In this study, the loss of ASXL1 protein expression was detected more frequently in adenocarcinomas and metastatic lymph nodes than in normal mucosa and adenomas. Furthermore, patients with negative and moderate expression (IRS 0–6) of ASXL1 protein showed frequent lymph node metastasis. Based on these results, we can assume that the ASXL1 gene might act as a tumor suppressor gene in CRC. To the best of our knowledge, this is the first study on ASXL1 expression in clinical samples of CRC. In previous studies, ASXL1 has been speculated to exhibit a tumor-suppressive or oncogenic role in a context-dependent manner . Most mutations detected in ASXL1 were truncations that are observed in various types of human cancers [5, 6, 9-11]. In addition, ASXL1 truncation is common in advanced CRC with infiltrative growth and peritoneal seeding . However, previous research has also shown that ASXL1 is overexpressed in cervical cancer . Therefore, similar to the case in most other cancers, the ASXL1 gene appears to exhibit a tumor suppressive function in CRC, except cervical cancer.
An ASXL1 mutation detected by NGS in eight patients was found to have a frameshift deletion at codon 1934delG (Gly645Val), resulting in the production of truncated ASXL1 protein . The truncated ASXL1 protein might exhibit a dominant-negative effect on gene regulation because frameshift mutations in ASXL1 disrupt the plant-homology domain . In this study, only the abundant expression group (IRS 8–12) exhibited distinctive features, such as larger tumors, less lymph node metastasis, and less lymphatic invasion than the negative and moderate expression groups (IRS 0–6). A possible explanation for this similarity in terms of the clinicopathological characteristics between the negative ASXL1 protein expression group and the moderate expression groups could be the dominant-negative effect of truncated ASXL1 protein in the moderate expression group. The truncated malfunctioning protein may interfere with the function of the normal ASXL1 protein.
The loss of ASXL1 protein expression was detected more frequently in metastatic lymph nodes than in adenocarcinomas. Furthermore, lymph node metastasis and lymphatic invasion were more frequent in CRC patients with negative and moderate expression (IRS 0–6) than in CRC patients with abundant expression (IRS 8–12). Thus, the loss of ASXL1 protein expression in CRC might be associated with cancer that spreads via a lymphatic route. The mechanism of this association requires further elucidation.
As ASXL1 truncations in human myeloid malignancies are misregulating mutations, which alter the transcriptional landscape, ASXL1 mutations are associated with a poor prognosis in patients with myeloproliferative neoplasms, myelodysplastic syndromes, chronic myelomonocytic leukemia, and acute myeloid leukemia . In this study, 5-year DFS rate was significantly different between the two groups, but the 5-year DFS rates of stage II and III cancers did not differ significantly. Considering the small number of abundant expression (IRS 8–12) groups when divided into stage II and III, a large-scale study on the role of ASXL1 protein as a prognostic factor is warranted.
ASXL1 truncation frequently occur in CRC cell lines with MSI . In addition, MSI is associated with advanced age because of epigenetic abnormalities in MLH1 promotor methylation . In the current study, MSI status was not conclusive based on the expression of ASXL1 protein in CRC. The reason for this result might be because MSI analysis was not conducted during the entire course of the study period, and therefore additional testing is required. In addition, the levels of ASXL1 protein expression in CRC patients were not found to be correlated with age.
Limitations of this study include the lack of cytogenetic analysis and difficulty in determining a clear cut-off score for ASXL1 protein expression because of the dominant-negative effect of the mutant protein. In addition, the results of MSI status were not evaluated for the entire study period. Despite these limitations, we believe that this is the first study on ASXL1 expression in clinical CRC samples.