7.1 Overview of Immunotherapy for Melanoma
Tumor immunotherapy is a method to achieve the purpose of inhibiting tumor growth and removing tumor cells in an active or passive way to enhance the body's anti-tumor immune response by applying immunological principles.It has gradually become a research hotspot in the medical field since 2011（Figure 7-1）.Tumor immunotherapy mainly consists of three parts：①Non-specific immunotherapy，mainly refers to the use of recombinant interleukin-2（IL-2），interferon and other cytokines，to activate or promote the killing function of the patient's immune cells.②Active specific immunotherapy，mainly refers to the use of tumor cells，tumor proteins，peptides or tumor carriers to make tumor vaccines to immunize patients，to activate autoimmunity and enhance anti-tumor effect.③Passive immunotherapy，including adoptive cell immunotherapy and monoclonal antibody therapy（see extended reading 1）.
The generation of anti-tumor immune response，which is known as the tumor immune cycle，is a series of progressive events from the release of tumor antigens to tumor-specific immune generation（Figure 7-2）.In brief，necrosis or apoptosis of tumor cells induces the release of tumor antigens，which are usually captured by antigen-presenting cells（APCs）.Then，APCs present antigenic peptides in major histocompatibility complex Ⅰ（MHC-Ⅰ）and major histocompatibility complex Ⅱ（MHC-Ⅱ）to activate immature T cells in lymph nodes.Activated T cells enter the tumor microenvironment（TME）and target tumor cells through the interaction between T cell receptors（TCRs）and MHC antigen-peptide complexes.Once being recognized，tumor-specific T cells can eliminate tumor cells by inducing apoptosis.The death of tumor cells leads to the release of additional tumor antigens，which further enhance the subsequent revolutions of this tumor immune cycle.Many factors that come into play in the tumor immune cycle provide a wide range of potential therapeutic targets for the treatment of tumors（see extended reading 2）.
The drugs approved by the Food and Drug Administration（FDA）of the United States for melanoma immunotherapy are IL-2 and immune checkpoint inhibitors，such as ipilimumab（trade name：Yervoy；target：cytotoxic T lymphocyte-associated protein 4，CTLA-4），pembrolizumab（trade name：Keytruda；target：programmed cell death protein 1，PD-1）and nivolumab（trade name：Opdivo；target：PD-1）.Please refer to chapter 2，section 2 for details in the previous article.
In addition to the US FDA-approved immunotherapies，several new melanoma immunotherapies are in research phase，such as the monoclonal antibody varlilumab，the monoclonal antibody MGA271，cytokines and tumor vaccine（see extended reading 3）.For example，the monoclonal antibody varlilumab，which targets CD27 to enhance the immune response of T cells and natural killer（NK）cells，is used in the treatment of tumors such as melanoma.CD27 is a costimulatory receptor of T cells and a member of tumor necrosis factor α（TNF-a）receptors superfamily.It plays an important role in the survival and activation of T cells and in the proliferation and cytotoxic activity of NK cells.Varlilumab is currently in phase Ⅰ of clinical trials for the treatment of metastatic melanoma.MGA271 is a monoclonal antibody against B7-H3，which is currently in phase Ⅰ of clinical trials for the treatment of melanoma.B7-H3，a member of B7 receptors family，expresses in APCs and other nonlymphoid tissue cells.The regulatory function of B7-H3 on T cells is still controversial.Some studies have shown that B7-H3 is a costimulatory receptor，while others have shown that B7-H3 has inhibitory effect on the receptor.In addition，some cytokines，such as granulocyte-macrophage colony-stimulating factor（GM-CSF），IL-2，interleukin-15，（IL-15），interleukin-18（IL-18），interleukin-21（IL-21），have anti-tumor effect and are currently in clinical trials for the treatment of melanoma.
The tumor vaccine is another important part of the melanoma immunotherapy，and some research has been done.Antigens on the surface of tumor cells are usually modified through a variety of mechanisms to inhibit the recognition by immune cells in the body，which hinders the clearance of tumor cells by the body.The melanoma vaccine is made of melanoma antigen such as tumor cells，tumor-associated proteins or peptides，genes that express tumor antigens.These vaccines are collected by APCs and presented to immune cells，further sensitize and activate T lymphocytes to overcome the immunosuppressive state caused by tumors，enhance immunogenicity and produce antigen-specific cellular immune response to kill melanoma cells，thereby inhibiting the growth，metastasis and recurrence of melanoma.MAGE，gp100，Melan A and NY-ESO are commonly used tumor antigens for the preparation of melanoma vaccine.They can activate the tumor-associated antigen（TAA）specific immune response to induce the anti-melanoma effect in the body，and they are expected to be effective targets for melanoma immunotherapy.
DCs are the most powerful antigenic antigens，which are thought to be the promoter of tumor immunotherapy as well as an important target for tumor immunotherapy，and they play a key role in the innate and adaptive immune activation.DCs are mainly used in two ways to mediate antigens presentation.In one way，DCs obtain exogenous antigen proteins through the cytoplasm.Then，the proteins molecules were decomposed into polypeptide fragments in the lysosomes and were proposed to the CD4+auxiliary T（T-helper，Th）cells，such as CD4+T lymphocytes by MHC-Ⅱ.In the other way，the endogenous antigen proteins in the DCs are degraded by protease.The peptide fragments of the degradation were combined with MHC-Ⅰ molecules and were proposed by the CD8+T lymphocytes to activate the antigen-specific cell toxicity T lymphocytes（CTLs）.CTLs can infiltrate tumor tissue and identify the MHC-Ⅰ antigen complexes expressed on the surface of the tumor cells，thus kill tumor cells.It is an important way to achieve effective immune treatment against tumors that tumor-specific CTLs are induced and play specific killing effect on tumor cells.Moreover，the activation of CD8+T lymphocytes not only directly kill tumor cells，but also obtain a persistent memory phenotype，which is of great significance to prevent the recurrence of tumors.In addition，CD4+T lymphocytes are also essential to the effective anti-tumor response for its effect of enhancing the cloning of CTLs and promoting the production and maintenance of memory phenotypes.DCs induce the production and activation of Th cells and CTLs by the antigen presentation of MHC-Ⅰ/Ⅱ，which has become the theoretical basis for the clinical trials of antigen based vaccines.
According to the antigen of melanoma，the melanoma vaccines can be divided into the antigen peptide vaccine，the tumor-associated antigen vaccine and the DNA vaccine.The vaccine，formed by polypeptide as an antigen，is called the melanoma antigen peptide vaccine，which can activate the patient's immune system by activating DCs，thus produce active immunity against melanoma cells.The melanoma antigen peptide vaccine has the advantages of high specificity and low toxicity in the process of killing tumor cells.TAA is an antigen protein produced by tumor cells that can trigger an immune response in the host.The tumor cell lysate（TCL）is used as a tumor antigen for the development of tumor vaccines.Compared with specific proteins or polypeptide tumor antigens，the antigen of the whole tumor cell source has many advantages as a vaccine.It can induce the activation of CTLs and CD4+T lymphocytes to provide a comprehensive source of potential tumor antigen，which can target tumor cells simultaneously，thereby avoiding the immune privilege caused by the absence of tumor antigens.However，the soluble tumor cell lysate that contains antigen and cell factor is essentially unstable.The TCL is easy to produce problems such as poor DCs intake，low antigen cross delivery efficiency and limitation of the induction of CTLs.(https://www.daowen.com)
Gene related tumor immunotherapy has become the focus of many cancer treatments.It can effectively treat disease from the source and eliminate or prevent hereditary diseases.Recent studies have shown that it has great potential in the treatment of melanoma.DNA vaccines are involved in the use of immune responses induced by DNA encoded by antigen.DNA vaccines have been widely used in the field of cancer and infectious diseases.The DNA encoded by antigen needs to be identified by APCs，such as DCs，macrophages and B lymphocytes，and then transfect APCs with its protease complex.The antigen protein is cut into small peptide fragments，which are presented to T lymphocytes and combined with MHC-Ⅰ and MHC-Ⅱ.Moreover，the surface related mannose receptors（MRs）of DCs can recognize a variety of sugar molecules on the cell surface or the cell wall of pathogens，which maintain the stability of the internal environment by participating in receptor-mediated phagocytosis，and combine innate immunity with acquired immunity to form the immune defense system of the body.In addition，studies on tumor signaling pathways indicate that molecular intervention of these signaling pathways provides a new idea for anti-tumor therapy.PUMA（the regulatory factors of apoptosis upregulated by p53），a member of the Bcl-2 protein family，is a downstream gene of p53 and has a pro-apoptotic effect.PUMA can induce the apoptosis of various tumor cells and inhibit the proliferation of tumor cells.
The recent strategies of preventive vaccines or therapeutic vaccines focus on improving the delivery of antigens and the extension of vaccine activity by the specific DCs.With the rapid development of the tumor immune response and tumor immunotherapy，immunoadjuvant as an immune enhancer has been widely used in the tumor vaccine.Immunoadjuvant can nonspecifically enhance the body's specific immunity by up-regulating the surface molecules expression of the immune cells and promoting the innate immune response.Effective tumor vaccines usually require appropriate adjuvants to enhance antigen presentation efficiency，antigen immunogenicity and antigen-induced immune effects.The codelivery of antigen and immunoadjuvant is expected to enhance the immune responses of the tumor-specific T lymphocytes，which can reduce the risk of immune escape and reduce the immune tolerance for single antigen peptides.Montanide ISA 51 and 720 are clinically approved as immunoadjuvants，which enhance the immunogenicity of tumor cells by stimulating humoral and cellular anti-tumor immune response.In addition，monophosphate A（MPLA），activator of Toll-like receptors 4（TLR4），is commonly used to enhance the immune response against viral infections and tumors.The stimulation of TLR could activate the internal signaling pathway in APCs and ultimately induce inflammatory cytokines，chemokines，interferons and synergistic costimulatory molecules，which provide the appropriate environment for the T lymphocytes stimulation.But the poor water solubility of MPLA resulted of the long hydrophobic alkyl chain has limited its extensive application in tumor vaccines.Therefore，it is of great significance to develop higher effect carriers for the co-delivery of tumor antigen and immunoadjuvant.
Compared with traditional therapies，immunotherapy has shown considerable improvement in objective response rates，time of disease progression and overall survival rate of patients.However，different immunotherapies also have their limitations.For example，immune checkpoint blocking has had unprecedented success，but unfortunately，only a small percentage of patients respond to immunotherapy.Clinical results indicate that only 15％of the patients respond to ipilimumab（a monoclonal antibody against human CTLA-4）and no more than 40％of the patients respond to anti-PD-1/PD-L1 antibodies.Tumor-associated antigen vaccine faces with instability，short half-life and easy to degrade in plasma，low uptake rate of gene fragments，easy to be degraded by nucleases，etc.
Furthermore，owing to the uncontrolled tissue accumulation and constitutive bioactivity of immunotherapeutic agents，most immunotherapy strategies share the risk of immunerelated adverse events（IRAEs），including hypokalemia，hypophysitis，myocarditis，diabetes mellitus and thyroid dysfunction.As a result，many patients have to discontinue treatment when IRAEs occur，which further limits the outcome of immunotherapy.Therefore，it is very necessary to improve the response rate of immunotherapy while reducing the side effects.
It is hopeful to remove various obstacles in the melanoma immunotherapy with the development of biomedical polymer materials.For example，biomedical polymer materials can prevent proteases from degrading antigenic peptides，and can control release antigen and prolong antigen presentation in the co-delivery of antigen and immunoadjuvant for tumor vaccine.As a gene carrier，the delivery system loaded gene fragments has the advantages of high plasticity，accurate structure，resistance to nuclease degradation，stability and low toxicity.In addition to the adjuvant immunotherapy of melanoma by encapsulating tumor antigens and genes，the adjuvant use of biomedical polymer materials for other melanoma components can also enhance the body's specific immune response and thus play an effective anti-tumor effect.Combining melanoma immunotherapy with biomedical polymer materials can optimize the existing methods of using melanoma antigens，gene fragments and other melanoma components for immunotherapy，promising to solve the existing problems of melanoma immunotherapy.