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Objectives: Colorectal cancer (CRC) is a major global health concern, ranking as the third most common cancer and the fourth leading cause of cancer-related deaths worldwide. Currently, the diagnostic accuracy of Lymph node metastasis (LNM) is currently unsatisfactory. Therefore, there is an urgent need to develop a reliable tool that can accurately predict lymph node metastasis (LNM) in patients diagnosed with CRC. Methods: We conducted an extensive proteomics investigation aimed at examining lymph node metastasis (LNM) in individuals diagnosed with colorectal cancer (CRC). In the discovery stage, employing a mass spectrometry-based proteomic approach, we analyzed a cohort of 60 colorectal cancer patients (NM=30, LNM=30), identifying distinct molecular profiles that differentiate patients with and without lymph node metastasis (LNM). Subsequently, we validated the protein classifier associated with lymph node metastasis. Results: We elucidated a combinatorial predictive protein biomarker that can distinguish patients with and without lymph node metastasis by LC-MS/MS. The classifier achieved an area under the curve (AUC) of 0.892 (95% CI, 0.842-0.941), while in the testing cohort, it attained an AUC of 0.929 (95% CI, 0.824-1.000). Furthermore, the four protein markers demonstrated an AUC of 0.84 (95% CI, 0.783–0.890) in the validation cohort. Additionally, we categorized patients into three types based on immunophenotyping. Type 1 primarily consisted of patients with negative lymph node metastasis (NM), characterized by immune cells such as NK cells, CD4 T effector memory cells, and memory B cells. Type 2 mainly included patients with positive lymph node metastasis (LNM), characterized by immune cells such as mesangial cells, epithelial cells, and mononuclear cells. In Type 1, a prominent upregulation observed in immune inflammation, as well as in glucose and lipid metabolism. In Type 2, significant upregulation was evident in pathways such as pyrimidine metabolism and cell cycle regulation. The findings of this study suggest that immune mechanisms may exert a pivotal role in the process of lymph node metastasis in CRC. Conclusions: Here, we present plasma protein signatures associated with lymph node metastasis in colorectal cancer (CRC). However, further validation across multiple centers is necessary to generalize these findings. [ABSTRACT FROM AUTHOR]

Objective: Forkhead box O1 (FoxO1) plays a crucial role in the development of many tumors. Cyclin D kinase (CDK) 1 could influence the nuclear export and activity of FoxO1 through phosphorylation of serine (S)249. However, the effects of S249 phosphorylation in the development of glioma remain unclear. The aim of the present study is to assess the function of FoxO1:S249V mutant, which was converted S249 phosphorylation site into valine (V) residues in the glioma development. Methods: FoxO1-knockdown U251 glioma cells (U251-KD cells) were established by infection of retrovirus particles with FoxO1 siRNA and FoxO1 restored cells (FoxO1:S249V) were obtained by re-introduction of FoxO1:S249V cDNA. We detected mRNA expression by real-time PCR, and cell cycle arrest and apoptosis by flow cytometric assay, and cell proliferation by BrdU assay and CCK-8 assay. The protective effects of FoxO1:S249V were detected by the xenograft tumor formation assay. Results: The FoxO1 mRNA expression was significantly decreased in the glioma specimens (n = 24). The U251-KD cells showed downregulation of p27 and Bim, while the phosphorylation of CDK1 was upregulated. FoxO1:S249V cells inhibited the phosphorylation of S249, and induced G2/M cell cycle arrest, following reduced cell growth and increased apoptosis. Moreover, FoxO1:S249V expression effectively inhibits the glioma growth. Conclusion: Our findings suggest that the forced FoxO1:S249V suppressed the cell growth through G2/M cell cycle arrests and increased apoptosis in glioma. [ABSTRACT FROM AUTHOR]

Objective: Forkhead box O1 (FoxO1) plays a crucial role in the development of many tumors. Cyclin D kinase (CDK) 1 could influence the nuclear export and activity of FoxO1 through phosphorylation of serine (S) 249 . However, the effects of S 249 phosphorylation in the development of glioma remain unclear. The aim of the present study is to assess the function of FoxO1:S 249 V mutant, which was converted S 249 phosphorylation site into valine (V) residues in the glioma development., Methods: FoxO1-knockdown U251 glioma cells (U251-KD cells) were established by infection of retrovirus particles with FoxO1 siRNA and FoxO1 restored cells (FoxO1:S 249 V) were obtained by re-introduction of FoxO1:S 249 V cDNA. We detected mRNA expression by real-time PCR, and cell cycle arrest and apoptosis by flow cytometric assay, and cell proliferation by BrdU assay and CCK-8 assay. The protective effects of FoxO1:S 249 V were detected by the xenograft tumor formation assay., Results: The FoxO1 mRNA expression was significantly decreased in the glioma specimens (n = 24). The U251-KD cells showed downregulation of p27 and Bim, while the phosphorylation of CDK1 was upregulated. FoxO1:S 249 V cells inhibited the phosphorylation of S 249 , and induced G2/M cell cycle arrest, following reduced cell growth and increased apoptosis. Moreover, FoxO1:S 249 V expression effectively inhibits the glioma growth., Conclusion: Our findings suggest that the forced FoxO1:S 249 V suppressed the cell growth through G2/M cell cycle arrests and increased apoptosis in glioma.