**11. Conclusions and Future Prospects**

CYP4 enzymes are responsible for the metabolism of fatty acids and play important roles in the homeostasis of fatty acids and fatty-acid-derived biomolecules such as leukotriene, prostanoid, and 20-HETE. Thus, CYP4 enzymes make important contributions to human health, including cardiovascular health, skin barrier maintenance, eye function, and cancer protection. However, the lack of research into certain aspects of the CYP4 family must be overcome. First, a specific antibody for the detection of each CYP4 protein and a specific substrate for each enzyme function must be developed to clearly determine the expression levels of these enzymes in different tissues under various induction, inhibition, and genetic conditions. High similarity of protein structures, overlapping substrates, co-expression in the same tissues, and genetic differences among individuals have interfered with the identification and characterization of *CYP4* genes. For targeted therapy and targeted delivery of drugs into cells or specific tissues, accurate measurement of CYP4 activity in tissues is essential. Second, further functional studies of *CYP4* genetic variants are needed. A growing number of genetic mutations of *CYP4* genes have been identified using high-throughput sequencing techniques. However, most of their functional changes compared to the wild type remain unknown. Only a small number of high-frequency genetic variants with known functional information have been investigated in multiple

populations, likely due to their high statistical power, which enables publication. Although in silico tools are useful for the prediction of functional changes, in silico prediction does not yet perfectly reflect in vivo conditions. Therefore, various commercial software programs often provide inconsistent predictions for the same genetic mutations. Development of high-throughput techniques for in vitro functional study and improvement of in silico methods are needed to elucidate the functional changes caused by mutations. Third, globally standardized values for CYP4 activity must be developed for application in artificial intelligence technology and algorithms used for the prediction of CYP4-related human diseases or the progression of disease states. As shown in Figure 2, large variations in CYP4-mediated metabolism, genetic variants of *CYP4* and other genes, and differing environmental conditions have been observed among individuals. Data integration to support correct diagnosis in humans is currently not possible, but is the ultimate goal of such research. To achieve this goal, accurate molecular tools for characterization of each CYP4 enzyme, functional information about *CYP4* genetic variants, and a standardized system for the application of CYP4 functional values in artificial intelligence or machine-learning tools are needed for personalized health care.

**Figure 2.** Correlation of *CYP4* genes with phenotypic outcomes. Most *CYP4* genes share similar structures and overlapping metabolic substrates. Phenotypic outcome prediction is difficult with a single or few *CYP4* genetic studies. Phenotypic outcomes are affected by genetic polymorphisms of various genes and dynamic environmental factors. Fundamental research into *CYP4* genes is essential to provide the data integration necessary for more accurate phenotype prediction than can be obtained using conventional methods.

**Funding:** This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Korea government (MSIT) (No.2018R1A5A2021242) and by the National Research Foundation of Korea grant funded by the Korea government (NRF-2017R1D1A3B03031007).

**Conflicts of Interest:** The authors declare no conflict of interest.
