Baculoviruses as Vectors for Gene Therapy against Human Prostate Cancer

Current curative strategies for prostate cancer are restricted to the primary tumour, and the effect of treatments to control metastatic disease is not sustained. Therefore, the application of gene therapy to prostate cancer is an attractive alternative. Baculoviruses are highly restricted insect viruses, which can enter, but not replicate in mammalian cells. Baculoviruses can incorporate large amounts of extra genetic material, and will express transgenes in mammalian cells when under the control of a mammalian or strong viral promoter. Successful gene delivery has been achieved both in vitro and in vivo and into both dividing and nondividing cells, which is important since prostate cancers divide relatively slowly. In addition, the envelope protein gp64 is sufficiently mutable to allow targeted transduction of particular cell types. In this review, the advantages of using baculoviruses for prostate cancer gene therapy are explored, and the mechanisms of viral entry and transgene expression are described

Resistance to Antiandrogens in Prostate Cancer : Is It Inevitable, Intrinsic or Induced?

Increasingly sophisticated therapies for chemical castration dominate first-line treatments for locally advanced prostate cancer. However, androgen deprivation therapy (ADT) offers little prospect of a cure, as resistant tumors emerge rather rapidly, normally within 30 months. Cells have multiple mechanisms of resistance to even the most sophisticated drug regimes, and both tumor cell heterogeneity in prostate cancer and the multiple salvage pathways result in castration-resistant disease related genetically to the original hormone-naive cancer. The timing and mechanisms of cell death after ADT for prostate cancer are not well understood, and off-target effects after long-term ADT due to functional extra-prostatic expression of the androgen receptor protein are now increasingly being recorded. Our knowledge of how these widely used treatments fail at a biological level in patients is deficient. In this review, I will discuss whether there are pre-existing drug-resistant cells in a tumor mass, or whether resistance is induced/selected by the ADT. Equally, what is the cell of origin of this resistance, and does it differ from the treatment-naïve tumor cells by differentiation or dedifferentiation? Conflicting evidence also emerges from studies in the range of biological systems and species employed to answer this key question. It is only by improving our understanding of this aspect of treatment and not simply devising another new means of androgen inhibition that we can improve patient outcomes

Characterization of Human Papillomavirus Type 16 E2 Protein and Subdomains Expressed in Insect Cells

AbstractThe E2 open reading frame of human papillomavirus type 16 (HPV-16) encodes a DNA-binding protein which modulates papillomavirus transcription and replication. To investigate the biological and biochemical properties of the HPV-16 E2 protein, we have constructed recombinant baculoviruses which express the full-length molecule and individual N- and C-terminal domains in Sf21 insect cells. In this system the full-length E2 protein was phosphorylated and targeted to the insect cell nucleus. A 93 amino acid C-terminal fragment encompassing the DNA binding and dimerization functions of E2 was also translocated to the nucleus but was not modified by phosphorylation. The E2 N-terminal protein accumulated in the insect cell cytoplasm but was not efficiently phosphorylated. The formation of heterodimers between full-length and N-terminally truncated E2 species was observed when Sf21 cells were co-infected with recombinant viruses and when homodimers were mixed in vitro, suggesting that the dimer interface is not sufficiently stable to prevent subunit exchange in vivo. Both homo- and heterodimeric E2 species were able to bind specifically and in any combination to tandem E2 binding sites from the HPV-16 regulatory region. Furthermore, the HPV-16 E2 protein bound to DNA exhibited a distinct susceptibility profile to pronase digestion, potentially contrasting with that reported for BPV-1 E2. These observations suggest that significant structural and functional differences may exist between the BPV/HPV E2 proteins and have implications for understanding E2-dependent regulation of transcription and replication

Low Temperature Plasma : A Novel Focal Therapy for Localized Prostate Cancer?

Despite considerable advances in recent years for the focal treatment of localized prostate cancer, high recurrence rates and detrimental side effects are still a cause for concern. In this review, we compare current focal therapies to a potentially novel approach for the treatment of early onset prostate cancer: low temperature plasma. The rapidly evolving plasma technology has the potential to deliver a wide range of promising medical applications via the delivery of plasma-induced reactive oxygen and nitrogen species. Studies assessing the effect of low temperature plasma on cell lines and xenografts have demonstrated DNA damage leading to apoptosis and reduction in cell viability. However, there have been no studies on prostate cancer, which is an obvious candidate for this novel therapy. We present here the potential of low temperature plasma as a focal therapy for prostate cancer

Mechanisms of growth inhibition of primary prostate epithelial cells following gamma irradiation or photodynamic therapy including senscence, necrosis, and autophagy, but not apoptosis

In comparison to more differentiated cells, prostate cancer stem-like cells are radioresistant, which could explain radio-recurrent prostate cancer. Improvement of radiotherapeutic efficacy may therefore require combination therapy. We have investigated the consequences of treating primary prostate epithelial cells with gamma irradiation and photodynamic therapy (PDT), both of which act through production of reactive oxygen species (ROS). Primary prostate epithelial cells were cultured from patient samples of benign prostatic hyperplasia and prostate cancer prior to treatment with PDT or gamma irradiation. Cell viability was measured using MTT and alamar blue assay, and cell recovery by colony-forming assays. Immunofluorescence of gamma-H2AX foci was used to quantify DNA damage, and autophagy and apoptosis were assessed using Western blots. Necrosis and senescence were measured by propidium iodide staining and beta-galactosidase staining, respectively. Both PDT and gamma irradiation reduced the colony-forming ability of primary prostate epithelial cells. PDT reduced the viability of all types of cells in the cultures, including stem-like cells and more differentiated cells. PDT induced necrosis and autophagy, whereas gamma irradiation induced senescence, but neither treatment induced apoptosis. PDT and gamma irradiation therefore inhibit cell growth by different mechanisms. We suggest these treatments would be suitable for use in combination as sequential treatments against prostate cancer

Epigenetic Control of Gene Expression in the Normal and Malignant Human Prostate : A Rapid Response Which Promotes Therapeutic Resistance

A successful prostate cancer must be capable of changing its phenotype in response to a variety of microenvironmental influences, such as adaptation to treatment or successful proliferation at a particular metastatic site. New cell phenotypes emerge by selection from the large, genotypically heterogeneous pool of candidate cells present within any tumor mass, including a distinct stem cell-like population. In such a multicellular model of human prostate cancer, flexible responses are primarily governed not only by de novo mutations but appear to be dominated by a combination of epigenetic controls, whose application results in treatment resistance and tumor relapse. Detailed studies of these individual cell populations have resulted in an epigenetic model for epithelial cell differentiation, which is also instructive in explaining the reported high and inevitable relapse rates of human prostate cancers to a multitude of treatment types

Retinoic acid and androgen receptors combine to achieve tissue specific control of human prostatic transglutaminase expression: a novel regulatory network with broader significance

In the human prostate, expression of prostate-specific genes is known to be directly regulated by the androgen–induced stimulation of the androgen receptor (AR). However, less is known about the expression control of the prostate-restricted TGM4 (hTGP) gene. In the present study we demonstrate that the regulation of the hTGP gene depends mainly on retinoic acid (RA). We provide evidence that the retinoic acid receptor gamma (RAR-G) plays a major role in the regulation of the hTGP gene and that presence of the AR, but not its transcriptional transactivation activity, is critical for hTGP transcription. RA and androgen responsive elements (RARE and ARE) were mapped to the hTGP promoter by chromatin immunoprecipitation (ChIP), which also indicated that the active ARE and RARE sites were adjacent, suggesting that the antagonistic effect of androgen and RA is related to the relative position of binding sites. Publicly available AR and RAR ChIP-seq data was used to find gene potentially regulated by AR and RAR. Four of these genes (CDCA7L, CDK6, BTG1 and SAMD3) were tested for RAR and AR binding and two of them (CDCA7L and CDK6) proved to be antagonistically regulated by androgens and RA confirming that this regulation is not particular of hTGP

Inhibition of the glucocorticoid receptor results in an enhanced miR-99a/100-mediated radiation response in stem-like cells from human prostate cancers

Radiation therapy is a major primary treatment option for both localized early stage prostate cancer, and for advanced, regionally un-resectable, cancer. However, around 30% of patients still experience biochemical recurrence after radiation therapy within 10 years. Thus, identification of better biomarkers and new targets are urgently required to improve current therapeutic strategies. The miR-99 family has been shown to play an important role in the regulation of the DNA damage response, via targeting of the SWI/SNF chromatin remodeling factors, SMARCA5 and SMARCD1 in cell line models. In the present study, we have demonstrated that low expression of miR-99a and miR-100 is present in cell populations which are relatively radiation insensitive, for example in prostate cancer stem cells and in castration-resistant prostate cancer. Additionally, treatment of cells with the synthetic glucocorticoid, Dexamethasone resulted in decreased miR-99a and 100 expression, suggesting a new mechanism of miR-99a and 100 regulation in androgen-independent prostate cells. Strikingly, treatment of prostate cells with the glucocorticoid receptor inhibitor, Mifepristone was found to sensitize prostate cells to radiation by increasing the levels of miR-99a and miR-100. These results qualify the miR99 family as markers of radiation sensitivity and as potential therapeutic targets to improve efficiency of radiotherapy

Low temperature plasmas as emerging cancer therapeutics : the state of play and thoughts for the future

The field of plasma medicine has seen substantial advances over the last decade, with applications developed for bacterial sterilisation, wound healing and cancer treatment. Low temperature plasmas (LTPs) are particularly suited for medical purposes since they are operated in the laboratory at atmospheric pressure and room temperature, providing a rich source of reactive oxygen and nitrogen species (RONS). A great deal of research has been conducted into the role of reactive species in both the growth and treatment of cancer, where long-established radio- and chemo-therapies exploit their ability to induce potent cytopathic effects. In addition to producing a plethora of RONS, LTPs can also create strong electroporative fields. From an application perspective, it has been shown that LTPs can be applied precisely to a small target area. On this basis, LTPs have been proposed as a promising future strategy to accurately and effectively control and eradicate tumours. This review aims to evaluate the current state of the literature in the field of plasma oncology and highlight the potential for the use of LTPs in combination therapy. We also present novel data on the effect of LTPs on cancer stem cells, and speculatively outline how LTPs could circumvent treatment resistance encountered with existing therapeutics