The Diagnostic Value of ACSL1, ACSL4, and ACSL5 and the Clinical Potential of an ACSL Inhibitor in Non-Small-Cell Lung Cancer

Abstract

Abnormal expression of ACSL members 1, 3, 4, 5, and 6 is frequently seen in human cancer; however, their clinical relevance is unclear. In this study, we analyzed the expression of ACSLs and investigated the effects of the ACSL inhibitor Triacsin C (TC) in lung cancer. We found that, compared to normal human bronchial epithelial (NHBE) cells, ACSL1, ACSL4, and ACSL6 were highly expressed, while ACSL3 and ACSL5 were lost in the majority of lung cancer cell lines. ACSL activity was associated with the expression levels of the ACSLs. In primary lung tumors, a higher expression of ACSL1, ACSL4, and ACSL5 was significantly correlated with adenocarcinoma (ADC). Moreover, ACSL5 was significantly reversely related to the proliferation marker Ki67 in low-grade tumors, while ACSL3 was positively associated with Ki67 in high-grade tumors. Combination therapy with TC and Gemcitabine enhanced the growth-inhibitory effect in EGFR wild-type cells, while TC combined with EGFR-TKIs sensitized the EGFR-mutant cells to EGFR-TKI treatment. Taken together, the data suggest that ACSL1 may be a biomarker for lung ADC, and ACSL1, ACSL4, and ACSL5 may be involved in lung cancer differentiation, and TC, in combination with chemotherapy or EGFR-TKIs, may help patients overcome drug resistance.

Keywords: ACSL inhibitor; ACSLs; combination therapy; diagnostic marker; lung cancer.

Figure 1

The expression of ACSL isoforms and ACSL enzymatic activity in normal bronchial epithelium (NHBE) cells and a panel of lung cancer cell lines. The expression of ACSL isoforms at both the mRNA and protein levels was analyzed by (A) RT-qPCR and (B) WB. Gene expression in comparison to the internal control GAPDH in NHBE cells was set to 1.0 for RT-qPCR analysis. GAPDH was used as a loading control for WB. (C) ACSL enzymatic activity was measured by liquid scintillation counting using [³H]-palmitic acid as the substrate. V: variant.

Figure 2

Tissue distribution and expression of ACSL1 and ACSL3 in normal human lung tissue. ACSL1 and ACSL3 were co-stained with cytokeratin (CK), surfactant protein C (SFPC), and DAPI in the same panel. (A) Alveolar type II cell. (B) Bronchial epithelial cells. (C) Macrophages.

Figure 3

Representative staining of ACSL isoforms in primary lung tumors.

Figure 4

The effect of Triacsin C (TC) on ACSL enzymatic activity and cell viability in lung cancer cell lines. (A) The effect of TC (2, 4, and 8 µM) on ACSL activity in H1650 and H1975 (upper panel) cells; the effect of 8 µM of TC on ACSL activity in five lung cancer cell lines (lower panel). (B) The effect of TC (8 µM) on ACSL5-overexpressing cells (upper panel) and ACSL5-knockdown cells (lower panel); (C). The effect of TC at serial concentrations on cell viability in six lung cancer cell lines. ** p < 0.01, *** p < 0.001 when analyzed using Student’s t-test.

Figure 5

The antitumor efficacy of Triacsin C combined with Gemcitabine or EGFR-TKIs in lung cancer cell lines, as revealed by a cell viability assay. (A) The effect of Gemcitabine alone or combined with TC in both EGFR-WT cell lines. (B) The effect of EGFR-TKI alone or combined with TC in EGFR-mutant cell lines.

Figure 6

The effect of ACSL1, ACSL4, or ACSL5 knockdown combined with Gemcitabine or Gefitinib on cell viability in lung cancer cell lines. (A) The effect of ACSL1 and ACSL4 knockdown combined with Gemcitabine on cell viability in H2170. (B) The effect of ACSL5 knockdown combined with Gemcitabine (left) or Gefitinib (right) on cell viability in H1650.