Epigenetic deregulation in breast cancer microenvironment: Implications for tumor progression and therapeutic strategies.

Breast cancer comprises a substantial proportion of cancer diagnoses in women and is a primary cause of cancer-related mortality. While hormone-responsive cases generally have a favorable prognosis, the aggressive nature of triple-negative breast cancer presents challenges, with intrinsic resistance to established treatments being a persistent issue. The complexity intensifies with the emergence of acquired resistance, further complicating the management of breast cancer. Epigenetic changes, encompassing DNA methylation, histone and RNA modifications, and non-coding RNAs, are acknowledged as crucial contributors to the heterogeneity of breast cancer. The unique epigenetic landscape harbored by each cellular component within the tumor microenvironment (TME) adds great diversity to the intricate regulations which influence therapeutic responses. The TME, a sophisticated ecosystem of cellular and non-cellular elements interacting with tumor cells, establishes an immunosuppressive microenvironment and fuels processes such as tumor growth, angiogenesis, and extracellular matrix remodeling. These factors contribute to challenging conditions in cancer treatment by fostering a hypoxic environment, inducing metabolic stress, and creating physical barriers to drug delivery. This article delves into the complex connections between breast cancer treatment response, underlying epigenetic changes, and vital interactions within the TME. To restore sensitivity to treatment, it emphasizes the need for combination therapies considering epigenetic changes specific to individual members of the TME. Recognizing the pivotal role of epigenetics in drug resistance and comprehending the specificities of breast TME is essential for devising more effective therapeutic strategies. The development of reliable biomarkers for patient stratification will facilitate tailored and precise treatment approaches.

The third dimension of tumor microenvironment-The importance of tumor stroma in 3D cancer models.

Recent advances in the three-dimensional (3D) cancer models give rise to a plethora of new possibilities in the development of anti-cancer drug therapies and bring us closer to personalized medicine. Three-dimensional models are undoubtedly more authentic than traditional two-dimensional (2D) cell cultures. Nowadays, they are becoming preferentially used in most cancer research fields due to their more accurate biomimetic characteristics. On the contrary, they still lack the cellular and matrix complexity of the native tumor microenvironment (TME). This review focuses on the description of existing 3D models, the incorporation of TME and fluidics into these models, and their perspective in the future research. It is clear that such an improvement would need not only biological but also technical progress. Therefore, the modern approach to anti-cancer drug discovery should involve various fields.

Nanomedicine and epigenetics: New alliances to increase the odds in pancreatic cancer survival.

Pancreatic ductal adenocarcinoma (PDAC) is among the deadliest cancers worldwide, primarily due to its robust desmoplastic stroma and immunosuppressive tumor microenvironment (TME), which facilitate tumor progression and metastasis. In addition, fibrous tissue leads to sparse vasculature, high interstitial fluid pressure, and hypoxia, thereby hindering effective systemic drug delivery and immune cell infiltration. Thus, remodeling the TME to enhance tumor perfusion, increase drug retention, and reverse immunosuppression has become a key therapeutic strategy. In recent years, targeting epigenetic pathways has emerged as a promising approach to overcome tumor immunosuppression and cancer progression. Moreover, the progress in nanotechnology has provided new opportunities for enhancing the efficacy of conventional and epigenetic drugs. Nano-based drug delivery systems (NDDSs) offer several advantages, including improved drug pharmacokinetics, enhanced tumor penetration, and reduced systemic toxicity. Smart NDDSs enable precise targeting of stromal components and augment the effectiveness of immunotherapy through multiple drug delivery options. This review offers an overview of the latest nano-based approaches developed to achieve superior therapeutic efficacy and overcome drug resistance. We specifically focus on the TME and epigenetic-targeted therapies in the context of PDAC, discussing the advantages and limitations of current strategies while highlighting promising new developments. By emphasizing the immense potential of NDDSs in improving therapeutic outcomes in PDAC, our review paves the way for future research in this rapidly evolving field.

Establishment of Pancreatic Cancer-Derived Tumor Organoids and Fibroblasts From Fresh Tissue.

Tumor organoids are three-dimensional (3D) ex vivo tumor models that recapitulate the biological key features of the original primary tumor tissues. Patient-derived tumor organoids have been used in translational cancer research and can be applied to assess treatment sensitivity and resistance, cell-cell interactions, and tumor cell interactions with the tumor microenvironment. Tumor organoids are complex culture systems that require advanced cell culture techniques and culture media with specific growth factor cocktails and a biological basement membrane that mimics the extracellular environment. The ability to establish primary tumor cultures highly depends on the tissue of origin, the cellularity, and the clinical features of the tumor, such as the tumor grade. Furthermore, tissue sample collection, material quality and quantity, as well as correct biobanking and storage are crucial elements of this procedure. The technical capabilities of the laboratory are also crucial factors to consider. Here, we report a validated SOP/protocol that is technically and economically feasible for the culture of ex vivo tumor organoids from fresh tissue samples of pancreatic adenocarcinoma origin, either from fresh primary resected patient donor tissue or patient-derived xenografts (PDX). The technique described herein can be performed in laboratories with basic tissue culture and mouse facilities and is tailored for wide application in the translational oncology field.

Liquid biopsy and preclinical tools for advancing diagnosis and treatment of patients with pancreatic neuroendocrine neoplasms.

Pancreatic neuroendocrine neoplasms (pNENs) are rare cancers with broad challenges for their management. The main clinical obstacles are the high rate of patients diagnosed at advanced stages, lack of prognostic markers for early detection of disease recurrence in resected patients, significant limitations in identifying those who will benefit from adjuvant therapy, and timely recognition of treatment response. Therefore, the discovery of new prognostic and predictive markers is necessary for patient stratification and clinical management. Liquid biopsy, which has revolutionized the field of clinical oncology, is extremely under-investigated in pNENs. This review highlights its potential and the recent advances in related technologies, as candidates for the delivery of the new tools that can help to refine pNEN diagnosis and to personalize treatment. In addition, the opportunities and limitations of available preclinical research models with regard to biomarker research are discussed in light of pNEN clinical needs.

Decitabine potentiates efficacy of doxorubicin in a preclinical trastuzumab-resistant HER2-positive breast cancer models.

Acquired drug resistance and metastasis in breast cancer (BC) are coupled with epigenetic deregulation of gene expression. Epigenetic drugs, aiming to reverse these aberrant transcriptional patterns and sensitize cancer cells to other therapies, provide a new treatment strategy for drug-resistant tumors. Here we investigated the ability of DNA methyltransferase (DNMT) inhibitor decitabine (DAC) to increase the sensitivity of BC cells to anthracycline antibiotic doxorubicin (DOX). Three cell lines representing different molecular BC subtypes, JIMT-1, MDA-MB-231 and T-47D, were used to evaluate the synergy of sequential DAC + DOX treatment in vitro. The cytotoxicity, genotoxicity, apoptosis, and migration capacity were tested in 2D and 3D cultures. Moreover, genome-wide DNA methylation and transcriptomic analyses were employed to understand the differences underlying DAC responsiveness. The ability of DAC to sensitize trastuzumab-resistant HER2-positive JIMT-1 cells to DOX was examined in vivo in an orthotopic xenograft mouse model. DAC and DOX synergistic effect was identified in all tested cell lines, with JIMT-1 cells being most sensitive to DAC. Based on the whole-genome data, we assume that the aggressive behavior of JIMT-1 cells can be related to the enrichment of epithelial-to-mesenchymal transition and stemness-associated pathways in this cell line. The four-week DAC + DOX sequential administration significantly reduced the tumor growth, DNMT1 expression, and global DNA methylation in xenograft tissues. The efficacy of combination therapy was comparable to effect of pegylated liposomal DOX, used exclusively for the treatment of metastatic BC. This work demonstrates the potential of epigenetic drugs to modulate cancer cells' sensitivity to other forms of anticancer therapy.

Chemotherapy-triggered changes in stromal compartment drive tumor invasiveness and progression of breast cancer.

BACKGROUND: Chemotherapy remains a standard treatment option for breast cancer despite its toxic effects to normal tissues. However, the long-lasting effects of chemotherapy on non-malignant cells may influence tumor cell behavior and response to treatment. Here, we have analyzed the effects of doxorubicin (DOX) and paclitaxel (PAC), commonly used chemotherapeutic agents, on the survival and cellular functions of mesenchymal stromal cells (MSC), which comprise an important part of breast tumor microenvironment. METHODS: Chemotherapy-exposed MSC (DOX-MSC, PAC-MSC) were co-cultured with three breast cancer cell (BCC) lines differing in molecular characteristics to study chemotherapy-triggered changes in stromal compartment of the breast tissue and its relevance to tumor progression in vitro and in vivo. Conditioned media from co-cultured cells were used to determine the cytokine content. Mixture of BCC and exposed or unexposed MSC were subcutaneously injected into the immunodeficient SCID/Beige mice to analyze invasion into the surrounding tissue and possible metastases. The same mixtures of cells were applied on the chorioallantoic membrane to study angiogenic potential. RESULTS: Therapy-educated MSC differed in cytokine production compared to un-exposed MSC and influenced proliferation and secretory phenotype of tumor cells in co-culture. Histochemical tumor xenograft analysis revealed increased invasive potential of tumor cells co-injected with DOX-MSC or PAC-MSC and also the presence of nerve fiber infiltration in tumors. Chemotherapy-exposed MSC have also influenced angiogenic potential in the model of chorioallantoic membrane. CONCLUSIONS: Data presented in this study suggest that neoadjuvant chemotherapy could possibly alter otherwise healthy stroma in breast tissue into a hostile tumor-promoting and metastasis favoring niche. Understanding of the tumor microenvironment and its complex net of signals brings us closer to the ability to recognize the mechanisms that prevent failure of standard therapy and accomplish the curative purpose.

The Role of BRCA1/2-Mutated Tumor Microenvironment in Breast Cancer.

Taking into account the factors of high incidence rate, prevalence and mortality, breast cancer represents a crucial social and economic burden. Most cases of breast cancer develop as a consequence of somatic mutations accumulating in mammary epithelial cells throughout lifetime and approximately 5-10% can be ascribed to monogenic predispositions. Even though the role of genetic predispositions in breast cancer is well described in the context of genetics, very little is known about the role of the microenvironment carrying the same aberrant cells impaired by the germline mutation in the breast cancer development and progression. Based on the clinical observations, carcinomas carrying mutations in hereditary tumor-suppressor genes involved in maintaining genome integrity such as BRCA1/2 have worse prognosis and aggressive behavior. One of the mechanisms clarifying the aggressive nature of BRCA-associated tumors implies alterations within the surrounding adipose tissue itself. The objective of this review is to look at the role of BRCA1/2 mutations in the context of breast tumor microenvironment and plausible mechanisms by which it contributes to the aggressive behavior of the tumor cells.

Decreased levels of circulating cytokines VEGF, TNF-ß and IL-15 indicate PD-L1 overexpression in tumours of primary breast cancer patients.

Programmed death ligand 1 (PD-L1) overexpression has been associated with poor clinical outcomes in several human cancers whose increased malignant behaviour might be related to PD-L1 mediated systemic immunological tolerance. This study aims to verify if circulating cytokines may serve as a proxy for non-invasive identification of sensitive prognostic biomarkers reflecting tumour and its microenvironment. Immunohistochemistry was used to measure PD-L1 expression in tumour tissue sections of 148 chemonaïve breast cancer (BC) patients. The panel of 51 cytokines was analysed using multiplex bead arrays. High PD-L1 expression in tumours was associated with shorter progression-free survival (HR 3.25; 95% CI 1.39-7.61; P = 0.006) and low circulating levels of three multifunctional molecules; VEGF, TNF-ß and IL-15 (P = 0.001). In multivariate analysis, patients with low VEGF had 4.6-fold increased risk of PD-L1 overexpression (P = 0.008), present in 76.5% of patients with all these three cytokines below the median (vs. 35.6% among the others; P = 0.002). The area under the curve value of 0.722 (95% CI 0.59-0.85; P = 0.004) shows that this combination of cytokines has a moderate ability to discriminate between PD-L1 high vs. PD-L1 low patients. Plasma cytokines, therefore, could serve as potential non-invasive biomarkers for the identification of high-risk BC cases.

Increased Stromal Infiltrating Lymphocytes Are Associated with the Risk of Disease Progression in Mesenchymal Circulating Tumor Cell-Positive Primary Breast Cancer Patients.

Circulating tumor cells (CTCs) and the immune infiltration of tumors are closely related to clinical outcomes. This study aimed to verify the influence of stromal lymphocyte infiltration and the immune context of tumor microenvironment on the hematogenous spread and prognosis of 282 chemotherapy naïve primary BC patients. To detect the presence of mesenchymal CTCs, RNA extracted from CD45-depleted peripheral blood was interrogated for the expression of mesenchymal gene transcripts. Tumor-infiltrating lymphocytes (TILs) were detected in the stromal areas by immunohistochemistry, using CD3, CD8, and CD45RO antibodies. The concentrations of 51 plasma cytokines were measured by multiplex bead arrays. TILs infiltration in mesenchymal CTC-positive patients significantly decreased their progression-free survival (HR = 4.88, 95% CI 2.30-10.37, p < 0.001 for CD3high; HR = 6.17, 95% CI 2.75-13.80, p < 0.001 for CD8high; HR = 6.93, 95% CI 2.86-16.81, p < 0.001 for CD45ROhigh). Moreover, the combination of elevated plasma concentrations of transforming growth factor beta-3 (cut-off 662 pg/mL), decreased monocyte chemotactic protein-3 (cut-off 52.5 pg/mL) and interleukin-15 (cut-off 17.1 pg/mL) significantly increased the risk of disease recurrence (HR = 4.838, 95% CI 2.048-11.427, p < 0.001). Our results suggest a strong impact of the immune tumor microenvironment on BC progression, especially through influencing the dissemination and survival of more aggressive, mesenchymal CTC subtypes.

Inflammation-Based Scores Increase the Prognostic Value of Circulating Tumor Cells in Primary Breast Cancer.

A correlation between circulating tumor cells (CTCs) and monocytes in metastatic breast cancer (BC), where CTCs and monocyte-to-lymphocyte ratio (MLR) were predictors of overall survival (OS), was recently shown. Herein, we aimed to assess the association between CTCs and the complete blood count (CBC)-derived inflammation-based scores in 284 primary BC patients. CTCs were determined in CD45-depleted peripheral blood mononuclear cells by real time-PCR. This method allowed us to detect a subset of CTCs with an epithelial-to-mesenchymal transition phenotype (CTC EMT), previously associated with inferior outcomes in primary BC. In the present study, CTC EMT positivity (hazard ratio (HR) = 2.4; 95% CI 1.20-4.66, p = 0.013) and elevated neutrophil-to-lymphocyte ratio (NLR) (HR = 2.20; 95% CI 1.07-4.55; p = 0.033) were associated with shorter progression-free survival (PFS) in primary BC patients. Multivariate analysis showed that CTC EMT-positive patients with NLR ≥ 3 had 8.6 times increased risk of disease recurrence (95% CI 2.35-31.48, p = 0.001) compared with CTC EMT-negative patients with NLR < 3. Similarly, disease recurrence was 13.14 times more likely in CTC EMT-positive patients with MLR ≥ 0.34 (95% CI 4.35-39.67, p < 0.001). Given its low methodological and financial demands, the CBC-derived inflammation-based score determination could, after broader validation, significantly improve the prognostication of BC patients.

Permanent Pro-Tumorigenic Shift in Adipose Tissue-Derived Mesenchymal Stromal Cells Induced by Breast Malignancy.

During cancer progression, breast tumor cells interact with adjacent adipose tissue, which has been shown to be engaged in cancer aggressiveness. However, the tumor-directed changes in adipose tissue-resident stromal cells affected by the tumor-stroma communication are still poorly understood. The acquired changes might remain in the tissue even after tumor removal and may contribute to tumor relapse. We investigated functional properties (migratory capacity, expression and secretion profile) of mesenchymal stromal cells isolated from healthy (n = 9) and tumor-distant breast adipose tissue (n = 32). Cancer patient-derived mesenchymal stromal cells (MSCs) (MSC-CA) exhibited a significantly disarranged secretion profile and proliferation potential. Co-culture with MDA-MB-231, T47D and JIMT-1, representing different subtypes of breast cancer, was used to analyze the effect of MSCs on proliferation, invasion and tumorigenicity. The MSC-CA enhanced tumorigenicity and altered xenograft composition in immunodeficient mice. Histological analysis revealed collective cell invasion with a specific invasive front of EMT-positive tumor cells as well as invasion of cancer cells to the nerve-surrounding space. This study identifies that adipose tissue-derived mesenchymal stromal cells are primed and permanently altered by tumor presence in breast tissue and have the potential to increase tumor cell invasive ability through the activation of epithelial-to-mesenchymal transition in tumor cells.

Recent advances in understanding tumor stroma-mediated chemoresistance in breast cancer.

Although solid tumors comprise malignant cells, they also contain many different non-malignant cell types in their micro-environment. The cellular components of the tumor stroma consist of immune and endothelial cells combined with a heterogeneous population of stromal cells which include cancer-associated fibroblasts. The bi-directional interactions between tumor and stromal cells therefore substantially affect tumor cell biology.Herein, we discuss current available information on these interactions in breast cancer chemo-resistance. It is acknowledged that stromal cells extrinsically alter tumor cell drug responses with profound consequences for therapy efficiency, and it is therefore essential to understand the molecular mechanisms which contribute to these substantial alterations because they provide potential targets for improved cancer therapy. Although breast cancer patient survival has improved over the last decades, chemo-resistance still remains a significant obstacle to successful treatment.Appreciating the important experimental evidence of mesenchymal stromal cells and cancer-associated fibroblast involvement in breast cancer clinical practice can therefore have important therapeutic implications.

A disintegrin and metalloprotease 23 hypermethylation predicts decreased disease-free survival in low-risk breast cancer patients.

A Disintegrin And Metalloprotease 23 (ADAM23), a member of the ADAM family, is involved in neuronal differentiation and cancer. ADAM23 is considered a possible tumor suppressor gene and is frequently downregulated in various types of malignancies. Its epigenetic silencing through promoter hypermethylation was observed in breast cancer (BC). In the present study, we evaluated the prognostic significance of ADAM23 promoter methylation for hematogenous spread and disease-free survival (DFS). Pyrosequencing was used to quantify ADAM23 methylation in tumors of 203 BC patients. Presence of circulating tumor cells (CTC) in their peripheral blood was detected by quantitative RT-PCR. Expression of epithelial (KRT19) or mesenchymal (epithelial-mesenchymal transition [EMT]-inducing transcription factors TWIST1, SNAI1, SLUG and ZEB1) mRNA transcripts was examined in CD45-depleted peripheral blood mononuclear cells. ADAM23 methylation was significantly lower in tumors of patients with the mesenchymal CTC (P = .006). It positively correlated with Ki-67 proliferation, especially in mesenchymal CTC-negative patients (P = .001). In low-risk patients, characterized by low Ki-67 and mesenchymal CTC absence, ADAM23 hypermethylation was an independent predictor of DFS (P = .006). Our results indicate that ADAM23 is likely involved in BC progression and dissemination of mesenchymal CTC. ADAM23 methylation has the potential to function as a novel prognostic marker and therapeutic target.

Type 3 inositol 1,4,5-trisphosphate receptor has antiapoptotic and proliferative role in cancer cells.

Although the involvement of type 1 (IP3R1) and type 2 (IP3R2) inositol 1,4,5-trisphosphate receptors in apoptosis induction has been well documented in different cancer cells and tissues, the function of type 3 IP3R (IP3R3) is still elusive. Therefore, in this work we focused on the role of IP3R3 in tumor cells in vitro and in vivo. We determined increased expression of this receptor in clear cell renal cell carcinoma compared to matched unaffected part of the kidney from the same patient. Thus, we hypothesized about different functions of IP3R3 compared to IP3R1 and IP3R2 in tumor cells. Silencing of IP3R1 prevented apoptosis induction in colorectal cancer DLD1 cells, ovarian cancer A2780 cells, and clear cell renal cell carcinoma RCC4 cells, compared to apoptosis in cells treated with scrambled siRNA. As expected, silencing of IP3R3 and subsequent apoptosis induction resulted in increased levels of apoptosis in all these cells. Further, we prepared a DLD1/IP3R3_del cell line using CRISPR/Cas9 gene editing method. These cells were injected into nude mice and tumor's volume was compared with tumors induced by DLD1 cells. Lower volume of tumors originated from DLD1/IP3R3_del cells was observed after 12 days, compared to wild type DLD1 cells. Also, the migration of these cells was lesser compared to wild type DLD1 cells. Apoptosis under hypoxic conditions was more pronounced in DLD1/IP3R3_del cells than in DLD1 cells. These results clearly show that IP3R3 has proliferative and anti-apoptotic effect in tumor cells, on contrary to the pro-apoptotic effect of IP3R1.

Targeted antitumor therapy mediated by prodrug-activating mesenchymal stromal cells.

Mesenchymal stromal cells (MSCs) were introduced as tumor-targeted vehicles suitable for delivery of the gene-directed enzyme/prodrug therapy more than 10 years ago. Over these years key properties of tumor cells and MSCs, which are crucial for the treatment efficiency, were examined; and there are some critical issues to be considered for the maximum antitumor effect. Moreover, engineered MSCs expressing enzymes capable of activating non-toxic prodrugs achieved long-term curative effect even in metastatic and hard-to-treat tumor types in pre-clinical scenario(s). These gene-modified MSCs are termed prodrug-activating MSCs throughout the text and represent promising approach for further clinical application. This review summarizes major determinants to be considered for the application of the prodrug-activating MSCs in antitumor therapy in order to maximize therapeutic efficiency.

Complete regression of glioblastoma by mesenchymal stem cells mediated prodrug gene therapy simulating clinical therapeutic scenario.

Suicide gene therapy mediated by mesenchymal stem cells with their ability to engraft into tumors makes these therapeutic stem cells an attractive tool to activate prodrugs directly within the tumor mass. In this study, we evaluated the therapeutic efficacy of human mesenchymal stem cells derived from bone marrow and from adipose tissue, engineered to express the suicide gene cytosine deaminase::uracil phosphoribosyltransferase to treat intracerebral rat C6 glioblastoma in a simulated clinical therapeutic scenario. Intracerebrally grown glioblastoma was treated by resection and subsequently with single or repeated intracerebral inoculations of therapeutic stem cells followed by a continuous intracerebroventricular delivery of 5-fluorocytosine using an osmotic pump. Kaplan-Meier survival curves revealed that surgical resection of the tumor increased the survival time of the resected animals depending on the extent of surgical intervention. However, direct injections of therapeutic stem cells into the brain tissue surrounding the postoperative resection cavity led to a curative outcome in a significant number of treated animals. Moreover, the continuous supply of therapeutic stem cells into the brain with growing glioblastoma by osmotic pumps together with continuous prodrug delivery also proved to be therapeutically efficient. We assume that observed curative therapy of glioblastoma by stem cell-mediated prodrug gene therapy might be caused by the destruction of both tumor cells and the niche where glioblastoma initiating cells reside.

Characterization of mesenchymal stem cells of "no-options" patients with critical limb ischemia treated by autologous bone marrow mononuclear cells.

BACKGROUND: Application of autologous bone marrow mononuclear cells to "no option" patients with advanced critical limb ischemia (CLI) prevented major limb amputation in 73% patients during the 6-month follow-up. We examined which properties of bone marrow stromal cells also known as bone-marrow derived mesenchymal stem cells of responding and non-responding patients are important for amputation-free survival. METHODS AND FINDINGS: Mesenchymal stem cells of 41 patients with CLI unsuitable for revascularisation were isolated from mononuclear bone marrow concentrate used for their treatment. Based on the clinical outcome of the treatment, we divided patients into two groups: responders and non-responders. Biological properties of responders' and non-responders' mesenchymal stem cells were characterized according to their ability to multiply, to differentiate in vitro, quantitative expression of cell surface markers, secretion of 27 cytokines, chemokines and growth factors, and to the relative expression of 15 mesenchymal stem cells important genes. Secretome comparison between responders (n=27) and non-responders (n=14) revealed significantly higher secretion values of IL-4, IL-6 and MIP-1b in the group of responders. The expression of cell markers CD44 and CD90 in mesenchymal stem cells from responders was significantly higher compared to non-responders (p<0.01). The expression of mesenchymal stem cells surface markers that was analyzed in 22 patients did not differ between diabetic (n=13) and non-diabetic (n=9) patient groups. Statistically significant higher expression of E-cadherin and PDX-1/IPF1 genes was found in non-responders, while expression of Snail was higher in responders. CONCLUSIONS: The quality of mesenchymal stem cells shown in the expression of cell surface markers, secreted factors and stem cell genes plays an important role in therapeutic outcome. Paracrine mechanisms are main drivers in the induction of reparatory processes in CLI patients. Differences in mesenchymal stem cells properties are discussed in relation to their involvement in the reparatory process.

Human adipose tissue-derived mesenchymal stem cells expressing yeast cytosinedeaminase::uracil phosphoribosyltransferase inhibit intracerebral rat glioblastoma.

Prodrug cancer gene therapy by mesenchymal stem cells (MSCs) targeted to tumors represents an attractive tool to activate prodrugs directly within the tumor mass, thus avoiding systemic toxicity. In this study, we tested the feasibility and efficacy of human adipose tissue-derived MSCs, engineered to express the suicide gene cytosine deaminase::uracil phosphoribosyltransferase to treat intracranial rat C6 glioblastoma. Experiments were designed to simulate conditions of future clinical application for high-grade glioblastoma therapy by direct injections of therapeutic stem cells into tumor. We demonstrated that genetically modified therapeutic stem cells still have the tumor tropism when injected to a distant intracranial site and effectively inhibited glioblastoma growth after 5-fluorocytosine (5-FC) therapy. Coadministration of C6 cells and therapeutic stem cells with delayed 5-FC therapy improved the survival in a therapeutic stem cell dose-dependent manner and induced complete tumor regression in a significant number of animals. Continuous intracerebroventricular delivery of 5-FC using osmotic pump reduced the dose of prodrug required for the same therapeutic effect, and along with repeated administration of therapeutic stem cells increased the survival time. Intracerebral injection of therapeutic stem cells and treatment with 5-FC did not show any detectable adverse effects. Results support the arguments to begin clinical studies for treatment of high-grade brain tumors.

Stem cell based cancer gene therapy.

The attractiveness of prodrug cancer gene therapy by stem cells targeted to tumors lies in activating the prodrug directly within the tumor mass, thus avoiding systemic toxicity. Suicide gene therapy using genetically engineered mesenchymal stem cells has the advantage of being safe, because prodrug administration not only eliminates tumor cells but consequently kills the more resistant therapeutic stem cells as well. This review provides an explanation of the stem cell-targeted prodrug cancer gene therapy principle, with focus on the choice of prodrug, properties of bone marrow and adipose tissue-derived mesenchymal stem and neural stem cells as well as the mechanisms of their tumor homing ability. Therapeutic achievements of the cytosine deaminase/5-fluorocytosine prodrug system and Herpes simplex virus thymidine kinase/ganciclovir are discussed. In addition, delivery of immunostimulatory cytokines, apoptosis inducing genes, nanoparticles and antiangiogenic proteins by stem cells to tumors and metastases is discussed as a promising approach for antitumor therapy. Combinations of traditional, targeted and stem cell-directed gene therapy could significantly advance the treatment of cancer.