Study On Screening And Theraputic Efficacy Evaluation Of Nasopharyngeal Carcinoma Using Raman Spectroscopy Technology

Nasopharyngeal carcinoma（NPC）is a malignant tumor with obvious ethnic and geographical distribution characteristics,with about 130,000 new cases worldwide every year,of which 85%of NPC patients occur in China and Southeast Asian countries.Due to the non-specificity of the symptoms,the depth of concealment of the site and the difficulty of visualization of the disease,70%of patients have reached the middle and advanced stages at the time of treatment（the five-year survival rate is only about 30.3%）.With the early detection and timely treatment,the total five-year survival rate can be increased to beyond 90%.Moreover,because of its hidden lesions and structural specialties,it is difficult to perform surgical treatment.Radiotherapy,as the main treatment method,plays an increasingly prominent role in the treatment of NPC.Therefore,the sensitivity of patient radiotherapy has become the key to the evaluation of radiotherapy efficacy and prognosis.In this project,Raman spectroscopy technology with molecular fingerprint detection capability was used to detect blood,cells and nucleic acids samples in NPC and radiotherapy benefit population,with the aim of developing a novel method for NPC screening and therapeutic effect evaluation.The main research contents include:1.Surface enhanced Raman scattering（SERS）technique was used to DNA detection at the single base level and applied to NPC screening.In this study,high quality DNA-SERS signals were successfully collected through the self-adsorption interaction between DNA molecules and nanoparticle surfaces and the use of efficient aggregators.More importantly,the SERS signal of the phosphate backbone was used as an internal standard,enabling quantitative DNA detection at the single-base level.On this basis,the technique was applied to the detection and analysis of total circulating DNA in NPC patients,and a combination of principal component analysis and linear discriminant analysis（PCA-LDA）was used to discriminate NPC patients from normal subjects,diagnostic sensitivity of83.3%and a specificity of 82.5%.The study showed that this method has the potential to be an efficient way to screen for NPC.2.A SERS sensor with self-calibration and dual signal amplification was developed for quantitative detection of NPC-related DNA tumor markers.In this study,carbonyl compounds(^Re-SCO and ^Os-SCO),whose spectral signals are in the"silent region(1800-2200cm^-1)",were used as SERS probes and internal standards,respectively,without interference from other biochemical substances.In addition,in this study,SERS probes(^Re-SCO)were equipped in a mesoporous silica-coated Au nanorods,which is released into the detection system in large quantities by photo-triggered,thus achieving secondary amplification of the SERS signal.This technology can realize the reliable and ultra sensitive detection of NPC related DNA（EB-DNA）,which is of great significance for the development of NPC screening clinical diagnosis tool based on SERS technology.3.The plasma SERS technique was combined with multivariate statistical algorithm（PCA-LDA）for radiotherapy efficacy assessment of NPC patient.In this study,plasma samples from 40 pre-and post-radiotherapy NPC patients and 30 healthy volunteers were tested for SERS.Due to the changes of biomolecules in plasma after radiotherapy,the spectral characteristics of plasma SERS post-radiotherapy and pre-radiotherapy are significantly different.Using PCA-LDA analysis,we can distinguish the samples pre-and post-radiotherapy,pre-radiotherapy and normal.The discrimination sensitivity is 83.3%and 95.1%,and the specificity is 91.2%and 93.0%,respectively.The physical condition of pre-radiotherapy patients is similar to that of normal people,and the difference of plasma SERS spectra is very small.The sensitivity and specificity of discriminant diagnosis are only 61.8%and 67.4%.The results show that plasma SERS technology,as a fast and simple tool,has great potential in the field of NPC radiotherapy evaluation and cancer screening.4.The NPC cells with radiotherapy resistance and radiotherapy sensitivity were analyzed by laser tweezer Raman spectroscopy.In this study,radiotherapy-resistant cells were successfully constructed by continuous fractionated irradiation of NPC cells（CNE2）withγ-rays.Then,the NPC cells were captured using an in-house built laser tweezer Raman spectroscopy system to obtain the Raman spectral of the cells under physiological environment in real time.Finally,through the analysis of PCA-LDA,and the diagnostic sensitivity and specificity were 97.0%and 89.0%,respectively.These results indicate that the system can more accurately reflect the disease information at the cellular level,and can provide methodological reference for the study of cell-based radiotherapy resistance detection.5.The SERS technology was used to profile the exosomes delivered by NPC cells with radiotherapy resistance.In this study,SERS substrates with hydrophobic characteristics and self-calibration function were prepared to obtain high-quality SERS spectra of exosomes and explore the differences of trace biological components between the two kinds of exosomes.And a diagnostic model with deep learning algorithm was established for discriminant analysis of secrete SERS spectra（the accuracy is 92.4%）,which is expected to realize the clinical application of SERS technology in the evaluation of radiotherapy efficacy of NPC.Based on the urgent need for screening and precise treatment of NPC,a novel technology based on SERS was developed for the study on the NPC screening and therapeutic effect assasment.This study not only has important theoretical and scientific significance,but also is expected to be extended for early screening,efficacy detection and prognosis assessment of other cancers in humans,having the potential of clinical application.