression of TRAIL as well adenoviral replication was limited to prostate-specific antigen (PSA) and prostate-specific membrane antigen (PSMA)-positive cells. Ad-IU2 induced 5-fold greater apoptosis selectively in PSA/PSMA-positive CWR22rv and C4-2 cells than an oncolytic adenoviral control. Furthermore, prolonged infection with Ad-IU2 reversed TRAIL resistance in LNCaP cells. Ad-IU2 exhibited superior killing efficiency in PSA/PSMA-positive prostate cancer cells at doses 5-to 8-fold lower than required by a PSRCA to produce a similar effect. This cytotoxic effect was not observed in non-prostatic cells, however. As an enhancement of its therapeutic efficacy, Ad-IU2 exerted a TRAIL-mediated bystander effect through direct cell-to-cell contact and soluble factors such as apoptotic bodies. In vivo, Ad-IU2 markedly suppressed the growth of subcutaneous androgenindependent CWR22rv xenografts compared to a PSRCA at six weeks post-treatment (3.1-vs.