The expression of PHYH have been linked to multiple diseases such as Refsum Disease and Retinitis Pigmentosa . Although there are no study associating human cancers to PHYH expression, The Human Protein Atlas have reported it to be a prognostic marker in renal cancer . To our knowledge, expression of PHYH and its impact on ccRCC has not yet been explored. Therefore, the potential role of PHYH in ccRCC was the main focus point of our study.
We applied bioinformatics analysis using high-throughput RNA-sequencing data from TCGA to examine PHYH expression in ccRCC patients and its association with various advanced pathologic characteristics. We demonstrated that a decrease PHYH expression is associated with presence of tumour, grade of cancer, stage of cancer, primary size of tumour, age and presence of distant metastasis. To further investigate the functions of PHYH in ccRCC, we performed GSEA and gene-gent network using TCGA data. GSEA showed that butanoate metabolism, histidine metabolism, propanoate metabolism, pyruvate metabolism, tryptophan metabolism, PPAR signalling pathway and Renin angiotensin system are differentially enriched in PHYH low expression phenotype. Gene-gen network analysis revealed association of PHYH with multiple PEX genes. These evidences highlighted the potential of PHYH serving as a prognostic marker of prognosis and therapeutic target in ccRCC.
Results from our study showed a decreased expression of PHYH gene in patients diagnosed with ccRCC. The PHYH gene encodes the enzyme phytanoyl-CoA hydroxylase, which is required for the alpha oxidation of branched chain [22, 23] and long chain  fatty acids such as phytanic acid in peroxisomes . Researchers suspect that phytanoyl-CoA hydroxylase potentially participate in determining the number of peroxisomes within cells and is involved in regulating their activities . Peroxisomes are membrane bound organelle within the cytoplasm that is conserved across eukaryotic cells , and plays a vital role in peroxisomal fatty acid beta-oxidation metabolism and ROS (reactive oxygen species) conversion . Diseases such as the Zellweger syndrome and other genetic diseases occurs due to implications in the fatty acid beta-oxidation[29, 30]. Study have found that many chemicals designated as peroxisome proliferators can induce peroxisome proliferation, resulting in increase in fatty acid oxidation in liver cells which leads to tumours growth in rodents [31–33]. A study has observed an absence of peroxisome in epithelial cells of proximal tubule in cancer cells of renal cell carcinoma . AS phytanoyl-CoA hydroxylase is coded by the PHYH gene and key component in peroxisome regulation, results of the present study agree with the provided evidence and suggests that decreased expression of PHYH gene is associated with absence of peroxisomes in ccRCC patients.
The gene-gene interactions form the results of our study have shown associations of multiple PEX genes (PEX2, PEX7, PEX10, PEX13, PEX 14) with PHYH. PEX genes encode peroxins, a class machinery protein required for proper peroxisome assembly . Autosomal recessive loss of function mutations in the PEX genes can result in peroxisome biogenesis disorders in the brain bone kidney and liver [36–39]. Overexpression of PEX genes such as PEX2 can result in accumulation of ubiquitinated PEX5 which can promote pexophagy (autophagosomal degradation of peroxisomes) . Decreased PEX5 levels are associated with both the onset of cancer in vivo , and sensitivity to exogenous H2O2 addition in hepatocarcinoma model systems in vitro . Identification of PEX14-containg vesicles has connected peroxisomes biogenesis to mitochondrial mediation . PEX7 facilitate matrix protein import, which significantly contributes to peroxisome membrane growth . Notably, PEX7 has primarily been documented to directly shuttle PHYH to the peroxisomal matrix . Given the importance of peroxisomal matrix protein import in normal cells it could be anticipated that the expression and/or function of peroxisome matrix proteins might become aberrant in tumour cells . Combining the results from our analysis with the evidence presented, a clear association can be observed in which PHYH expression affects the expression of PEX genes. This in turn causes perturbations in peroxisomes biogenesis, function, and structure.
Results from the network analysis also revealed that alteration of PHYH expression in ccRCC phenotype implicates the alpha oxidation pathway. Genes HACL1, SLC27A2 (shown to be associated with PHYH) are genes that code for protein 2-hydroxyacyl-CoA lyase 1 and very long-chain acyl-CoA synthetase, both enzymes along with PHYH are critical enzymes in converting phytanic acid to pristanic acid. Recent review have highlighted that peroxisomal disorders affect phytanic acid and alpha oxidation . As most metabolism of phyantic acid occurs in the liver and kidney via alpha-oxidation, an alteration in PHYH expression will mostly likely implicate peroxisomal and subsequent alpha oxidation. The highlights the alpha oxidation as a target pathway for furfure studies in ccRCC.
GESA pathway analysis of TCGA data reveal multiple differentially expressed pathways in PHYH low expression phenotype. Among these altered pathways, the key peroxisome proliferator-activated receptor gamma (PPARγ) pathway has been shown to be functionally expressed  in ccRCC and that increased PPARγ abundance correlates with reduced patient survival . Gluconeogenesis associated pathways pyruvate, and butanoate metabolism have also been shown to be downregulated in kidney cancer . The renin angiotensin system (RAS) was also demonstrated to be under expressed in ccRCC. RAS is a hormone system known to maintain blood pressure and body fluids . Recent literature has implicated a crucial role of the RAS in the development and maintenance of cancer, particularly its effects on cancer stem cells [49–52]. In addition, RAS deregulation was demonstrated as a renal cancer risk factor . Collectively, evidences suggest that these altered pathways and metabolism are good association factors with ccRCC and starting points for understating in depth underlying pathophysiological mechanism of ccRCC phenotype.
In conclusion, PHYH expression may be a potential prognostic molecular marker of poor survival in ccRCC. Low PHYH expression in ccRCC patients is closely related with dysfunction, degradation, and absence of peroxides. This occurs alters alpha -oxidation pathway which may potentially be a targeted pathway for future studies, Moreover, PPAR signalling, pyruvate metabolism, butanoate metabolism and RAS may be the key pathway regulated by PHYH in ccRCC. Further experimental validation should be performed to prove the biologic impact of PHYH.