Trophinin Is an Important Biomarker and Prognostic Factor in Osteosarcoma: Data Mining from Oncomine and the Cancer Genome Atlas Databases

Osteosarcoma (OS) is a type of bone malignancy with a high rate of treatment failure. To date, few evident biomarkers for the prognostic significance of OS have been established. Oncomine was used to integrate RNA and DNA-seq data from the Gene Expression Omnibus (GEO), The Cancer Genome Atlas (TCGA), and the published literature. The correlation of the gene Trophinin (TRO) and different types of cancers was generated using the Cancer Cell Line Encyclopedia (CCLE) online tool. Prognostic values of featured Melanoma Antigen Gene (MAGE) members were further assessed by establishing the overall survival using the Kaplan-Meier plotter. Moreover, the online tool, Database for Annotation, Visualization and Integrated Discovery version (DAVID), was used to understand the biological meaning list of the genes. MAGEB10, MAGED2, TRO, MAGEH1, MAGEB18, MAGEB6, MAGEB4, MAGEB1, MAGED4B, MAGED1, MAGEB2, and MAGEB3 were significantly overexpressed in sarcoma. TRO was further demonstrated to be distinctively upregulated in osteosarcoma cell lines and associated with shorter overall survival. TRO may play an important role in the development of OS and may be a promising potential biomarker and prognostic factor.


Introduction
Osteosarcoma (OS) is the most common bone malignancy in young people, with over 50% of cases occurring in the first two decades of life [1]. Tremendous progress has been achieved in the past few decades in the field of OS therapy development including advances in surgical and diagnostic strategies and the use of combined chemotherapy [2]. Despite this progress, curative treatment of OS remains challenging with subsequent poor prognosis and shortened life expectancy [3]. Early diagnosis and treatment of OS, particularly with nonmetastatic OS, is crucial for improving the prognosis of OS patients [4]. Unfortunately, few distinctive diagnostic and prognostic biomarkers have been identified. A better understanding of the molecular and cellular biology and an identification of effective biomarkers involved in tumor initiation and progression are crucial for optimizing diagnosis and treatment of OS.
The TRO gene encodes for Trophinin, a member of the Melanoma Antigen Gene (MAGE) family, mediate the initial attachment of the blastocyst to the uterine epithelial cells during embryo implantation through a cell adhesion molecule complex in combination with bystin and tastin [5]. Furthermore, a prior study indicated that TRO is overexpressed in many trophoblastic cancers, which is of great importance for the progression of trophinin-expressing cancers [6]. TRO may therefore have a critical role in the development of different cancers, with a possible role in OS initiation and prognosis remaining elusive.

CCLE
Analysis. CCLE analysis agreed with the Oncomine results demonstrating that TRO was distinctively upregulated in the osteosarcoma cell lines (Figure 4(a)). In addition, the DNA copy number was relatively high in osteosarcoma (Figure 4(b)). Moreover, the Achilles shRNA knockdown results showed osteosarcoma was ranked second when ordered by dependency probability with TRO (Figure 4(c)).  (13) Table 1).

Discussion
OS is the most common type of bone malignancy with a high incidence in teenagers [7]. Early diagnosis and effective treatment of OS could remarkably decrease the high OS-related mortality [8,9]. However, current therapies for advanced OS remain poor due to the lack of specific molecular targets. In this study, we first aimed to investigate the diagnostic role and prognostic value of TRO in OS.
TRO is a member of MAGE family members of which have previously been reported as effective targets for cancer immunotherapy [10]. TRO is a type of adhesion molecule with specific expression on human trophoblast cells [11]. Trophoblastic cancer patients with high expression of this molecule often have poor prognosis [12]. Among those with positive expression of TRO, HMGB1/RAGE are often coexpressed, suggesting that TRO can promote tumor invasion through an HMGB1/RAGE signaling pathway [13]. Similarly, high levels of TRO were found to be related with poor prognosis in liver and gallbladder cancer [14,15]. It has also been demonstrated that TRO enhanced invasion and promoted colorectal cancer through a mechanism involving HMGB1/RAGE [11]. However, the function of TRO in the prognosis and progression of OS had not yet been fully elucidated.
This study first investigated the function and clinical significance of TRO in OS patients via the clinical data of OS patients and public expression profiles. The results indicated that the members of the MAGE family, including MAGEB10, MAGED2, TRO, MAGEH1, MAGEB18, MAGEB6, MAGEB4, MAGEB1, MAGED4B, MAGED1, MAGEB2, and MAGEB3, were significantly overexpressed in sarcoma tissues. And TRO was further demonstrated to be distinctively upregulated in OS cell lines and associated with shorter overall survival. Moreover, it was indicated from the statistical analysis that there was an evident correlation between TRO expression levels and a relatively poor prognosis. It was verified by further univariate and multivariate analyses that TRO could be used as a potential prognostic biomarker for OS patients.
To the best of our knowledge, the regulation of the expression of TRO in OS patients has not yet been explored. A limited number of studies have reported that overexpression of other MAGE family numbers, such as melanoma antigen family A (MAGEA), and preferentially expressed antigen in melanoma (PRAME) are related to OS progression [16]. However, the specific mechanism of MAGE family contribution to the occurrence, development, and prognosis of OS is still unknown. The present study suggested that TRO can act as a promising biomarker for OS diagnosis and prognosis. Nevertheless, the current study presented results derived solely from bioinformatics analyses, and further experimental evidence is required to validate these findings.

Conclusions
Overall, as TRO is overexpressed in human OS tissues, a poor prognosis can be predicted by a high level of TRO. Therefore, our findings highlight that TRO may be an important biomarker for the prognosis and progression of OS.

Data Availability
The data used to support the findings of this study are available from the corresponding author upon request.