Chronological evolution of immune tumor microenvironment (TME) from esophageal dysplasia (ED) to esophageal squamous cell carcinoma (ESCC)

Background : Previous studies have shown that ED, a precancerous stage of ESCC, exhibits genetic alterations and genomic instability similar to those observed in ESCC. Other factors-such as alterations in immune surveillance-may also contribute to the progression from dysplasia to invasive cancer. Methods: Patients with a history of ED prior to the diagnosis of ESCC were enrolled for gene expression profiling (GEP) study of esophageal lesions and patients with early stage ESCC treated with endoscopic submucosal dissection (ESD) were enrolled for immunohistochemistry (IHC) study. Formalin-fixed paraffin-embedded tissues (FFPEs) from ED diagnosed more than 6 months prior to the diagnosis of ESCC (denoted as dysplasia-1), ED diagnosed within 6 months prior to the diagnosis of ESCC (denoted as dysplasia-2), and ESCC were retrieved. Transcriptomic data were generated by NanoString nCounter platform with Human PanCancer Immune Profiling panel and were further analyzed for the expression levels of infiltrating immune cells by CIBERSORT. FFPEs from another cohort of patients with early stage ESCC treated with ESD were stained with anti-CD68 (Rabbit polyclonal, abcam, UK) and anti-CD163 (Rabbit polyclonal, Synaptic Systems GmbH, Göttingen, Germany). Their expression levels were semi-quantitatively determined in the stroma and intratumoral fields and were compared among the areas of normal mucosa, dysplasia, and ESCC in the same patient. Results: Seventeen ESCC patients were enrolled for GEP study. The median time periods from the diagnosis of dysplasia-1 to that of ESCC and from the diagnosis of dysplasia-2 to that of ESCC were 8.6 months and 3.0 months, respectively. The analysis of immune cell signatures defined by NanoString platform revealed that multiple cell types were significantly increased in ESCC compared with dysplasia-1 (mast cell, macrophage, and dendritic cell, all P < 0.05) and in ESCC compared with dysplasia-2 (macrophage, mast cell, CD8 T cell, dendritic cell, total TIL, T cell, and regulatory T cell, all P < 0.05). No significant difference was found between dysplasia-1 and dysplasia-2. Immune cells classified by CIBERSORT showed an increase of M2 macrophage and a decrease of M1/M2 ratio in ESCC compared with dysplasia-1 or dysplasia-2 (both P < 0.05). Eighty-two early ESCC patients were enrolled for IHC study. A progressive increase in the expression levels of CD68 and CD163 was found from mucosa to dysplasia then to ESCC (all P < 0.001) Conclusions: The increased expression of M2 macrophages in the ESCC TME compared with precancerous dysplasia supports the hypothesis that immune TME alterations may drive the progression from dysplasia to ESCC. (Funded by MOST 107-2314-B-002-199-, MOST 109-2314-B-002-231-, MOHW114-TDU-B-221-144006, NSTC 114-2314-B-002 -206 -MY3, NTUCCS-110-10, and NTUCCS-111-05)