Communication between cells: exosomes as a delivery system in prostate cancer

Despite the considerable efforts in screening and diagnostic protocols, prostate cancer still represents the second leading cause of cancer-related death in men. Many patients with localized disease and low risk of recurrence have a favourable outcome. In a substantial proportion of patients, however, the disease progresses and becomes aggressive. The mechanisms that promote prostate cancer progression remain still debated. Many findings point to the role of cross-communication between prostate tumor cells and their surrounding microenvironment during the disease progression. Such a connection fosters survival, proliferation, angiogenesis, metastatic spreading and drug-resistance of prostate cancer. Recent years have seen a profound interest in understanding the way by which prostate cancer cells communicate with the surrounding cells in the microenvironment. In this regard, direct cell-to-cell contacts and soluble factors have been identified. Increasing evidence indicates that PC cells communicate with the surrounding cells through the release of extracellular vesicles, mainly the exosomes. By directly acting in stromal or prostate cancer epithelial cells, exosomes represent a critical intercellular communication system. By querying the public database (https://pubmed.ncbi.nlm.nih.gov) for the past 10 years, we have found more than four hundred papers. Among them, we have extrapolated the most relevant about the role of exosomes in prostate cancer malignancy and progression. Emerging data concerning the use of these vesicles in diagnostic management and therapeutic guidance of PC patients are also presented. Graphical Abstract

Bone Marrow-Derived Mesenchymal Stem Cells Migrate toward Hormone-Insensitive Prostate Tumor Cells Expressing TGF-β via N-Cadherin

The prostate tumor microenvironment plays important roles in the metastasis and hormone-insensitive re-growth of tumor cells. Bone marrow-derived mesenchymal stem cells (BM-MSCs) are recruited into prostate tumors to facilitate tumor microenvironment formation. However, the specific intrinsic molecules mediating BM-MSCs’ migration to prostate tumors are unknown. BM-MSCs’ migration toward a conditioned medium (CM) of hormone-insensitive (PC3 and DU145) or hormone-sensitive (LNCaP) prostate tumor cells was investigated using a three-dimensional cell migration assay and a transwell migration assay. PC3 and DU145 expressed transforming growth factor-β (TGF-β), but LNCaP did not. Regardless of TGF-β expression, BM-MSCs migrated toward the CM of PC3, DU145, or LNCaP. The CM of PC3 or DU145 expressing TGF-β increased the phosphorylation of Smad2/3 in BM-MSCs. Inactivation of TGF-β signaling in BM-MSCs using TGF-β type 1 receptor (TGFBR1) inhibitors, SB505124, or SB431542 did not allow BM-MSCs to migrate toward the CM. The CM of PC3 or DU145 enhanced N-cadherin expression on BM-MSCs, but the LNCaP CM did not. SB505124, SB431542, and TGFBR1 knockdown prevented an increase in N-cadherin expression. N-cadherin knockdown inhibited the collective migration of BM-MSCs toward the PC3 CM. We identified N-cadherin as a mediator of BM-MSCs’ migration toward hormone-insensitive prostate tumor cells expressing TGF-β and introduced a novel strategy for controlling and re-engineering the prostate tumor microenvironment.

Advances in Magnetic Nanoparticle-Driven Delivery of Gene Therapies towards Prostate Cancer

Conventional treatment possibilities for one of the most common diseases among men, i.e., prostate cancer has several side effects. Gene-based therapy such as siRNA, CRISPR/Cas9, pDNA, and miRNA have emerged as an alternative, combating posttherapy side effects and drug resistance. Magnetic nanoparticles have been appropriately modified and functionalized to efficiently deliver the gene therapy-based active compounds to prostate tumor cells. The main purpose of this review article is to highlight the strategies currently being utilized for the treatment of prostate cancer using magnetic nanoparticles for delivery of genetic material using both the passive and active (ligand-based) targeting. It further discusses the challenges in efficient delivery of therapeutics to tumor sites and their remedial approaches. Finally, it provides a glimpse of future advances for tumor-specific modifications of magnetic nanoparticles to combat prostate cancer.

Antitumor Activity and Mechanism of Action of the Hormonotoxin, an LHRH Analog Conjugated to Dermaseptin-B2, a Multifunctional Antimicrobial Peptide

Prostate cancer represents the most common cancer in men. For patients with advanced or metastatic form, treatments will be able to slow down the progression but cannot cure it even with the used of new targeted therapies. In this context, the development of innovative drugs resulting from the exploration of biodiversity could open new therapeutic alternatives. Dermaseptin-B2 (DRS-B2), a natural multifunctional antimicrobial peptide isolated from the Amazonian frog skin, has been reported to possess antitumor and antiangiogenic activities. To improve DRS-B2 pharmacological properties and target prostate tumor cells more specifically, we have developed a chimeric molecule, called Hormonotoxin (H-B2) which is composed of a DRS-B2 combined with a hormonal analog, d-Lys6-LHRH, to target LHRH-Receptor which is overexpressed in more than 85% of prostate cancers. In vitro H-B2 has a significant antiproliferative effect on the PC3 tumor cell line, with an IC50 value close to that of DRS-B2. The antitumor activity of H-B2 was confirmed in vivo in mouse model xenografted with PC3 tumors and appears to be better tolerated than DRS-B2. Biophysical experiments showed that the addition of the hormonal analog to DRS-B2 did not alter either its secondary structure or its biological activity. Combination of different experimental approaches indicated that H-B2 induces cell death by an apoptotic mechanism whereas DRS-B2 was shown to induce it by necrosis. These results could explain the H-B2 less toxicity compared to DRS-B2. H-B2 represents a promising targeting approach for cancer therapy.

Attenuating CD3 affinity in a PSMAxCD3 bispecific antibody enables killing of prostate tumor cells with reduced cytokine release

BackgroundTherapeutic options currently available for metastatic castration-resistant prostate cancer (mCRPC) do not extend median overall survival >6 months. Therefore, the development of novel and effective therapies for mCRPC represents an urgent medical need. T cell engagers (TCEs) have emerged as a promising approach for the treatment of mCRPC due to their targeted mechanism of action. However, challenges remain in the clinic due to the limited efficacy of TCEs observed thus far in solid tumors as well as the toxicities associated with cytokine release syndrome (CRS) due to the usage of high-affinity anti-CD3 moieties such as OKT3.MethodsUsing genetically engineered transgenic rats (UniRat and OmniFlic) that express fully human IgG antibodies together with an NGS-based antibody discovery pipeline, we developed TNB-585, an anti-CD3xPSMA TCE for the treatment of mCRPC. TNB-585 pairs a tumor-targeting anti-PSMA arm together with a unique, low-affinity anti-CD3 arm in bispecific format. We tested TNB-585 in T cell-redirected cytotoxicity assays against PSMA+ tumor cells in both two-dimensional (2D) cultures and three-dimensional (3D) spheroids as well as against patient-derived prostate tumor cells. Cytokines were measured in culture supernatants to assess the ability of TNB-585 to induce tumor killing with low cytokine release. TNB-585-mediated T cell activation, proliferation, and cytotoxic granule formation were measured to investigate the mechanism of action. Additionally, TNB-585 efficacy was evaluated in vivo against C4-2 tumor-bearing NCG mice.ResultsIn vitro, TNB-585 induced activation and proliferation of human T cells resulting in the killing of PSMA+ prostate tumor cells in both 2D cultures and 3D spheroids with minimal cytokine release and reduced regulatory T cell activation compared with a positive control antibody that contains the same anti-PSMA arm but a higher affinity anti-CD3 arm (comparable with OKT3). In addition, TNB-585 demonstrated potent efficacy against patient-derived prostate tumors ex vivo and induced immune cell infiltration and dose-dependent tumor regression in vivo.ConclusionsOur data suggest that TNB-585, with its low-affinity anti-CD3, may be efficacious while inducing a lower incidence and severity of CRS in patients with prostate cancer compared with TCEs that incorporate high-affinity anti-CD3 domains.

Control of Surface Properties of Hyaluronan/Chitosan Multilayered Coatings for Tumor Cell Capture

Prostate cancer (PCa) is a slow-growing neoplasm that has, when diagnosed in its early stages, great chances of cure. During initial tumor development, current diagnostic methods fail to have the desired accuracy, thus, it is necessary to develop or improve current detection methods and prognostic markers for PCa. In this scenario, films composed of hyaluronic acid (HA) and chitosan (CHI) have demonstrated significant capture potential of prostate tumor cells (PC3 line), exploring HA as a CD44 receptor ligand and direct mediator in cell-film adhesion. Here, we present a strategy to control structural and cell adhesion properties of HA/CHI films based on film assembly conditions. Films were built via Layer-by-layer (LbL) deposition, where the pH conditions (3.0 and 5.0) and number of bilayers (3.5, 10.5, and 20.5) were controlled. The characterization of these films was carried out using profilometry, ultraviolet-visible (UV-VIS), atomic force microscopy (AFM) and contact angle measurements. Multilayer HA/CHI films produced at pH 3.0 gave optimum surface wettability and availability of free carboxyl groups. In turn, at pH 5.0, the coverings were thinner and presented a smoother surface. Films prepared with 3.5 bilayers showed greater tumor cell capture regardless of the pH condition, while films containing 10.5 and 20.5 bilayers presented a significant swelling process, which compromised their cell adhesion potential. This study shows that surface chemistry and morphology are critical factors for the development of biomaterials designed for several cell adhesion applications, such as rapid diagnostic, cell signaling, and biosensing mechanisms.

Monomethyl Auristatin E Grafted-Liposomes to Target Prostate Tumor Cell Lines

Novel nanomedicines have been engineered to deliver molecules with therapeutic potentials, overcoming drawbacks such as poor solubility, toxicity or short half-life. Lipid-based carriers such as liposomes represent one of the most advanced classes of drug delivery systems. A Monomethyl Auristatin E (MMAE) warhead was grafted on a lipid derivative and integrated in fusogenic liposomes, following the model of antibody drug conjugates. By modulating the liposome composition, we designed a set of particles characterized by different membrane fluidities as a key parameter to obtain selective uptake from fibroblast or prostate tumor cells. Only the fluid liposomes made of palmitoyl-oleoyl-phosphatidylcholine and dioleoyl-phosphatidylethanolamine, integrating the MMAE-lipid derivative, showed an effect on prostate tumor PC-3 and LNCaP cell viability. On the other hand, they exhibited negligible effects on the fibroblast NIH-3T3 cells, which only interacted with rigid liposomes. Therefore, fluid liposomes grafted with MMAE represent an interesting example of drug carriers, as they can be easily engineered to promote liposome fusion with the target membrane and ensure drug selectivity.

X-ray Fluorescence Uptake Measurement of Functionalized Gold Nanoparticles in Tumor Cell Microsamples

Quantitative cellular in vitro nanoparticle uptake measurements are possible with a large number of different techniques, however, all have their respective restrictions. Here, we demonstrate the application of synchrotron-based X-ray fluorescence imaging (XFI) on prostate tumor cells, which have internalized differently functionalized gold nanoparticles. Total nanoparticle uptake on the order of a few hundred picograms could be conveniently observed with microsamples consisting of only a few hundreds of cells. A comparison with mass spectroscopy quantification is provided, experimental results are both supported and sensitivity limits of this XFI approach extrapolated by Monte-Carlo simulations, yielding a minimum detectable nanoparticle mass of just 5 pg. This study demonstrates the high sensitivity level of XFI, allowing non-destructive uptake measurements with very small microsamples within just seconds of irradiation time.

Unbiased Phenotype-Based Screen Identifies Therapeutic Agents Selective for Metastatic Prostate Cancer

In American men, prostate cancer is the second leading cause of cancer-related death. Dissemination of prostate cancer cells to distant organs significantly worsens patients’ prognosis, and currently there are no effective treatment options that can cure advanced-stage prostate cancer. In an effort to identify compounds selective for metastatic prostate cancer cells over benign prostate cancer cells or normal prostate epithelial cells, we applied a phenotype-based in vitro drug screening method utilizing multiple prostate cancer cell lines to test 1,120 different compounds from a commercial drug library. Top drug candidates were then examined in multiple mouse xenograft models including subcutaneous tumor growth, experimental lung metastasis, and experimental bone metastasis assays. A subset of compounds including fenbendazole, fluspirilene, clofazimine, niclosamide, and suloctidil showed preferential cytotoxicity and apoptosis towards metastatic prostate cancer cells in vitro and in vivo. The bioavailability of the most discerning agents, especially fenbendazole and albendazole, was improved by formulating as micelles or nanoparticles. The enhanced forms of fenbendazole and albendazole significantly prolonged survival in mice bearing metastases, and albendazole-treated mice displayed significantly longer median survival times than paclitaxel-treated mice. Importantly, these drugs effectively targeted taxane-resistant tumors and bone metastases – two common clinical conditions in patients with aggressive prostate cancer. In summary, we find that metastatic prostate tumor cells differ from benign prostate tumor cells in their sensitivity to certain drug classes. Taken together, our results strongly suggest that albendazole, an anthelmintic medication, may represent a potential adjuvant or neoadjuvant to standard therapy in the treatment of disseminated prostate cancer.