A Guide for Selection of Delivery Vectors for Cancer Vaccine (p

    1. Bacterial Vectors

    2.1 Listeria Monocytogenes

    Listeria monocytogenes (LM) is a Gram-positive bacterium, in the division Firmicutes, named after Joseph Lister. It is the species of pathogenic bacteria that causes the infection listeriosis. It is a facultative anaerobic bacterium, capable of surviving in the presence or absence of oxygen. It can grow and reproduce inside the host's cells and is one of the most virulent foodborne pathogens, with 20 to 30% of food borne listeriosis infections in high-risk individuals may be fatal. Its ability to grow at temperatures as low as 0℃ permits multiplication at typical refrigeration temperatures, greatly increasing its ability to evade control in human foodstuffs.

    Listeria monocytogenes is a Gram-positive facultative intracellular parasite with a wide range of hosts, with special intracellular infection and immune mechanisms. After LM infects the host, it can exist in both lysosomes and in the cytoplasm of the cells, so that the foreign antigens transported can enter the MHC class I and MHC class II antigen presentation pathways, and induce CD+8T cells and CD+4T cell immunity. Response; at the same time, LM's natural adjuvant characteristics make it an ideal vaccine carrier, especially in the delivery of tumor antigens.

    2.1.1 Immune response induced after LM infection Innate immune response

     LM infection of host cells can lead to the production of a variety of pro-inflammatory factors and inflammatory chemokines. The most important early cytokines are IL-12p70, γ-interferon (IFN-γ) and interleukin 18 (IL‐ 18). These cytokines and cascades produce chemokines that induce neutrophil infiltration into the site of infection to activate cell-mediated killing. Interleukin 12 (IL-12) stimulates NK cells and T cells to produce IFN-γ, which in turn activates macrophages and stimulates aggregation of dendritic cells (DCs) carrying bacterial antigens. To the site of infection, further induction of antigen-specific cytotoxic T lymphocyte effect (CTL) is induced. IFN-γ induced by LM infection in a host can trigger DC cells carrying bacterial antigens to regulate co-stimulatory molecules and produce functional IL-12, and IL-12 as an important signaling molecule can induce unsensitized helper T cells differentiate into helper T cells to advance cell-mediated immune responses. Cellular immune response

    After LM carries a foreign antigen into the host, the bacteria present in the lysosome are degraded into short peptides. The short peptide binds directly to the MHC class II molecule and is then presented to CD+4T cells. CD+4T cells produce a large number of Th1 type cytokines. For example, IFN-γ and tumor necrosis factor alpha (TNF-α), these cytokines not only promote the differentiation of Th0 cells into Th1 type cells, but also play a role in assisting CD+8T cell responses. When LM escapes to the cytoplasm of the cell, the secreted protein enters the proteasome structure and is degraded into a peptide substance. The peptide substance is transported to the endoplasmic reticulum and then binds to the MHC class I molecule and is presented to CD+8T cells. After LM infection, effector CD+8 cytotoxic T lymphocytes rapidly expand, mediating the recognition and killing of infected cells, and then transforming into protective T lymphocytes.

    2.1.2 Advantages of attenuated LM as a foreign antigen expression vector

    The researchers engineered the attenuated LM into a vector that can accommodate exogenous antigens, making it a prophylactic or therapeutic live vector vaccine that induces a strong cellular immune response. As a vaccine vector, attenuated Listeria has the following advantages: direct infection of antigen-presenting cells, with two types of antigen processing and presentation pathways; chromosomal system can stably express multiple gene products, easy to operate; as Gram-positive bacteria, no Contains endotoxin and is easy to use. In view of these advantages of LM as a foreign antigen expression vector, LM has become a research hotspot in molecular biology and immunology in recent years. Listeriosis vaccines has been widely used as a vector for carrying viral and tumor antigens to stimulate cell-mediated immune responses. With the attenuated LM as the carrier, many foreign antigens have been studied in experimental animal models such as mice, rabbits, cats, chickens and monkeys, and good results have been obtained.

    2.2. Salmonella

        Salmonella belongs to the Enterobactericae family, a group of Gram-negative, facultatively anaerobic and facultatively intracellular pathogenic bacteria. Currently, based on genome sequence similarity, the genus Salmonella is categorized into two species S. bongori and S. enterica which in turn is divided into six subspecies including S. enterica subsp. enterica.

    Salmonella sp. has an ability to multiply inside phagocytic and nonphagocytic cells including macrophages, dendritic cells (DCs), neutrophils, M cells and epithelial cells. The ability of Salmonella to invade and survive within a host cell is dependent on two Type III Secretion Systems (T3SS), the multiprotein complexes with a needle-like structure present on the bacteria cell wall. Proteins involved in the assembly of the two major T3SSs of Salmonella are encoded by SPI1 and SPI2.

    The vast majority of tumors express proteins or other antigens that are absent (or present only in very low quantities) in healthy adult tissues. These tumor-associated antigens (TAAs) are potentially immunogenic and tumor development is usually accompanied by specific, although often ineffective, anti-TAA immune response. TAA vaccines used for cancer therapy often fail, probably due to inadequate antigen presentation and insufficient activation of innate immunity. The application of Salmonella as a vector for TAAs should result in overcoming both impediments. The first attempts to deliver TAA via Salmonella were undertaken in late 1990s. From that time numerous studies utilizing natural (mPSCA, mAFP, survivin, endoglin) or artificial (β-galactosidase) tumor antigens have proved that placing a TAA-coding transgene under strong cytomegalovirus promoter in a plasmid carried by Salmonella allows for TAA expression in the cytoplasm of infected cells or dendritic cells which engulfed the infected, apoptotic cells; TAA expression elicits efficient cell-mediated or both cell-mediated and humoral immune responses.

    Attenuated Salmonella mediates the reaction with the host through the type III secretion system. It can transmit effector genes and express a variety of therapeutic proteins on the cell surface or cytoplasm, induce the body to produce corresponding specific humoral immunity, cellular immunity and local mucosal immune response, and resist the invasion of pathogens carrying the corresponding foreign proteins. Attenuated Salmonella carrying a foreign gene enters macrophages and dendritic cells and then disintegrates and dies, releasing multiple copies of the foreign gene, exogenous gene expression. Inducing MHC-1 molecule-mediated antigen-specific CD4+ and CD8+ T cell tumor protective immune responses. CTL and antigen-presenting dendritic cells were activated, with a decisive increase in the respective activation markers CD2, CD25, CD28, CD48 and CD80.

     Because attenuated Salmonella can not only grow under aerobic and anaerobic conditions. It can also selectively accumulate in tumor tissues. The hypoxic necrotic areas in the tumor can make these bacteria easily spread in tumor tissues: at the same time, they are sensitive to antibiotics. If necessary, it can be eliminated by antibiotics without tolerance; plus good invasiveness and specific tendency. Can be displaced from the distant inoculation section and accumulate in the tumor site. The level of replication in tumor tissues is 1000-10000 times higher than that of normal tissues; and the cost is low, which can be administered by oral route, with few side effects, suitable for long-term treatment and can express multiple effector genes, and combined with other methods to exert anti-tumor Maximum effect. Long-term maintenance until tumor regression and many other features as an anti-tumor ideal carrier delivery vector, which provides the possibility of using Salmonella as a targeting vector for tumor gene therapy. Attenuated Salmonella cancer vectors, currently experts have used attenuated Salmonella as a gene transfer vector to treat solid tumors, deep tumors and metastases, and have achieved satisfactory therapeutic effects in both in vitro and animal model experiments. It has been shown to have high tumor targeting and safety in clinical phase I experiments. And some of the results have been initially applied to the clinic. It shows a strong development prospect.


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