Developing cancer vaccine: Neoantigen identification and beyond
Immunotherapy has been the hot spot in cancer treatment in recent years. As a key component, neoantigen identification is usually done bioinformatically. If you are not familiar with concept of immunotherapy or neoantigen, please read THIS article first. Here Ghost wanna talk about general workflow of neoantigen prediction and its usage in developing cancer vaccine.
Just like any antigens in general, neoantigens are the containing tumor mutations peptides that are only expressed in tumor cells. Some of them can be recognized by major histocompatibility complex (MHC) and presented onto the surface of tumor cell. These neoantigens are immunogenic and can be recognized by immune cell. Cancer vaccine in general is a way to manufacture these immunogenic peptides and inject back to the patients hoping to boost their immune response.
Neoantigens are highly personalized
The tricky part of developing such vaccine is that the neoantigens differ between cancer patients. In addition, neoantigen's efficacy in triggering immune response depends on multiple other personalized biological property of the patient, such as HLA type and TCR type. Here is what has to be met to develop effective vaccine:
Somatic mutation: Neoantigen must somatic mutation.
Mutation expression: Neoantigen must be expressed in tumor cell.
MHC - peptide binding: MHC must be able to recognize and bind the neoantigen with minimal restriction. This is the most restrictive step (only a few peptideS can bind to MHC).
Neoantigen presentation: The neoantigen must be processed and presented onto the tumor cell surface by APC.
Immuno recognition: This is another heavily restrictive step. For TCR - neoantigen binding, the TCR must be able to recognize both neoantigen and MHC. First, the neoantigen must have the right anchor residue for TCR to bind. Besides, not every TCR has the able to every MHC. MHC restriction dictates that TCR can only recognize the MHC they have encountered in the thymus during lymphocyte development. This filtering step is usually done in vivo.
neoantigen candidate prediction
Now let's see how High Throughput Sequencing (HTS) technique is optimized for such task (Figure B).
To find potential neoantigen, we identify somatic mutations from tumor WES data.
To ensure the expression of neoantigen, gene expression is estimated on RNA-Seq data.
To ensure the neoantigen can be bind and processed by MHC, HLA (the genes codes MHC) typing is performed to get patient's HLA haplotype. For somatic mutations identified earlier, the 10mers and 15mers peptides with somatic mutation in them are generated and fetched together with HLA haplotype to MHC - neoantigen binding prediction tool, such as pVAC-seq.
At this point, we should be able to obtain a list of neoantigens candidates. The neoantigens in this list must meet multiple criteria such as variant allele depth (frequency), MHC binding affinity, locus (gene) expression, self similarity, immunogenicity. We once analyzed leukemia samples. Initially over 200 somatic mutations were identified with WES data as figure shown below (leukemia tends to have low mutation burden). Mutations were called using both VarScan and MuTect. MuTect usually gets less mutations due to the algorithm difference of these tools.
However, after predicting their bind affinity to HLA haplotype, only over 30 neoantigens made their way to the final candicate list.
In vivo neoantigen validation
We assume these neoantigen candidates have potential to be presented on the cell surface. One question left is whether patient's immune system can recognize them. This part is usually assessed by wet-lab. One way to validate these neoantigen is using MHC-humanized mice (To ensure the mice has the same HLA type as patient). these neoantigens can be further prioritized by MHC binding score and then synthesized and injected into mices. The vaccine efficacy can then be assessed by comparing mices with and without vaccination.
Problem with cancer vaccine
Current ongoing cancer vaccine research seems to limit to vaccine of single neoantigen. Tumor cells without such neoantigen will most likely resist to the boosted immune response in the way that very much like standard targeted therapy. In the future, vaccine mixed with multiple potential neoantigens may pose a solution to overcome the tumor heterogeneity issue.