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Dendritc cells (DCs) are professional antigen-presenting cells. Dendritic cells in the periphery capture and process antigens, express co-stimulatory molecules, migrate to lymphoid organs such as lymph nodes and secrete cytokines to cause immune responses. There are successful models of DCs mediated immunotherapy of cancer. The cancer immunotherapy aims to induce tumor specific effector T cells that can reduce the tumors, and also activate tumor-specific memory T cells that controls the tumor. The DCs are capable of regulation of the T-cell mediated immunity. This attribute of DCs can be developed as adjuvant in cancer immunotherapy.
Keywords: Dendritic cell, T cell, immunostimulation, tumor antigen.
Dendritic cells (DCs) are a heterogeneous group of professional antigen-presenting cells (APCs). DC undergoes maturation in their development, upon interaction with the signal, which expresses co-stimulatory molecules such as B.7 on their surface. It has been showed that the DC possess the ability to prime T cell to maintain the immune response against viruses, bacteria, foreign particles and tumor cells (1).
Cancer is one of the leading death causing disease worldwide and is responsible for 13% of deaths in 2007. The development of cancer includes uncontrolled growth of the cells followed by metastasis which makes it more life-threatening. Current studies using DC in cancer immunotherapy are in the interesting phase. The efficiency of the pulsed DCs with tumor specific or tumor associated antigen (TAA), in animal models as well as in human clinical trials against cancer has been assessed. Gong et al. showed that the fusion of DCs with patient derived tumor cell induces activation of the T-cell against specific tumor (1). Other approach by Xing et al; explains the usefulness of DCs in the treatment of cancer. The fusion of DCs and Herpes Simplex Virus/ Ganciclovir(HSV TK/GCV) induced dying cell results in antitumor activity (2).
Generation and Development of DCs:
Dendritic cells are ideally positioned in the body parts having large external environmental interface. This gives them the ability to encounter and process antigen easily. The DCs are mainly divided into two types: conventional DCs (cDCs) and plasmocytoid DCs (pDCs) (1-4).
In the first step, DCs derived from hematopoietic bone marrow progenitor cells are immature and express pattern recognition receptors (PRRs) knows as Toll-like receptors (TLR) on their surface. The pDCs are found to have TLR-7, TLR-9 while cDCs possesses TLR-4. The DCs process and present antigen on the surface using Major Histo-compatibility Complex I and II (MHC) and upregulate costimulatory molecule such as CD80 (B7.1), CD83, CD86 (B7.2) and CD40 which act as ligand for CD28 present on T cell and assist in the activation of T cell (1-6). Granulocyte Monocyte- colony stimulating factor (GM-CSF) is also involved in the maturation and activation of other antigen presenting cells such as macrophages and monocyte (3). The result of chromium release assay proves that there is marked increase in cytotoxic T cells in GM-CSF secreting cells. This shows that GM-CSF causes significant improvement in maturation and activation of DCs. The other factor which is found to have significant effect on DC development is FLt3 tyrosine kinase (4). Claudia et al. showed that Flt3 tyrosine kinase plays important role in DCs homeostasis in the periphery (4).
Immunomodulation of DCs for cancer immunotherapy:
Immature DCs engulf antigen and undergo maturation. These mature DCs leave their site of maturation and move on to the secondary lymphoid organ such as spleen and present antigen to T and B cell. DC processes this antigen by two different types: exogenous pathway and endogenous pathway. In exogenous pathway, DCs present antigen using MHC-I to CD4 cell whereas in endogenous pathway, DCs present antigen using MHC-II to CD8 or CTL cell (3,4). The aim of cancer therapy using DCs involves eliciting this T cell activation against tumor antigen. It can be achieved by loading of DCs with the patient specific tumor antigen, by fusion of DCs and tumor cell. This fused DC/Tumor cell will then be incubated with the T cell. This cause priming of T-cell due to proper presentation of antigen via fused cell resulting into anti-tumor response.
The generation of DCs for cancer immunotherapy involves isolation and culture of the hematopoietic CD34+ progenitor cells with GM-CSF and IL-4 (5). This isolation of DCs can be done using Ficoll-Hypaque density-gradient centrifugation. The maturation of the obtained DCs can be carried by various methods. The commonly used method involves incubating DCs with antigen specific tumor cells. It consist tumor specific antigen or tumor associated antigen (TAA). TAA consists of RNAs, tumor peptides, exosomes, and tumor lysates. One or two TAAs are insufficient for eliciting anti-tumor activity. Shigeki Ohata et al. developed an experimental model for preclinical studies of cancer immunotherapy (5).They showed that Bone marrow (BM) derived DCs from common marmosets express CD11c, which is similar to human. They found that GM-CSF, IL-4 and TNF-ï¡ cytokines have significant effect on maturation of DCs (5). In this study, yield of BM derived DCs is high as compared to that of DCs derived from spleen and peripheral blood, which should be sufficient of clinical studies.
Loading DCs with tumor antigen
So far there are two trends for loading of DCs. One is using TAA peptide, RNA for pulsing DCs or using entire tumor cell for pulsing DCs. The later is found to be more effective in pulsing DCs. Moreover these are just loading of DCs and do not consider any MHC haplotype specificity. Jianlin Gong et al., showed that marked inhibition of mucin-1 (MUC-1) specific tumor when treated with MUC-1 specific T cell induced by MUC-1 transfected DCs. It confirms that there is need of tumor specific loading of DCs (1).
Models for DCs Immunotherapy:
There are many studies available on how the pulsing of DCs can be done for cancer treatment. It can be achieved by pulsing DCs with tumor peptide, tumor RNA or DNA. This approach results in narrow range immunogenicity of DCs which can be evaded by tumor cells. This can be circumvented by fusion DCs with entire tumor cell which will posses wide range of TAAs and can be effectively used to target tumor. Here we discussed the two studies that show the effective ways of using antigen pulsed DCs for tumor therapy.
First, Gong et al. showed that fusion of the cancer cell and DCs can efficiently prime T cell (1). In this study they isolated Peripheral blood mono-nuclear cells (PBMC) from breast cancer patient and gown them with granulocyte-macrophage colony-stimulating factor (GM-CSF) and IL-4. It causes expression of MHC-I and MHC-II along with other co-stimulatory molecule such as CD80, CD83 on the surface of DCs except MUC1. The fused DCs/breast tumor (BT) expresses both MUC-1 and MHC-II along with MHC-I and other molecules. This confirms the formation of heterokaryon of fused DCs/BT (Fig.1). This fused DCs/BT is co-cultured with autologous T cell.
The DCs/BT response for activation of T cell was found to be much higher than as compared to only DCs, DCs mixed with BT cell and only BT cells. The cytotoxicity of T cell against BT cell was markedly high as compared to other controls. In their later studies they showed that the effectiveness of fusion of DCs is independent of HLA type of tumor.
The dying apoptotic or necrotic cells can be derived ex vivo by using irradiation of heat shock proteins and can found to induce specific cytotoxic activity (2). But this procedure cannot be used for clinical studies. This problem can be circumvented by fusion of DCs with dying cell induced by HSV-TK. HSV-TK itself is not safe to use in vivo due to its irregular and uncontrolled expression.
Secondly, Wei Xing et al. showed that certain changes during fusion of DCs with tumor cell increases its effectiveness (2). The fusion of DCs with dying tumor cells produced with HSV-TK along with GCV provides new approach for clinical studies of cancer treatment. The HSV-TK/GCV system is found to have marked effect on cell death by apoptosis and necrosis as compared to controls (Fig.2). In this model this DCs/HSV-TK/GCV is used I vivo for activation of CTL activity.
Role of Interferons (INF):
Interferon- ï§ï€ is secreted specifically by the Th1 cell, DCs and NK cell which help in both innate and adaptive immunity. INF-ï§ also activate macrophages and up regulate pro-inflammatory factors such as CXCL9, CXCL 11 (7).Priming of DCs is important activity of INF-ï§. Loredana Frasca et.al proved that INF-ï§ï€ is highly important in maturation and activation of DCs. Their experiment shows that INF-ï§ helps maturation of DCs by up regulating IL-27 and IL-12 (7).
Route of administration and efficiency of DCs treatment:
Route of administration can affect efficiency of tumor cell pulsed DCs treatment. The fusion of DCs with tumor specific antigen causes maturation of DCs. This maturation of DCs will result in upregulation of co stimulatory molecule such as CD80, CD86 and CD83 and adhesion molecules such as LFA-1, CD62. The adhesion molecules are necessary for interaction DCs with High Endothelial Venules (HEV). Fong et.al showed that this DCs response is independent of route of administration (8).
Conclusion and future prospect:
DCs are an attractive target for therapeutic manipulation of the immune system to increase otherwise insufficient immune responses to tumor antigens. The studies on DCs using murine model and clinical studies against caner have proved that it can be used as an efficient tool in development of novel cancer immunotherapy. In vivo activation and targeting of DCs, as well as exploitation of DCs to reduce immune responses, will expand the target use of DCs in treatment of variety of immune-mediated diseases.