N1, N12-Diacetylspermine Is Elevated in Colorectal Cancer and Promotes Proliferation through the miR-559/CBS Axis in Cancer Cell Lines

Abstract

N1, N12-Diacetylspermine (DiAcSpm) has been reported to be upregulated in the urine of cancer patients. Mass spectrometry has shown elevated DiAcSpm expressions in colorectal cancer (CRC) tissues. However, the diagnostic application of DiAcSpm is not available due to a lack of diagnostic grade antibodies. Also, its biological roles in CRC cells remain unexplored. In the present study, we developed an antibody that directly detected DiAcSpm expression in paraffin-embedded tissues. We also characterized its biological characteristics and underlying mechanisms. Polyclonal antibodies were generated by immunizing animals with a synthetic product of DiAcSpm. Antibody DAS AB016 showed strong sensitivity against DiAcSpm in CRC tissues. Immunohistochemistry results showed that DiAcSpm expression was significantly elevated in CRC tissues. High levels of DiAcSpm correlated with the clinical stage and Ki67 index. DiAcSpm treatment increased levels of proliferation, cell cycle progression, and cyclin D1 and cyclin E proteins in CRC cell lines, SW480 and Caco-2. DiAcSpm also upregulated ATP production in these two cell lines. RNA-sequencing showed that DiAcSpm downregulated miR-559, which was confirmed using RT-qPCR. The luciferase reporter assay, western blotting, and RT-qPCR showed that cystathionine β-synthase (CBS) was the target of miR-559. miR-559 inhibited, while CBS accelerated, CRC proliferation. In addition, CBS siRNA knockdown blocked the biological effects of DiAcSpm on CRC cells. In conclusion, DiAcSpm was found to be increased in CRC tissues using a newly developed antibody. DiAcSpm accelerated CRC proliferation by regulating the miR-559/CBS axis.

Emerging roles of cystathionine β-synthase in various forms of cancer

“…Several independent studies have confirmed CBS protein expression in the above colon cancer lines, and extended the findings to additional colon cancer cell lines (e.g. DLD-1 and SW480) as well [ 27 , 38 , 49 , 51 , 52 , [90] , [91] , [92] , [93] , [94] , [95] , [96] , [97] , [98] , [99] , [100] ].…”

“…However, clearly, high CBS expression has not been observed in all rapidly proliferating cell types studied, so proliferation, by itself , can not be the sole driving factor. Nevertheless, in some cases, a close correlation has been found between CBS upregulation and increased cell proliferation; for instance, in human colon cancer cell lines treated with increasing concentrations of the polyamine N12-diacetylspermine [ 98 ]. The potential role of hypoxia as a CBS-inducing or enhancing factor, has been explored by multiple investigators, and yielded variable results.…”

The Potential Role of Spermine and Its Acetylated Derivative in Human Malignancies

“…One of the well explored spermine derivative in cancer is N 1 , N 12 -Diacetylspermine (DiAcSpm), which is produced by two rounds of acetylation of spermine, catalyzed by SAT1/2. Studies have demonstrated the prognostic value of DiAcSpm in lung cancer, colorectal cancer and breast cancer [14][15][16]. In this review, we are going to review the changes in spermine level in different human cancers and the possible applications of spermine in cancer treatment regimes in a clinical setup.…”

Recent advances in the role of endogenous hydrogen sulphide in cancer cells

“…89 N1, N12-Diacetylspermine can up-regulate the expression of CBS and promote the proliferation of colorectal cancer cell lines SW480 and Caco2, while miR-559 can target and inhibit CBS and inhibit the proliferation of the cells. 138 These opposite results of CBS may be caused by the difference in cell lines, and AOAA, as a pyridoxal phosphate-dependent enzymes inhibitor, showed an inhibitory effect on both CBS and CSE. 139,140 3-MST is highly expressed in murine colon cancer cell line CT26.…”

“…MiR-24-3p, miR-203, and miR-376a can target CBS to induce apoptosis, and miR-559 can also target CBS to inhibit cell proliferation. 130,138,173,174 MiR-30 family can induce oxidative stress by inhibiting CSE, 175,176 while miR-216a also directly targeted CSE to inhibit its expression. 177 And miR-4137 can inhibit both CBS and CSE in BC to yield a tumour-suppressing effect.…”