PMC8511698_01
Radiologic images before and after nivolumab showing distinct TGR differences. (A) Representative computed tomography scans of the patient's lung before, at baseline, and during treatment. (B) Second immunohistochemistry (IHC) showing CDX2 positivity, indicating that the possible original tumor site may be the gastrointestinal duct. (C) Second IHC indicating a PD-L1 expression rate of 5-10%. (D) Second IHC showing loss of MMR protein expression; *loss of MLH1 protein expression; **loss of expression of PMS2 protein. (E) Positron emission tomography/computed tomography (PET-CT) scan suggesting extensive tumor burden.
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Two months later, follow-up CT revealed tumor progression in both the lungs ( Figure 1A ). Positron-emission tomography-CT revealed increased intake of 18F-fluorodeoxyglucose in the intestinal mesentery, cervicothoracic, abdominal lymph nodes, and both the lungs, indicating significant tumor load once again ( Figure 1D ). Immunohistochemistry (IHC) of the second biopsy indicated that the tumor most likely originated from the gastrointestinal duct (IHC showed CDX2 positivity as shown in  Figure 1B , microscopic magnification x100). A 5-10% loss of MLH1 and PMS2 protein expression and PD-L1 expression was also observed ( Figures 1C, D , microscopic magnification x100), suggesting a dMMR status in this patient. A follow-up chest CT scan confirmed rapid metastatic progression in the lungs and lymph nodes ( Figure 1A ). This value was calculated with reference to a single lesion, but the CT scan revealed a large number of newly developed lesions ( Figure 1A ). The tumor growth curve as shown in  Figure 1E  also indicated explosive tumor progression. The time to failure of the patient was 1.5 months. The ratio of TGKexp and TGKref was 2.18.
Results showing tumor microenvironment and genomic mutation copies alteration in autopsy specimens. (A) Representative images of PanCK (pan cytokeratin), CD56, CD8, CD68, and HLA-DR as shown by mIHC. Nuclei (blue) and counterstained by DAPI. Original magnification x200. (B) Tumor proportion score (TPS) and combined positive score (CPS). (C) Quantification results of TIL percentages by mIHC in the tumor stroma and margin. (D) Alteration of the genomic mutation copies before and after immunotherapy.
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TGFBR2 p.K128Sfs (66.46%), and TSC1 p.T635Rfs (5.41%). Tumor NGS was detected via the punctured tissues. Three mutation genes were inspected ( Figure 2D ), including MLH1 p.N287Kfs (34.3%), TP53 p.H179Y (38.4%), and TSC1 p.T635Rfs (1.7%). Tumor mutation burden (TMB) was also calculated. Quantitative analysis of the tumor microenvironment showed that the degree of CD8+, CD68+, and CD56+ cells in the tumor stroma, and tumor region, were 0.59% and 1.28%, respectively, along with 0.15%, 0.30%, and 0.63% and 0.49%, respectively ( Figure 2C ).