Mesenchymal Stem Cells : vector for targeted cancer therapy

Mesenchymal stem cells (MSCs) have been studied extensively due to their potential to differentiate to cell types of varying lineages.Adipose tissue and umbilical cord blood are two tissues frequently used to obtain MSCs. Due to tumor tropism of MSCs and their ability to protect encoded cytotoxic genes, MSCs have garnered interest as a potential vector for targeted therapy, with limited damage to normal tissues. The tumor microenvironment plays a critical role in ensuring the survival of cancer cells through promotion of MSCs to differentiate into cancerassociated fibroblasts (CAFs), which promote tumor growth and metastasis. Through specific interactions between ligands and receptors expressed on MSCs and cancer cells, respectively,MSCs can home to necrotic tissues or inflamed sites in the body, including the tumor microenvironment. In fact, an inflammatory tumor environment is similar to a wound healing environment. This review discusses the preeminent characteristics of MSCs and their influence ontumor cell growth and metastasis. MSCs may represent an encouraging platform for cancer treatment. The combination of MSC and gene therapy represents a potentially outstanding strategy to specifically target and effectively destroy tumor.


Introduction
Cancer continues to be a huge disease burden across the globe, even despite more improvements in prevention, diagnosis and treatment of cancer.One of the main obstacles in cancer treatment remains the fact that the cancer cells cannot be completely destroyed and that specific target of cancer cells is still incomplete.
Conventional cancer therapies remain limited.Surgery is invasive and only effective with large tumors; but most cancers are known to become metastatic and surgery is futile for those metastatic cancer cells in the bloodstream).standard cell source in research applications.However, the process of BM-derived MSC acquisition is highly complicated, which adversely affects the ability to receive MSC donors (Bentzon et al., 2005;Kern et al., 2006;Mueller and Glowacki, 2001).Therefore, there is a need for alternative sources of MSCs, without greatly affecting MSC number, differentiation potential and lifespan.
Adipose tissue and umbilical cord blood are the two most promising tissues.Adipose tissue has the following advantages: (i) it is usually obtained from subcutaneous tissue and represents reduced invasion; (ii) it maintains MSCs called adipose-derived stem cells (ADSC), which is considered to be an autologous stem cell source in personalized cell-based therapies.Mononuclear cell populations obtained from umbilical cord blood have been shown to also be a rich source of MSCs.Umbilical cord blood derived MSC have two main advantages: (i) the number of stem cells in cord blood are higher than other adult tissues; and (ii) the majority of stem cells in cord blood are naive, which may be due in part to higher telomerase activity (Chang et al., 2006), thus they have higher proliferation (Goodwin et al., 2001).MSCs derived from adipose tissue or umbilical cord blood are mostly identical with those from BM (Bieback et al., 2004;Erices et al., 2000).
Particularly, MSCs have characteristics that are superior to other types of stem cells, making MSCs rise as a superior candidate for stem cell therapy.These characteristics include: (i) MSCs are adult stem cells and therefore their application in autologous therapy is easily implemented and with reduced possibility of causing graft versus host disease (GvHD).Even in the case when autologous MSCs are not available, umbilical cord blood can serve as a qualified source for acquiring MSCs to downregulated (Stagg, 2007).
MSCs have the ability to home to the tumor location through a specific interactions between ligands and their respective receptors expressed on MSCs and on tumor cells in their microenvironment (Massague et al., 2000;Mishra et al., 2005).Many researchers have been focusing on the use of MSCs as vehicles to deliver targeted cytotoxic agents to cancer cells.This innovative approach has several advantages: (i) compared to naked or polymer-enclosed anti-cancer drugs, MSCs can deliver drugs to the desired location without damaging normal tissues; (ii) MSCs can secrete drugs through exosomes, which enable drug delivery to take place over a longer period, avoiding acute severe side effects associated when a high dose of drugs is injected in the body.This allows the lifetime of the drug flow to be extended, thereby restricting repeated drug delivery into the body in order to achieve clinical efficacy.

MSCs have been collected from various tissues.
There are many reports on the expression of molecular markers on their cell surface which makes it easier to acquire and evaluate the MSC population.and CXCR4 (Corcoran et al., 2008), and through GD2 and CD2, expressed on bone marrow mesenchymal stem cells (BM-MSCs) (De Giorgi et al., 2011).
MSCs are attracted to tumors because the tumors have a similar environment as wound healing.
MSCs increase the motility of cancer cells by activating ADAM10 (also known asCDw156 or CD156c) (Dittmer, 2010).TNF-alpha is the most potent inflammatory cytokine present in the tumor microenvironment.survival (Sasportas et al., 2009).In addition to virus-based gene delivery system, it is feasible to use non-viral vectors to transfer therapeutic genes to MSCs (Hu et al., 2012).bearing mice (Hu et al., 2011).Lentivirus-mediated TNFalpha-expressing MSCs induced apoptosis in cancer cells via antiangiogenic effects (Zhang et al., 2011).Moreover, MSCs and protein vaccination, in combination, demonstrated a significant inhibition of tumor growth and lung metastasis (Wei et al., 2011).suppress tumor growth (Uchibori et al., 2009) and significantly prolong survival of tumor-inoculated animals (Bak et al., 2010), by creating a toxic tumor-specific environment without transduction of suicide gene to normal organs (Conrad et al., 2011).
Angiogenic prevention is a way to limit tumor Prevention of adipocyte differentiation is one of approach that cancer cells use to control MSC differentiation into cell types which support tumor cell homing and progression.High presence of matrix metalloprotease-9 (MMP-9) and urokinase plasminogen activator (uPA) in cancer cell-conditioned media confirmed this effect (Xu et al., 2009).MSCs also upregulate MMP-2 and MMP-9 in prostate cancer cells, which in turn promote cancer cell proliferation, migration and invasion (Ye et al., 2012).
Cancer cells interact with each other and with the surrounding environment through direct interaction; additionally, communication among cells in the tumor microenvironment is mediated through soluble molecules.
These molecules can be directly secreted and have autocrine or paracrine influence once they interact with compatible receptors expressed on target cells.When surface receptors are not available, to ensure that the signal to stimulate cancer cell proliferation is maintained, cancer cells secrete these molecules through exosomes.
Exosomes supply a diverse source of cytokines, chemokines, and growth factors, and enter target cells more easily through membrane fusion.Ovarian cancer-, breast cancer-, gastric cancer-derived exosomes contribute to the development of tumor-associated myofibroblasts, derived from MSCs.The exosomes also increase expression of tumor-promoting factors, SDF-1, VEGF, CCL5 and TGF-beta, by activating both SMADdependent and SMAD-independent intracellular signaling pathway (Cho et al., 2011;Cho et al., 2012;Gu et al., 2012).which significantly enhanced the migration of mammary cancer cell lines (Halpern et al., 2011).
Tumor associated MSC (TAMC) secrete protumoral cytokines; of these, CXCL12 is considered to display strong protective effects through CXCL12/CXCR4 interaction in ovarian cancer cells (Lis et al., 2011).TGFbeta-mediated salivary gland cancer cells recruit MSCs and, in turn, MSCs disperse cancer cell connections and reduce the expression of E-cadherin in cancer cells.
Donor MSCs can inhibit tumor progression and prevented osteoclastogenesis (Chanda et al., 2009;Secchiero et al., 2010).Human MSC-conditioned medium downregulate expression of VEGF in tumor cells, thereby suppressing tumorigenesis and tumor angiogenesis (Li et al., 2011).During co-culture, human umbilical cord mesenchymal stem cells (HUMSCs) induced apoptosis of breast cancer cells through both cell-cell contact and formation of a novel cell-in-cell phenomenon.In the animal models, HUMSC injection also efficiently inhibited metastatic breast cancer (Chao et al., 2012).Xu, F., Gomillion, C., Maxson, S., and Burg, K.J. (2009).In vitro interaction between mouse breast cancer cells and mouse mesenchymal stem cells during adipocyte differentiation.
adequate quantity.In addition, since MSCs have immunosuppressive properties, one can use allogenic MSCs with appropriate HLA compatibility; (ii) in vivo MSCs are involved in the recovery of damaged tissues because of the ability to secrete and recruit tissue regenerative stimulants.Due to their trophic nature to direct factors to damaged tissues, MSCs have been increasingly evaluated as a potential vector for targeted therapies, with minimal effects on normal tissues; (iii) MSCs are one of the few kinds of stem cells which can be obtained from the body's tissues, cultured and manipulated in vitro, and then transplanted back into the body without causing serious complications, and under appropriate control.In addition, manipulation of MSCs is easy, without being extensively time-, labor-, and costconsuming.Prochymal is a MSC-containing product approved in Canada, which is also the first product in the world applying stem cell therapy in the disease treatment.Although these MSCs are derived from allogenic individuals their efficiency remains high, as expected, due to the fact that MSCs have the ability to suppress immune responses.MSC grafts modulate the immune system via cell-cell interaction and through soluble factors.MSCs secrete immunosuppressive molecules, including hepatocyte growth factor, prostaglandin E2, TGF-β1, indoleamine 2,3-dioxygenase, nitric oxide and IL-10.Contact-dependent mechanism, via B7H1 (also known as PD-L1 or CD274) interactions, can also explain MSCmediated immune suppression.As a result, the function of a broad range of immune cells, including T cells, B cells, Exposure to BM-MSCs to TNF-alpha promotes locomotion of cancer cells through CXCR3 ligand(Shin et al., 2010).IL-1alpha, secreted by inflammatory cells, also induces MSCs to promote the growth of cancer cells(Cheng et al., 2012).TGF-beta1-dependent immunosuppression is one of mechanism by which MSCs support tumor growth.MSCs increase the frequency of regulatory T cells (Tregs), and as a result, NK cell and CTL functions are inhibited and granzyme B production is decreased(Patel   et al., 2010).MSCs migrate and differentiate into tumorstroma cells, including CAFs and even vascular endothelial cells, which then enhance angiogenesis in the tumor microenvironment(Albarenque et al., 2011;Shinagawa et al., 2010;Zhang et al., 2010).Cancer stem cells, which are also known as tumor-initiating cells (TICs) are responsible for tumor initiation, maintenance and metastasis.Tumor-derived MSCs significantly increase tumor initiation caused by TICs, even transplanted TICs (Lanza et al., 2012).Through expression of matrix metalloproteinases 1 and 3, BM-MSCs support metastasis of invasive cancer cells.They also efficiently chemoattract endothelial cells (Zhao et al., 2012).MSC/gene therapy combination: "Trojan Horse" to target specific tumor locations MSCs represent an outstanding vehicle for gene delivery.Cytosine deaminase (CD), which converts prodrug 5-FC (5-fluorocytosine) to toxic 5-FU (5fluorouracil), is one cytotoxic gene that has been extensively investigated in gene therapy applications.Interestingly, MSCs engineered to deliver CD (CD-MSC) were not sensitized to 5-FC, thus showing they can overcome the often deleterious effect of a suicide gene expression by a cellular vehicle.CD-MSC can deliver the transgene to the site of tumor formation and mediate strong antitumor responses (Kucerova et al., 2007).Lentiviral (LV)-transduced MSCs do not show altered features in cell growth, differentiation capacity or migration preferences (Kallifatidis et al., 2008).MSCs delivering recombinant tumor necrosis factor-related apoptosisinducing ligand (TRAIL) showed profound anti-tumor effects without affecting their "stem-like" properties or MSCs act as a shield to protect the tumoricidal gene therapy vector in the bloodstream.Compared to viral-based gene transfer methods, transduction of an adenoviral vector expressing TRAIL into MSCs showed stability in circulation, as well as induction of high levels of virus-neutralizing antibodies(Mohr et al., 2008).Importantly, TRAIL-engineered MSCs can produce anticancer agents locally and continuously(Moniri et al.,   2012), resulting in decreased bone and lung metastases(Reagan et al., 2012).Adipose-derived mesenchymal stem cells (ADSC) are available for personalized medicine.ADSC have also been engineered to express TRAIL, and were capable of mediating malignant cell apoptosis without any significant or apparent toxicity to normal tissues(Grisendi et al., 2010).MSCs armed with TRAIL migrate to tumor and reduce tumor growth.Interestingly, MSC/gene combined therapy has a synergistic effect when combined with traditional treatments(Loebinger et al., 2010).Adeno-associated virus (rAAV) have also been evaluated for gene modification of MSCs, including to constitutively express interferon (IFN)-beta.The most notable results of IFN-beta-expressing MSC therapy was the ability to trigger anti-cancer immune activity(Ren et al., 2008).Particularly, high levels of IFN-beta were detectable only in the tumor microenvironment, and normal levels of dendritic cells (DC), CD8 + T cells and CD4 + /Foxp3 + regulatory T-cells (Tregs) were detected.Cancer cell-replication-related signaling pathways, including those of Stat3, Src, Akt, cMyc and MMP2, are down-regulated.As a result, primary tumor growth and metastasis are inhibited(Ling et al., 2010).The amniotic fluid plays a very important role in the development of the fetus.Human amniotic fluid-derived mesenchymal stem cells (AF-MSCs) were successfully expanded and manipulated by lentivirus to express interferon beta (IFN-beta).The gene modified AF-MSCs had the ability to reduce tumor growth(Wang et al., 2012), as well as prolong survival of tumor-bearing mice(Bitsika et al., 2012).Immunotherapy has many advantages in cancer treatment; perhaps the most important one is the ability to harness and activate the immune system to destroy cancer cells.Immunotherapy remains still a relatively new therapy compared to conventional therapies.The strategy relies on effector cells to generate immune reactions strong enough to eliminate the tumor, but without causing hypersensitivity or autoimmune reactions.Immunotherapy includes the cellular components, such as effector cells, and non-cellular components, such as cytokines and antigens, which stimulate immune responses.Many kinds of cytokines have been used in immune therapy but their anti-tumor therapeutic efficacy is still low.This is due to the fact that most cytokines are injected into the bloodstream, causing systemic toxicity.Umbilical blood mononuclear cell (UBMC)-derived mesenchymal stem cells (UBMC-MSCs) expressing IL-21 showed delayed tumor growth and prolonged survival in ovarian-cancer- Suicide gene therapy have been attracting great attention from the scientific community.Its principle is based on therapeutic (usually cytotoxic) gene transfer to specific tumor locations in the body.A harmless prodrug is administered into the body; when the pro-drug reaches the tumor site, enzymatic activity encoded from the suicide gene converts the prodrug into a toxic drug, which then destroy cancer cells at the site.MSCs can be engineered to selectively express herpes simplex virusthymidine kinase (HSV-tk) gene; these engineered MSCs accumulate at the tumor site.Following ganciclovir infusion, MSCs expressing HSV-tk are activated and can growth and control cancer cell metastasis.Endostatin and angiostatin are potent inhibitors of tumor angiogenesis.Endostatin-engineered MSCs can overcome the limitation of short half-time and toxicity related to virus-mediated endostatin gene therapy(Jiang et al., 2010).Zheng et al.    demonstrated that adenoviral transduction of human placenta-derived mesenchymal stem cells (hpMSCs) to deliver endostatin has many promising results; for instance, the toxic effects were specific to the tumor site with significantly decreased blood sprouts and dramatically increased tumor apoptosis index(Zheng et al., 2012).Tumor vasculature plays an extremely important role in tumor cell maintenance and metastasis.Antiangiogenesis mediators contribute to new era of hope for cancer patients.Expression of pigment epithelial-derived factor (PEDF), a potent anti-angiogenesis mediator, did not adversely affect MSC biology.MSC-MDA7 reduced tumor growth.Moreover, MSC-PEDF reduced growth in even more aggressive cancer models, and were able to completely prevent prostate tumor establishment in vivo(Zolochevska et al., 2012).The interaction between tumor cell and MSCs: The interaction between two opposite charges Stimulated by molecules secreted in tumor microenvironment, human adipose tissue-derived mesenchymal stem cells (hADSCs) differentiate into cancer-associated myofibroblasts or fibroblasts, which play a pivotal role in tumorigenesis.Cancer-derived lysophosphatidic acid (LPA) induces the differentiation process through multiple signaling pathways involving TGF-beta1-Smad (Jeon et al., 2008), Rho kinase, ERK, PLC and phosphoinositide-3-kinase (Jeon et al., 2010).Once MSCs differentiate into osteoblasts, CCL2 level is significantly elevated.The bone-derived CCL2 induces migration of breast cancer cells.After co-culture with MSCs, breast cancer cells also overexpress genes involved in CCL2-mediated signaling pathways (Molloy et al., 2009).
The impact of isolation and culture of MSCs on cancer cells is unclear.Studies have focused on identifying how conditioned media-derived from MSCs alters the characteristics of the cancer cells.Halpern et al. analyzed MSC-conditioned media and revealed the presence of numerous cytokines, most notably CXCL1 and CXCL5, The existence of cancer stem cells (CSCs) has been demonstrated.Like all tissues that contain stem cells, the tumor consists of heterogeneous stem cell populations.The CSCs are organized based mainly on the state of cell differentiation, e.g. on expression of pluripotency markers, such as aldehyde dehydrogenase.MSCs can accelerate tumor growth by increasing the breast CSC population, through cytokine loops involving IL6 and CXCL7.Primary human breast cancers and tumorxenografts represent MSC-CSC niches, enhancing drug resistance and pluripotency through activation of the IL-6/JAK2/STAT3 pathway(Hsu et al., 2012;Liu et al., 2011a).Carcinoma-associated MSCs (CA-MSCs) have been identified in a majority of human ovarian tumor samples.CA-MSCs increase expression of BMP2, BMP4 and BMP6, which promote tumor growth by increasing the number of CSCs(McLean et al., 2011).Cancer cells and MSCs can interact in a reciprocal fashion.For example, cancer cells secrete interleukin-1 (IL-1), which stimulates prostaglandin E(2) (PGE(2)) secretion from MSCs.IL-1 and PGE-2 induce MSCs to produce a variety of cytokines.MSC-secreted PGE-2 and cytokines , in turn, can act on cancer cells to induce activation of beta-catenin signaling and formation of cancer stem cells(Li et al., 2012).Co-culture of cancer cells with MSCs increased the population of CD133+ gastric carcinoma cells (known as CSCs) through the wingless-type MMTV integration site (WNT) family member 5A (WNT5A) and transforming growth factor-beta (TGF-beta)-induced (TGFBI) genes (Nishimura et al., 2012).The epithelial-mesenchymal transition (EMT) was detected very early during embryonic development; the EMT process helps facilitate cell rearrangement and organ formation.The process of carcinoma development through metastatic initiation is thought to mimic the EMT process.Cell-cell connections, via firm bonds, are what limits movement of epithelial cells.In a microenvironment where nutrients and oxygen are deficient, as in the tumor microenvironment, the epithelial-like cancer cells undergo EMT to establish mesenchymal-like cancer cells, liberating the cell-cell and cell-ECM interactions, and initiating metastasis.Cancer cells which undergo EMT are considered to be tumorinitiating cells.MCF7 cells underwent EMT after co-culture with human adipose-derived MSCs (hAD-MSCs).The mechanism which MSCs induce cancer cell EMT is primarily mediated by transforming growth factor-beta1(TGF-b1)(Xu et al., 2012).It is obvious that the tumor microenvironment (which is nutrient-deprived and hypoxic) and the inflammatory factors present in the microenvironment play a pivotal role in enhancing the hypoxia-inducible factor 1alpha (HIF-1alpha)-dependent pathway.IFN-gamma and TNF-alpha, known as two of most potent inflammatory molecules, further accelerate tumor angiogenesis through VEGFproduced by MSCs(Liu et al., 2011b).Breast cancer-associated MSCs secrete EGF and provide a favorable microenvironment for tumor cell growth via the EGF/EGFR/Akt pathway(Yan et al., 2012).Anti-tumor cell property of MSCsAlthough many studies have showed that MSCs support tumor formation, survival and malignancy, other reports have indicated that MSCs have the ability to kill cancer cells.The latter observation came from studies of MSCs (isolated from cancerous tissues) which had the ability to promote tumor growth.By chemoattractant mechanisms and employment of molecules generated from MSCs, cancer cells sustain growth and escape from attack by the immune system.MSCs isolated from healthy tissues have become a powerful tool for gene therapeutic transfer to cancer tissues based the ability of MSC to show tumor tropism.MSCs have an inhibitory effect on MCF-7 cells as well as K562 cells, via expression of dickkopf-1 (Dkk-1, a well-known negative regulator of WNT signaling pathway) (Qiao AbbreviationsMSCs Mesenchymal stem cells; CAFs cancerassociated fibroblast; ES embryonic stem cells; iPS induced pluripotent stem cells; BM bone marrow; ADSC adipose-derived mesenchymal stem cell; GvHD graft versus host disease; MHC major histocompatibility complex; GF growth factor;SDF-1α stromal cell-derived 1α; ECM extracellular matrix; MMP matrix metalloproteinase; BMP bone morphogenic protein; Tregs regulatory T cells; TIC tumor-initiating cells; CD cytosine deaminase; 5-FC 5-fluorocytosine; 5-FU 5-fluorouracil; LV Lentiviral; TRAIL tumor necrosis factor-related apoptosisinducing ligand; IFN interferon; DC dendritic cell; UBMC Umbilical blood mononuclear cell; HSV-TK herpes simplex virus-thymidine kinase; IL interleukin; TGF-b transforming growth factor-beta; EMT epithelialmesenchymal transition; VEGF vascular endothelial growth factor; bFGF basic fibroblast growth factor; TICs tumor-initiating cells; BM-MSCs bone marrow