HNCs rank as the sixth most common type of cancers worldwide. The cancers are often at an advanced stage at the time of diagnosis and display frequent recurrence and metastasis. Thus, prognosis and patient survival are poor. Radiotherapy and chemotherapy have largely improved the treatment of HNCs in recent decades12, 33, 34, 41, 50, 56. However, the 5-year survival rate is still very low. Improving the accuracy of early diagnosis could significantly improve the disease-free survival rate of patients.
Compared with other detection methods, liquid biopsy has become the preferred choice for disease screening because of its non-invasiveness, low cost, ease of use, and high stability. Some biomarkers for HNCs, including proteins, miRNAs, and EBV DNA, have been identified using liquid biopsies13, 54. However, each of these markers has its own disadvantages, including low positive rates, high false positive rate, need for experienced operators, and instrumental limitations. Therefore, finding effective early diagnostic markers in serum is critical for the treatment of HNCs.
LncRNAs have been reported to participate in the pathogenesis of HNCs. LncRNAs circulating in the serum or other bodily fluids present promising biomarkers for clinical diagnostic and prognostic applications. For example, serum MALAT1, AFAP1-AS1, and AL359062 can function as diagnostic and prognostic biomarkers for NPC 22. Notably, the upregulation of the ATB lncRNA can accurately predict papillary thyroid carcinoma and its prognosis6. However, few studies have examined novel lncRNAs expression in serum in HNCs.
The LOC284454 lncRNA is located on 19p13.12 and the miR-23-a~27a~24-2 cluster is present upstream of the same transcript. LOC284454 is a nuclear localized and chromatin associated lncRNA. LOC284454 RNA is found only in primates and is highly conserved. In our previous study, we demonstrated that LOC284454 promotes migration and invasion of NPC cells invitro and in vivo, and is associated with skeletal remodeling and adhesion signal pathways11. In this study, based on the feasibility of SYBR-qPCR and TaqMan-qPCR tests of serum LOC284454, we found that compared with healthy controls, the expression of LOC284454 was higher in NPC, oral cancer, and thyroid cancer, indicating that LOC284454 might be very important for the diagnosis of HNCs. To confirm this, we used ROC curve analysis to evaluate the diagnostic value of LOC284454. The AUC values of LOC284454 in NPC, oral cancer, and thyroid cancer were 0.931, 0.698, and 0.834, respectively, indicating that LOC284454 might be an appropriate diagnostic biomarker for these cancers. Even though we found that LOC284454 is highly expressed in NPC, oral cancer, and thyroid cancer, that does not mean LOC284454 can be generalized to all cancers. Study have shown that LOC284454 is significantly reduced in prostate, uterus, breast, and kidney cancer7, suggesting that LOC284454 is specificly highly expressed in HNC.
Real-time PCR can sensitively detect small changes in nucleic acids based on fluorescent dyes and fluorescently labeled probes. In TaqMan-PCR, a fluorescent reporter group and a fluorescence quenching group are labeled on both ends of the probe1, 14, 16, 55. When amplified, the 5'-3' exonuclease activity of the Taq enzyme degrades the probe. The fluorescent reporter group and the fluorescence quenching group are separated, so that the fluorescence monitoring system can receive the fluorescent signal, and the accumulation of fluorescent signal is completely synchronized with the formation of the PCR product 20, 24, 37. Since the qPCR instrument has a multicolor fluorescent channel, the experimental group and the control group are allowed to react in the same tube with the same cDNA template, which can reduce systematic errors and improve the specificity and sensitivity of the experiment40. This is also one of the highlights of this study and might be very useful for future detection of biomarkers.
We found that LOC284454 is highly expressed in the peripheral blood of HNCs. Why it remains stable in the peripheral blood is still unclear. We suspect that this may be related to exosomes or vesicles. Exosomes can encapsulate proteins, lipids, and nucleic acids, remain stable in the tumor microenvironment, and are important in tumor metastasis46. Recent studies have shown that non-coding RNAs exist in exosomes. Exosomes can carry non-coding RNAs to non-adjacent cells for information communication and can participate in tumor development26, 27, 35, 44. More research is needed to elucidate these mechanisms.
In summary, our results verified that LOC284454 is significantly upregulated in the serum of patients with NPC, oral cancer, and thyroid cancer based on SYBR-qPCR and TaqMan-qPCR. Moreover, ROC curve data indicate that LOC284454 could be used as a novel diagnostic biomarker for HNCs. Further research should focus on follow-up investigations to study the prognostic value of LOC284454. It is hoped that the development of new technologies, such as digital PCR, will make it easier to detect phenotypic specific molecular changes, and will increase the sensitivity and specificity of biomarkers.