A Study Of Quantitative MRI Parameters For Prediction Of Immune Phenotype Of Oral Tongue Squamous Cell Carcinoma

[Research background]The incidence of oral tongue squamous cell carcinoma(OTSCC)has gradually increased in recent years.Immunotherapy has become a new field of treatment for cancer.The tumor immune microenvironment is an essential participant in promoting the proliferation,survival,invasion and migration of cancer.Studies have confirmed that tumor immune microenvironment is closely related to tumor immune escape.It is observed there are different tumor-immune phenotypes generally assumed :immune-flamed tumors,immune-excluded tumors and immune-desert tumors.The immune-flamed tumors will play a better role in immunotherapy.Study has found the use of magnetic resonance imaging can provide biologic tumor date of tumors,which can be predictable biomakers of patient response before treatment.Diffusion-weighted imaging technology(DWI)is a functional magnetic resonance imaging technology.Its principle is the diffuse movement of water molecules.The apparent diffusion coefficient(ADC)can reflect the dispersion of water molecules in the tissue and provide information on the pathophysiological changes in the disease,which may become a non-invasive biological predictor of tumor immune microenvironment.[Research objects]This project will study the feasibility of using MRI quantitative parameters to predict immune phenotype of tongue squamous cell carcinoma.To analyze the expression of PD-L1+ on tumor cells and TILs in different immune phenotypes.And the difference of ADC and Volume values in different subgroups.[Materials and Methods]1.Case date A total of 29 patients between August 2012 to April 2020 were enrolled in this study,including 16 males and 13 females,aged 26-85 years,with an average age of(56.55±11.07)years.All of the tumors were diagnosed as OTSCC by experienced pathologists.2.MRI technology MR imaging was performed using a 1.5-T magnetic resonance unit(Philips R2SH7A5 1.5T SE MRI scanner).All patients were received by preoperative magnetic resonance imaging(MRI),diffusion-weighted imaging(DWI)and enhanced MRI examination.On the ADC images with a b value of 1000s/mm~2,the ROI was manually drawn over the he largest level of primary tumor from which subtracted evidently necrotic and cystic areas referring to T2-weighted contrast enhancing areas.We used 3D Slicer software on a workstation to automatically calculate the primary tumor volume.3.Immunohistochemical analysis CD3+,CD8+ and PD-L1+ expression in tumor tissues obtained from patients with OTSCC were examined by Immunohistochemical analysis on serial histological sections(3-μm-thick).All slices were scanned with a digital pathological slice scanner(3DHISTECH),and a semi-quantitative scoring method was used on the Case Viewer software to evaluate the immunohistochemical staining.4.Statistical analysis Statistical analysis was performed by using SPSS.v.22.0 software.The Shapiro-Wilk(S-W)test was used for diagnosis normality of data distribution.Correlations between expression of PD-L1+,CD3+ and CD8+ were analyzed using spearman correlation.The Fisher’s exact test was used to examine the association between the expression of PD-L1+,CD3+,CD8+ and clinicopathological characteristics.The independent samples t-test and the Mann-Whitney U-test was used to compare between the two groups.A one-way analysis of variance(ANOVA)and the Kruskal-Wallis H test was used to compare more than two groups.P-values below 0.05 were considered statistically significant.[Results]1.MRI manifestations of OTSCC The 29 cases of OTSCC mainly showed patchy,massive,nodular or strip-shaped irregular soft tissue masses on MRI images.T1 W images were all low,slightly low or intermediate signal.Hypointense,slightly high or mixed hyper-hypointense signal were showed on T2 WI with fat suppression.The diffusion was limited on DWI images,which showed increased signal intensity.ADC maps showed low or slightly low signal.2.Quantitative parameter values of DWI The ADC values(X ± SD)of the 29 subjects were(1.161±0.185)x10-3mm~2/s.The mean Volume value was 2264.42 mm3,with a range of 1068.74 to6950.63 mm3.3.The expression of CD3+,CD8+ and PD-L1+CD3+ expression score was 6.8±2.9,CD8+ expression score was 4.0(2.6 ～8.4).The score of PD-L1+ on tumor cells was 2.0(0～7.0).The score of PD-L1 on TILs was 0.6(0～1.9).CD3+ expression score was positively correlated with CD8+expression score(γ=0.655,P<0.001).The PD-L1+ expression score of tumor cells was positively correlated with the PD-L1+ expression score of TILs(γ=0.619,P<0.001).The CD8+ expression score was positively correlated with the PD-L1+expression score of TILs(γ=0.426,P<0.05).4.The difference of PD-L1+ expression on TILs in different immune phenotype.There was no difference in the expression of PD-L1+ on TILs between low CD3+ and high CD3+ groups.The expression of PD-L1+ on TILs was statistical significance in different immune phenotype(P<0.05).Among them,the expression of PD-L1+ on TILs was significantly higher in immune-inflamed than immune-desert group.5.Differences in ADC and Volume values of clinicopathological characteristics There was no statistically significant difference in ADC values between the groups of different age,gender,depth of invasion,metastasis.There was significantly difference in Volume values between female group and male group(P=0.01).The significant difference was also observed in metastasis and non-metastasis group(P=0.003).There was no statistically significant difference in Volume values among groups of different ages and depth of invasion.6.ADC and Volume values in different grades of OTSCC The difference in ADC values among well,moderately and poorly differentiated groups is statistically significant(P=0.001).There was a statistically significant between the well and moderately differentiated group(P<0.05).It was also observed between well and poorly differentiated group,while the difference between the moderately and the poorly differentiated group was not statistically significant(P>0.05).There was no significant difference in the Volume values of the different grades in OTSCC.7.The difference of ADC and Volume values in different immune phenotype.The difference in ADC values between low CD3+ and high CD3+ groups were not statistically significant,and the difference between low CD8+ and high CD8+groups were not either.There were significant differences in the Volume values of low CD3+ and high CD3+ groups(P<0.05),and the difference of the Volume values in low CD8+ and high CD8+ groups were not statistically significant.[Conclusion]1.The expression of PD-L1+ on tumor-infiltrating lymphocytes is correlated with the CD8+ expression score in OTSCC.The expression of PD-L1+ is different in tumor microenvironments based on CD8+ expression,which will produce different responses in immunotherapy.2.Apparent diffusion coefficient(ADC)values have significant differences in OTSCC with different pathological differentiation,which can be used to evaluate the tumoe malignancy before treatment.The ADC value was lower in immune-flamed group but it was underpowered to show statistical significance,which was probably due to small sample size.The tumor volume value is higher in the cervical lymph node metastasis group.The tumoe volume can be used to predict the cervical lymph node metastasis of OTSCC.Tumor volume has a certain value in predicting clinical parameters,which play a certain role in the evaluation of tumor infiltrating lymphocytes(TILs)by providing complementary information.