Antisense Bcl-2 Oligonucleotide Research Articles

A Phase I Pharmacokinetic and Pharmacodynamic Correlative Study of the Antisense Bcl-2 Oligonucleotide G3139, in Combination with Carboplatin and Paclitaxel, in Patients with Advanced Solid Tumors

This phase I trial assessed the safety and tolerability of G3139 when given in combination with carboplatin and paclitaxel chemotherapy. The effect of G3139 treatment on Bcl-2 expression in peripheral blood mononuclear cells (PBMC) and paired tumor biopsies was also determined. Patients with advanced solid malignancies received various doses of G3139 (continuous i.v. infusion days 1-7), carboplatin (day 4), and paclitaxel (day 4), repeated in 3-week cycles, in a standard cohort-of-three dose-escalation schema. Changes in Bcl-2/Bax transcription/expression were assessed at baseline and day 4 (prechemotherapy) in both PBMCs and paired tumor biopsies. The pharmacokinetic interactions between G3139 and carboplatin/paclitaxel were measured. Forty-two patients were evaluable for safety analysis. Primary toxicities were hematologic (myelosuppression and thrombocytopenia). Dose escalation was stopped with G3139 at 7 mg/kg/d, carboplatin at area under the curve of 6, and paclitaxel at 175 mg/m(2) due to significant neutropenia seen in cycle 1 and safety concerns in further escalating chemotherapy in this phase I population. With G3139 at 7 mg/kg/d, 13 patients underwent planned tumor biopsies, of which 12 matched pairs were obtained. Quantitative increases in intratumoral G3139 with decreases in intratumoral Bcl-2 gene expression were seen. This paralleled a decrease in Bcl-2 protein expression observed in PBMCs. Although the maximal tolerated dose was not reached, the observed toxicities were consistent with what one would expect from carboplatin and paclitaxel alone. In addition, we show that achievable intratumoral G3139 concentrations can result in Bcl-2 down-regulation in solid tumors and PBMCs.

A phase I study of antisense Bcl-2 oligonucleotide (G3139) in combination with carboplatin (C) and paclitaxel (T) in patients with advanced solid tumors

13006 Background: G3139 is an 18-mer oligonucleotide that targets the mRNA of Bcl-2, a protein that inhibits apoptosis conferring treatment resistance, and is synergistic with taxanes in studies using Bcl-2 over-expressing xenografts. Here we conduct a study combining G3139 with C + T in order to define the recommended phase II dose, as well as evaluate the direct biologic effect of G3139 on tumor. Methods: Patients with advanced solid tumors were treated with a dose escalation of G3139 (3–7 mg/kg/day) by continuous infusion days 1–7, with C (AUC 5) and T (150 mg/m2) given on day 4, repeated in 3 week cycles. At the planned expanded cohort (G3139 7 mg/kg/day dose), 12 patients underwent paired tumor biopsies (baseline and day 4 pre-chemotherapy) for assessment of Bcl-2/Bax expression by RT-PCR and IHC, as well as intratumoral G3139 levels using ELISA. Likewise, peripheral blood mononuclear cells (PBLs) were assessed using flow cytometry and RT-PCR for changes in Bcl-2/Bax expression. Results: 34 patients have been treated: 11 melanoma, 7 bladder, 3 prostate, 2 esophageal, 3 non-small cell lung, and 1 head/neck, kidney, breast, pancreas, liver, gastrointestinal stromal, biliary, and unknown primary cancers. G3139/C/T appeared well tolerated at the doses administered with the main toxicities observed being attributed to the C + T chemotherapy (myelosuppression and thrombocytopenia). The maximal tolerated dose is yet to be achieved. Assessment of paired tumor biopsies on 12 patients show decreases in Bcl-2 transcription in laser capture microdissected tumor using RT-PCR, with concordance observed using IHC. Likewise, decrease in Bcl-2 gene expression in PBLs was seen. 2 confirmed PRs were observed, including a bladder patient with baseline Bcl-2 tumor expression 10-fold greater than any other patient assessed. Conclusions: G3139/C/T is well tolerated, with main toxicities attributable to the chemotherapy alone. Bcl-2 is suppressed in tumor and PBLs supporting that a biologically active dose of G3139 is being achieved. Currently, the C + T is being escalated to define a MTD. Complete clinical data, G3139 pharmacokinetics, and Bcl-2/Bax data will be presented in detail. No significant financial relationships to disclose.

B cell translocation gene 1 contributes to antisense Bcl-2-mediated apoptosis in breast cancer cells

The antiapoptotic protein Bcl-2 is overexpressed in a majority of breast cancers, and is associated with a diminished apoptotic response and resistance to various antitumor agents. Bcl-2 inhibition is currently being explored as a possible strategy for sensitizing breast cancer cells to standard chemotherapeutic agents. Antisense Bcl-2 oligonucleotides represent one method for blocking the antiapoptotic effects of Bcl-2. In this study, we show that antisense Bcl-2 efficiently blocks Bcl-2 expression, resulting in the apoptosis of breast cancer cells. Antisense Bcl-2-mediated cytotoxicity was associated with the induction of the B cell translocation gene 1 (BTG1). Importantly, knockdown of BTG1 reduced antisense Bcl-2-mediated cytotoxicity in breast cancer cells. Furthermore, BTG1 expression seems to be negatively regulated by Bcl-2, and exogenous expression of BTG1 induced apoptosis. These results suggest that BTG1 is a Bcl-2-regulated mediator of apoptosis in breast cancer cells, and that its induction contributes to antisense Bcl-2-mediated cytotoxic effects.

Targeting Apoptosis Pathways in Cancer Therapy

Apoptosis, or programmed cell death, is a mechanism by which cells undergo death to control cell proliferation or in response to DNA damage. The understanding of apoptosis has provided the basis for novel targeted therapies that can induce death in cancer cells or sensitize them to established cytotoxic agents and radiation therapy. These novel agents include those targeting the extrinsic pathway such as tumor necrosis factor-related apoptosis-inducing ligand receptor 1, and those targeting the intrinsic Bcl-2 family pathway such as antisense bcl-2 oligonucleotides. Many pathways and proteins control the apoptosis machinery. Examples include p53, the nuclear factor kappa B, the phosphatidylinositol 3 kinase pathway, and the ubiquitin/proteosome pathway. These can be targeted by specific modulators such as bortezomib, and mammalian target of rapamycin inhibitors such as CCI-779 and RAD 001. Because these pathways may be preferentially altered in tumor cells, there is potential for a selective effect in tumors sparing normal tissue. This article reviews the current understanding of the apoptotic pathways, including the extrinsic (cytoplasmic) and intrinsic (mitochondrial) pathways, and the agents being developed to target these pathways.

APOPTOSIS AND CHEMOTHERAPY FOR BLADDER CANCER

We discuss the role of apoptosis, that is gene directed self-destruction of a cell, in the response of bladder transitional cell carcinoma cells to chemotherapy. A directed MEDLINE literature search of apoptosis, bladder cancer and chemotherapy was performed to extract relevant information for review. The characteristics of apoptotic cells were defined and the methods in common use to detect these traits is described. The role of the key mediators of the apoptotic process in bladder cancer is discussed in the context of chemosensitivity and disease stage. The importance of the apoptosis induction after chemotherapy is highlighted. On stimulus by appropriate external or internal signals a cell may alter the expression of genes encoding for proteins associated with the apoptotic process. The development of apoptosis depends on the balance between pro-apoptotic and anti-apoptotic proteins. Key alterations in genes and proteins related to apoptosis within bladder cancer result in a shift away from the default state of apoptosis toward a cell with increased survival properties that is chemoresistant. Much current research in bladder cancer is aimed at restoring chemosensitivity by shifting the cell toward a pro-apoptotic phenotype. Successful translation of this work into clinical practice may improve survival in patients in whom prognosis is currently poor.

Enhanced efficacy of therapy with antisense BCL-2 oligonucleotides plus anti-CD20 monoclonal antibody in scid mouse/human lymphoma xenografts

Abstract Monoclonal anti-CD20 antibody (rituximab) is active, but not curative, therapy for B-cell non-Hodgkin's lymphoma. BCL-2 is an antiapoptotic protein whose expression is dysregulated in most indolent B-cell malignancies. Antisense oligonucleotides (AS-ODNs) that down-regulate BCL-2 expression induce apoptosis and chemosensitize B-cell lymphoma cells. We hypothesized that BCL-2 down-regulation by AS-ODNs would sensitize cells to rituximab and improve therapeutic results. There is enhanced apoptosis and reduction in cell numbers when DoHH2 cells are treated in vitro with rituximab plus BCL-2 AS-ODNs, compared with either agent alone. There is little in vitro effect on WSU-FSCCL cells by rituximab, AS-ODNs that down-regulate BCL-2 by targeting the immunoglobulin portions of the BCL-2-immunoglobulin fusion molecule, or a combination of the two. The combination is more effective than either agent alone in clearing DoHH2 cells from ascites in scid mice. Combination therapy with AS-BCL-2-ODNs and rituximab significantly prolongs survival in both the DoHH2 and WSU-FSCCL models. With higher and repeated doses, this combination could be curative. We conclude that the combination of rituximab and antisense-mediated down-regulation of BCL-2 has enhanced activity against human lymphoma, prolongs survival, and could cure mice bearing human lymphoma. This merits investigation in clinical trials.

Bcl-2 Controls Caspase Activation Following a p53-dependent Cyclin D1-induced Death Signal

MCF-7 and ZR-75 breast cancer cells infected with an adenovirus constitutively expressing high levels of cyclin D1 demonstrated widespread mitochondrial translocation of Bax and cytochrome c release that was approximately doubled after the addition of all-trans retinoic acid (RA) or Bcl-2 antisense oligonucleotide. By comparison, the percentage of cells in Lac Z virus-infected cultures containing translocated Bax and cytoplasmic cytochrome c was markedly less even after RA treatment. Despite this, RA-treated Lac Z and untreated cyclin D1 virus-infected cultures contained similarly low proportions of cells with active caspase or cells that were permeable to propidium iodide. Bax activation was p53-dependent and accompanied by arrest in G(2) phase. Although constitutive Bcl-2 expression prevented Bax activation, it did not alter cyclin D1-induced cell cycle arrest, illustrating the independence of these events. Both RA and antisense Bcl-2 oligonucleotide decreased Bcl-2 protein levels and markedly increased caspase activity and apoptosis in cyclin D1-infected cells. Thus amplified cyclin D1 expression initiates an apoptotic signal inhibited by different levels of cellular Bcl-2 at two points. Whereas high cellular levels of Bcl-2 prevent mitochondrial Bax translocation, lower levels can prevent apoptosis by inhibition of caspase activation.

Induction of programmed cell death with an antisense Bcl-2 oligonucleotide.

AbstractLimited advances have occurred in lung cancer prevention and treatment (1), which has necessitated the investigation of other means of intervention.As cancer is a disease of altered cellular homeostasis and genetic damage, programmed cell death (PCD) has gained increasing importance as a physiologic mechanism that has relevance to cancer-treatment strategies (2). Dysregulation of PCD in lung cancer renders these cells resistant to chemotherapeutic agents that induce this process (3,4). Therefore, genetic manipulation of PCD regulatory genes may restore the altered homeostasis in these cells and permit these genetically damaged cells to enter PCD.A known regulator of the PCD pathway is Bcl-2, a protein capable of suppressing cell death, which is upregulated in human premalignant and malignant lung lesions (5,6). Bcl-2 has been localized to the nuclear membrane, the endoplasmic reticulum, the cytosol, and the outer mitochondrial membrane (7–11). The exact molecular mechanisms by which Bcl-2 inhibits PCD have not been unequivocally established.Initial results indicate Bcl-2 inactivates inducers of PCD, such as Bax, through heterodimerization and that it is the ratio of Bcl-2, or other functionally similar genes, to the expression of PCD inducers that determines whether a cell will undergo PCD (12,13).When Bcl-2 heterodimerizes with Bax within the outer mitochondria membrane it forms an ion channel, which is functionally different from the ion channel formed by Bax homodimerization (14,15). KeywordsReverse Transcription Polymerase Chain ReactionProgramme Cell Death Regulatory GeneCell Death Detection ELISAPLUSOuter Mitochondrion MembraneMalignant Lung LesionThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Antisense Bcl2 oligonucleotide in cisplatin-resistant bladder cancer cell lines.

To determine the change of expression of Bcl2 in cisplatin-resistant bladder cancer cell lines and the reversibility of chemoresistance to cisplatin with antisense oligonucleotide against Bcl2, as higher expression of Bcl2 is associated with drug resistance in many different cancer cell lines. In the cisplatin-resistant bladder tumour cell lines T24R1 and T24R2, the expression of Bcl2 was determined by reverse transcription-polymerase chain reaction and Western blot assay, and antisense oligonucleotide targeting of the Bcl2 coding sequence was administered with lipofectin. The expression of Bcl2 mRNA and protein was greater in T24R1 and T24R2 cells than in the parent T24 cells. Short-term exposure to cisplatin up-regulated Bcl2 mRNA and protein expression in parent T24 cells. Treatment with antisense oligonucleotide down-regulated Bcl2 protein expression and significantly enhanced the cytotoxicity of cisplatin. Up-regulation of Bcl2 protein expression might be one of the mechanisms of cisplatin resistance in bladder cancer cells, and antisense Bcl2 oligonucleotide may be helpful in chemotherapy for bladder cancer by reversing cisplatin resistance.

Liposomal delivery of antisense oligonucleotides for efficient downregulation of Bcl-2 and induction of apoptosis.

The aim of this study was to enhance the delivery and thus anti-tumoral efficiency of antisense bcl-2 oligonucleotides (ODN's). Bcl-2 overexpressing DoHH2 lymphoma and HeLa-cells were transfected with ODN's using a polycationic liposome preparation. Specific hybridization of antisense ODN's was demonstrated by gel-shift assays and in vitro transcription/translation studies. Cellular uptake of oligonucleotides was evaluated by fluorescence microscopy. Inhibition of bcl-2 translation was demonstrated by quantitative RT-PCR and Western Blot. TUNEL assay, ANNEXIN V-binding and Apo-2.7 expression were performed to evaluate induction of apoptosis. Using polycationic liposomes, a ODN transfection rate of 95% in HeLa and 45% in DoHH2 cells were demonstrated by fluorescence microscopy. 24 hours after transfection quantitative RT-PCR detected a 56% decrease of bcl-2 mRNA in antisense and a 7% decrease in sense transfected DoHH2 cells (p < 0.05). In HeLa-cells, bcl-2 expression was almost completely inhibited 72 hours after antisense ODN transfection. Antisense treated cells also showed significant induction of apoptosis. Polycationic liposome-mediated transfection of bcl-2 antisense ODN's causes enhanced cellular uptake and efficient bcl-2 downregulation in bcl-2 overexpressing cell lines. This delivery strategy may explain why significant induction of apoptosis was achieved at low oligonucleotide concentrations (approximately 200 pmol/5 x 10(5) tumor cells).

Depletion of Bcl-2 by an antisense oligonucleotide induces apoptosis accompanied by oxidation and externalization of phosphatidylserine in NCI-H226 lung carcinoma cells.

Oxidant-induced apoptosis involves oxidation of many different and essential molecules including phospholipids. As a result of this non-specific oxidation, any signaling role of a particular phospholipid-class of molecules is difficult to elucidate. To determine whether preferential oxidation of phosphatidylserine (PS) is an early event in apoptotic signaling related to PS externalization and is independent of direct oxidant exposure, we chose a genetic-based induction of apoptosis. Apoptosis was induced in the lung cancer cell line NCI-H226 by decreasing the amount of Bcl-2 protein expression by preventing the translation of bcl-2 mRNA using an antisense bcl-2 oligonucleotide. Peroxidation of phospholipids was assayed using a fluorescent technique based on metabolic integration of an oxidation-sensitive and fluorescent fatty acid, cis-parinaric acid (PnA), into cellular phospholipids and subsequent HPLC separation of cis-PnA-labeled phospholipids. We found a decrease in Bcl-2 was associated with a selective oxidation of PS in a sub-population of the cells with externalized PS. No significant difference in oxidation of cis-PnA-labeled phospholipids was observed in cells treated with medium alone or a nonsense oligonucleotide. Treatment with either nonsensc or antisense bcl-2 oligonucleotides was not associated with changes in the pattern of individual phospholipid classes as determined by HPTLC. These metabolic and topographical changes in PS arrangement in plasma membrane appear to be early responses to antisense bcl-2 exposure that trigger a PS-dependent apoptotic signaling pathway. This observed externalization of PS may facilitate the 'labeling' of apoptotic cells for recognition by macrophage scavenger receptors and subsequent phagocytic clearance.

Inhibition of progression to androgen-independence by combined adjuvant treatment with antisense BCL-XL and antisense Bcl-2 oligonucleotides plus taxol after castration in the Shionogi tumor model.

We have reported that antisense Bcl-2 oligodeoxynucleotide (ODN) delays progression to androgen independence in the androgen-dependent (AD) mouse Shionogi tumor model. Here, we characterize changes in bcl-xL, another important anti-apoptotic gene, and test the efficacy of adjuvant antisense Bcl-xL ODN therapy either alone or in combination with antisense Bcl-2 ODN and chemotherapy after castration in the Shionogi tumor model. Bcl-xL mRNA levels increased up to 3-fold postcastration and remained 1. 5-fold higher in androgen-independent (AI) recurrent tumors compared with AD tumors before castration. Treatment of Shionogi cells with antisense Bcl-xL ODN inhibited Bcl-xL expression in a dose-dependent and sequence-specific manner. Systemic administration of antisense Bcl-xL ODN in mice bearing Shionogi tumors after castration delayed emergence of AI recurrent tumors. We then examined whether combined adjuvant antisense Bcl-xL and/or Bcl-2 ODNs plus taxol (paclitaxel) therapy further delays time to AI progression. Combined treatment of Shionogi cells with antisense Bcl-xL and Bcl-2 ODNs significantly enhanced taxol chemosensitivity compared with either agent alone, reducing the IC(50) of taxol by more than 1 log. Apoptotic DNA laddering and cleavage of poly(ADP-ribose) polymerase were more substantial after treatment with combined antisense Bcl-2 and Bcl-xL ODNs plus taxol than that with either 2 agents. Adjuvant administration of antisense Bcl-xL and Bcl-2 ODNs plus micellar taxol resulted in a significantly delayed time to AI recurrence compared with administration of either 2 agents. Our findings suggest that Bcl-xL represents a suitable molecular target for antisense ODN strategy and illustrate the potential additive effects of multi-target pharmacology for cancer therapy.

The antisense BCL-2 oligonucleotide and CDNA expressing vector induce apoptosis in BCL-2-positive human hepatoma cells

The antisense BCL-2 oligonucleotide and CDNA expressing vector induce apoptosis in BCL-2-positive human hepatoma cells

Antisense-mediated inhibition of the bcl-2 gene induces apoptosis in human malignant glioma

BACKGROUND The bcl-2 protooncogene represses a number of cellular apoptotic pathways and is known to be expressed in increasing amounts in glial tumors of higher malignancy. We tested whether antisense oligonucleotides to the bcl-2 gene would affect glioma cell viability. METHODS Antisense oligonucleotides directed to the first six codons of the human bcl-2 gene, and nonsense oligonucleotides as a control, were transfected into malignant glioma cells. Two human Bcl-2 positive glioblastoma cell lines from our tumor bank (Jon52 and Roc) were both transfected in vitro with bcl-2 antisense (AS) and nonsense (NS) oligonucleotides at 1 μm and 5 μm concentrations for 5 and 24 hr. Cell viability was assessed at 2, 4, 5, and 7 days by using an MTT mitogenic assay and by cell counting via direct visualization using a hemocytometer. RESULTS There was up to a log-fold decrease in cell growth of the bcl-2 AS treated cells compared to the NS transfected cells for both Roc ( p = 0.007 and p = 0.004) and Jon52 ( p = 0.02 and p = 0.004) at 5 and 24 hr of transfection. There was as much as 50% cytotoxicity in both glioblastoma cell lines at 1 μm and 5 μm concentrations after 24 hr transfection with AS bcl-2 oligonucleotides (all p < 0.01). Western blot analysis demonstrated a decrease in the expression of the Bcl-2 protein in one cell line, whereas there was a statistically significant increase in the apoptotic index of both cell lines ( p < 0.05 by chi square analysis). CONCLUSIONS Our results suggest that transfection of human glioma cells with antisense bcl-2 results in an increase in apoptotic death. This provides evidence that Bcl-2 plays a role in tumor progression of glioma by acting as an oncogene, and suggests that inhibition of the bcl-2 gene could have a therapeutic effect.

Both HPV and carcinogen contribute to the development of resistance to apoptosis during oral carcinogenesis.

Oral carcinomas frequently contain human papilloma virus (HPV)-16/18. As p53 is degraded through interaction with HPV-16/18 products (E6/E7), p53 dysfunction may contribute to oral carcinogenesis. Furthermore, epidemiological studies suggest that smoking history may be critical for oral carcinogenesis. To delineate the involvement of HPV-16 infection and carcinogen in oral carcinogenesis, Park et al have established a multistep oral carcinogenesis model. Overexpression of p53 altered the expression of Fas antigen (Fas-R), Bax and Bcl-2; however, it remains unclear how the loss of p53 modifies the expression of these molecules. Using the multistep oral carcinogenesis model, we analyzed how the loss of p53 and carcinogen modified the expression of these molecules and their role in the development of resistance to apoptosis of oral carcinomas. The HOK-16B cell line was immortalized by HPV-16 transfection of normal human oral keratinocytes (NHOK). HOK-16B-BaP and HOK-16B-BaP-T1 were established from HOK-16B following short-term and long-term stimulation with the chemical carcinogen, benzo(a)pyrene, respectively. The malignant phenotype develops in sequence from HOK-16B, HOK-16B-BaP and HOK-16B-BaP-T1. The expression of apoptosis-related molecules was examined by Western blot analysis or by flow cytometry. Fas-mediated cytotoxicity was assessed using CH-11, an agonistic anti-Fas-R IgM monoclonal antibody. The apoptosis-related molecules examined were the Fas-R, Bcl-2, Bax, and Fas-associated phosphatase 1 (FAP-1). Downregulation of Fas-R and upregulation of Bcl-2 in HOK-16B-BaP were observed in HOK-16B-BaP and HOK-16B-BaPT1. Bax was downregulated in HOK-16B, HOK-16B-BaP and HOK-16B-BaP-T1. The expression of FAP-1 was increased with progression towards malignancy. NHOK and HOK-16B were relatively sensitive to CH-11, whereas HOK-BaP and HOK-BaP-T1 were resistant to CH-11. Treatment of HOK-16B-BaP with antisense bcl-2 oligonucleotide rendered the cells more sensitive to CH-11-induced apoptosis. These data demonstrate that both the loss of p53 and carcinogen stimulation are associated with altered expression of Fas-R, Bcl-2 and FAP-1, although the loss of p53 is sufficient for altered expression of Bax. Thus, both HPV infection and smoking contribute to acquisition of anti-apoptotic characteristics by oral carcinomas.

Antisense bcl-2 treatment increases programmed cell death in non-small cell lung cancer cell lines

Programmed cell death (PCD) is a genetically regulated pathway that is altered in many cancers. This process is, in part, regulated by the ratio of PCD inducers (Bax) or inhibitors (Bcl-2). An abnormally high ratio of Bcl-2 to Bax prevents PCD, thus contributing to resistance to chemotherapeutic agents, many of which are capable of inducing PCD. Non-small cell lung cancer (NSCLC) cells demonstrate resistance to these PCD-inducing agents. If Bcl-2 prevents NSCLC cells from entering the PCD pathway, then reducing the amount of endogenous Bcl-2 product may allow these cells to spontaneously enter the PCD pathway. Our purpose was to determine the effects of bcl-2 antisense treatment on the levels of programmed cell death in NSCLC cells. First, we determined whether bcl-2 and bax mRNA were expressed in three morphologically distinct NSCLC cell lines: NCI-H226 (squamous), NCI-H358 (adenocarcinoma), and NCI-H596 (adenosquamous). Cells were then exposed to synthetic antisense bcl-2 oligonucleotide treatment, after which programmed cell death was determined, as evidenced by DNA fragmentation. Bcl-2 protein expression was detected immunohistochemically. All three NSCLC cell lines expressed both bcl-2 and bax mRNA and had functional PCD pathways. Synthetic antisense bcl-2 oligonucleotide treatment resulted in decreased Bcl-2 levels, reduced cell proliferation, decreased cell viability, and increased levels of spontaneous PCD. This represents the first evidence that decreasing Bcl-2 in three morphologically distinct NSCLC cell lines allows the cells to spontaneously enter a PCD pathway. It also indicates the potential therapeutic use of antisense bcl-2 in the treatment of NSCLC.

Growth inhibition of DU-145 prostate cancer cells by a Bcl-2 antisense oligonucleotide is enhanced by N-(2-hydroxyphenyl)all-trans retinamide.

Hormonally insensitive prostate cancer is a relatively slow-growing, but usually fatal, disease with no long-term treatment options. Transformation of normal prostate cells to a malignant phenotype often involves corruption of the apoptotic machineries. Bcl-2 protein is one of the key inhibitors of apoptosis and is often unregulated in advanced prostate cancer. The prostate cancer cell line DU-145 was used as a model of a hormonally insensitive, advanced prostate cancer. Cell growth in liquid culture was significantly inhibited by antisense Bcl-2 oligonucleotides compared with control sense oligonucleotides; inhibition by these oligonucleotides was significantly enhanced on combination with the synthetic retinoid N-(2-hydroxyphenyl)all-trans-retinamide (2-HPR). Interestingly, growth inhibition occurred in the absence of apoptosis as measured using two assay techniques. We hypothesize that in these recalcitrant cells the apoptotic pathway is compromised at several levels, and Bcl-2 may play another role in promoting cell growth. The use of Bcl-2 antisense oligonucleotides plus 2-HPR may provide a novel approach to therapy of hormone-resistant prostate cancer.