# Design of Outer Membrane Vesicles as Cancer Vaccines: A New Toolkit for Cancer Therapy
## 4 Conclusions and Future Directions  
Currently, OMVs have immense potential to be used as cancer vaccines, but many studies focus only on the insertion of cancer antigens into OMVs and directly use OMVs derived from conventional engineering of _E. coli_. Genetic engineering of OMVs presents enormous potential to artificially modify different intracellular bacteria such as _Salmonella_ and _Listeria_ to maximize the ability of OMVs to stimulate immune responses, thereby designing an ideal cancer vaccine. In addition, tumor-targeted Gram-positive bacteria such as _Clostridium_ and _Bifidobacterium_ also spontaneously produced EVs through the exfoliation of cell membranes [110; 111]. Furthermore, bacterial EVs derived from Gram-positive bacteria _L. acidophilus_ and _S. aureus_ showed significant antitumor effects [112]. Thus, Gram-positive bacteria can also be used as targets for the design of vesicle-based cancer vaccines (Table 2).  
The challenges that may have to be addressed in OMV cancer vaccine development are as follows: (1) selection of suitable antigens for different types of cancer, (2) enhancing tissue phagocytosis through genetic engineering, and (3) refining the mechanisms of vaccine-induced immune responses and their clinical efficacy [28]. Similarly, the following issues remain to be addressed for OMVs: (1) the high reactogenicity of PAMPs, such as LPS, (2) low expression levels of relevant protective antigens, (3) immuno-dominant antigens that misdirect the immune response, and (4) molecules that are immunosuppressive or otherwise interfering with a protective immune response. Furthermore, among all challenges, immune responses directed to the carrier itself may affect the repeated use of a vaccine based on the same antigen delivery platform. It was reported that pre-existing anti-carrier antibodies may mediate antibody-dependent phagocytosis and promote antigen clearance, thus reducing antigen uptake by APCs and weakening the subsequent immune responses. In addition, undesired responses to OMV components may excessively consume the immune response and thus attenuate specific responses to the target antigen [101]. Therefore, finding a balance between the immune response induced by the OMV itself and the efficiency of antigen delivery is the most important task in designing OMVs as cancer vaccines.  
In conclusion, there are still many opportunities and challenges in the development of OMVs, which require more effort and clinical trials. We believe that cancer vaccines based on OMVs can become a safe and effective therapeutic option with prospects for broad application prospects.  
[PERSON], [PERSON], and [PERSON] wrote the manuscript. [PERSON] and [PERSON] revised the manuscript for its integrity and accuracy. [PERSON] approved the final version of this manuscript and takes responsibility for its content.  
This study was supported by the National Natural Science Foundation of China (31760261 and 31660035), the Science and Technology Research project of Jiangxi Provincial Education Department (60224), National Innovation and Entrepreneurship Program for College Students (201910403031), and the Key projects of Jiangxi Natural Science Foundation (20171 ACB20003).  
The authors declare no conflict of interest.