Functional expression of choline transporter-like protein 1 (CTL1) in small cell lung carcinoma cells: A target molecule for lung cancer therapy

Abstract

Choline is essential for the synthesis of the major membrane phospholipid phosphatidylcholine and the neurotransmitter acetylcholine (ACh). Elevated levels of choline and up-regulated choline kinase activity have been detected in cancer cells. Thus, the intracellular accumulation of choline through choline transporters is the rate-limiting step in phospholipid metabolism and a prerequisite for cancer cell proliferation. However, the uptake system for choline and the functional expression of choline transporters in lung cancer cells are poorly understood. We examined the molecular and functional characterization of choline uptake in the small cell lung carcinoma cell line NCI-H69. Choline uptake was saturable and mediated by a single transport system. Interestingly, removal of Na(+) from the uptake buffer strongly enhanced choline uptake. This increase in choline uptake under the Na(+)-free conditions was inhibited by dimethylamiloride (DMA), a Na(+)/H(+) exchanger (NHE) inhibitor. Various organic cations and the choline analog hemicholinium-3 (HC-3) inhibited the choline uptake and cell viability. A correlation analysis of the potencies of organic cations for the inhibition of choline uptake and cell viability showed a strong correlation (R=0.8077). RT-PCR revealed that choline transporter-like protein 1 (CTL1) mRNA and NHE1 are mainly expressed. HC-3 and CTL1 siRNA inhibited choline uptake and cell viability, and increased caspase-3/7 activity. The conversion of choline to ACh was confirmed, and this conversion was enhanced under Na(+)-free conditions, which in turn was sensitive to HC-3. These results indicate that choline uptake through CTL1 is used for ACh synthesis. Both an acetylcholinesterase inhibitor (eserine) and a butyrylcholinesterase inhibitor (ethopropazine) increased cell proliferation, and these effects were inhibited by 4-DAMP, a mAChR3 antagonist. We conclude that NCI-H69 cells express the choline transporter CTL1 which uses a directed H(+) gradient as a driving force, and its transport functions in co-operation with NHE1. This system primarily supplies choline for the synthesis of ACh and secretes ACh to act as an autocrine/paracrine growth factor, and the functional inhibition of CTL1 could promote apoptotic cell death. Identification of this new CTL1-mediated choline transport system provides a potential new target for therapeutic intervention.

Metabolic Imaging of Pancreatic Ductal Adenocarcinoma Detects Altered Choline Metabolism

“…Increased expression of CHT1 and CTL1 in other cancers has been previously reported (10, 13, 14). Other choline transporters and phospholipases (7) may also have contributed to the altered choline metabolism and should be further investigated.…”

“…Increased expression of the high-affinity choline transporter CHT1 with a K m of ~ 2 µM, also called solute carrier family 5 member 7 (SLC5A7) (12), has been previously observed in breast cancer cells (10). Another choline transporter, choline transporter-like protein 1 (CTL1) with a K m of ~ 68 µM, was found to be overexpressed in human lung and colon carcinoma cells (13, 14). …”

Knockdown by shRNA identifies SLC44A5 as a potential therapeutic target in hepatocellular carcinoma

“…Notably, the SLC44A1-5 family has been shown to be functionally important in the development of human lung, prostate and colon carcinoma (17)(18)(19)(20), however, the mechanisms underlying the effects of the SLC44A1-5 family remain to be elucidated. One of these proteins, SLC44A1, has previously been found to stimulate NCI-H69 cell growth and choline uptake, suggesting that SLC44A1 may function as a lung carcinogenic gene (17).…”

“…One of these proteins, SLC44A1, has previously been found to stimulate NCI-H69 cell growth and choline uptake, suggesting that SLC44A1 may function as a lung carcinogenic gene (17). The possible correlation between choline uptake and viability was also assessed (the effect of choline transporter inhibitors on the survival of various cancer cells).…”

Targeting Phospholipid Metabolism in Cancer

“…The choline analog hemicholinium-3 (HC-3) and tetrahexylammonium chloride were reported to inhibit choline uptake and reduce cell proliferation when treating the human colon carcinoma cell line HT-29 (144). The Na + /H + exchanger inhibitor dimethylamiloride and various organic cations such as quinine, quinidine, desipramine, imipramine, clomipramine, diphenhydramine, and fluvoxamine, among others were shown to inhibit choline uptake and cell viability in the small cell lung carcinoma cell line NCI-H69 (145). In the same study, HC-3 and CTL1 siRNA were shown to inhibit choline uptake and cell viability as well, along with increasing caspase-3/7 activity (145).…”