Cellular irinotecan resistance in colorectal cancer and overcoming irinotecan refractoriness through various combination trials including DNA methyltransferase inhibitors: a review

Abstract

Treatment with pharmacological drugs for colorectal cancer (CRC) remains unsatisfactory. A major cause of failure in pharmacotherapy is the resistance of colon cancer cells to the drugs, creating an urgent issue. In this review, we summarize previous studies on the resistance of CRC cells to irinotecan and discuss possible reasons for refractoriness. Our review presents the following five major causes of irinotecan resistance in human CRC: (1) cellular irinotecan resistance is induced mainly through the increased expression of the drug efflux transporter, ABCG2; (2) cellular irinotecan resistance is also induced in association with a nuclear receptor, pregnane/steroid X receptor (PXR/SXR), which is enriched in the CYP3A4 gene enhancer region in CRC cells by exposing the cells to SN-38; (3) irinotecan-resistant cells possess either reduced DNA topoisomerase I (Top1) expression at both the mRNA and protein levels or Top1 missense mutations; (4) alterations in the tumor microenvironment lead to drug resistance through intercellular vesicle-mediated transmission of miRNAs; and (5) CRC stem cells are the most difficult targets to successfully treat CRC. In the clinical setting, CRC gradually develops resistance to initially effective irinotecan-based therapy. To solve this problem, several clinical trials, such as irinotecan plus cetuximab vs. cetuximab monotherapy, have been conducted. Another clinical trial on irinotecan plus guadecitabine, a DNA-methyltransferase inhibitor, has also been conducted.

Keywords: ABCG2; DNA topoisomerase I; Drug resistance; anti-cancer drugs; cancer stem cells; colorectal cancer; epigenetics; irinotecan.

Figure 1

Irinotecan, the active metabolite, SN-38, and its glucuronide (A), and formation of inhibitory complex of Top1-DNA-SN-38 much more potent than irinotecan (B). In liver, irinotecan is bioactivated through hydrolysis catalyzed by carboxylesterases to be converted into SN-38, an active metabolite of irinotecan. SN-38 undergoes glucuronide conjugation to be SN-38 glucuronide by a UDP-glucuronosyltransferase, UGT1A1, for the detoxification pathway. SN-38 glucuronide is hydrolyzed by b-glucuronidase after being excreted in the gut. In the tumor cells, SN-38 can target the Top1-DNA covalent reaction intermediates and reversibly stabilize the Top1-DNA-SN-38 complex. As illustrated in (B), the collision of the DNA replication fork, together with this ternary complex formation, ultimately results in lethal and irreversible double-strand breaks^[4]. Top1: Topoisomerase I.

Figure 2

Factors causing resistance to anti-cancer drugs in ABCB1- or ABCG2-overexpressing colon cancer cell lines and an ABCB1-overexpressing HeLa cell line. Resistance factors for SN-38, irinotecan, topotecan, mitoxantrone, anthracyclines, 5-fluorouracil, and platinum anticancer drugs are shown relative to the respective parental cell lines. Cell lines are HCT116-SN6 [2004 (COL), ABCG2-overexpressing colon cancer cell^[10] (orange)], S1-IR20 [2021 (COL), ABCG2-overexpressing colon cancer cell^[11] (gray)], S1-B1-20 [2000 (COL), ABCB1-overexpressing colon cancer cell^[12] (yellow)], and Hvr100-6 [2002 (HeLa), ABCB1-overexpressing HeLa cell^[13] (blue)]. Establishments of these resistant sublines are described in Table 1. ND: Not determined.

Figure 3

Epigenetic regulation mechanisms in cancer cells that result in acquired irinotecan resistance. Methylation in a promoter region of a drug efflux transporter, ABCG2, and stabilization of a mRNA binding protein, HuR, through miR-519c function are depicted as epigenetic regulation mechanisms. When positive regulation mechanisms are prevailing in cancer cells, ABCG2 gene expression will be enhanced. Thus, cancer cells will become resistant to irinotecan.

Figure 4

Elevated cellular drug metabolism in relation to pregnane X receptor or steroid and xenobiotic receptor. Induction of glucuronidation of SN-38 within colorectal cancer cells renders cells resistant to irinotecan-based therapy. PXR and SXR are well known as they are expressed mainly in mammalian livers and gastrointestinal tracts and are involved in the induction of various classes of drug-metabolizing enzymes and drug transporters to detoxify therapeutic drugs such as irinotecan and SN-38. PXR: Pregnane X receptor; SXR: steroid and xenobiotic receptor; RXR: retinoid X receptor.

Figure 5

Factors affecting levels of the Top1-DNA cleavage complex and irinotecan resistance. Levels of irinotecan-causing DNA damage response depends upon formation of the Top1-DNA cleavage complex through Top1 function. Decreases in cellular Top1 activities, Top1 gene copy number, and Top1 protein levels result in cellular irinotecan resistance. Top1: Topoisomerase I.

Figure 6

Transition of colon cancer cell phenotypes in relation to alteration of status of CXCR4 and CRC markers for acquiring irinotecan resistance. The G-protein coupled chemokine receptor, CXCR4, co-localizes with CRC stem cell markers, such as Lgr5, CD133, and CD44, which are thought to be associated with the epithelial-mesenchymal transition process and resistance to cancer drug therapy. CSC: Cancer stem cell; SDF-1α: stromal cell-derived factor-1α; TGFα: tumor growth factor-α; ERK1/2: extracellular signal-regulated kinase 1/2.

Figure 7

The tumor microenvironment may affect cancer cell properties and drug sensitivity/resistance. Scaffolds of cancer tissues were prepared by decellularization. Human colorectal cancer cells, HT-29, were inoculated on the resultant scaffolds to form 2D or 3D cultures of HT-29 cells. HT-29 cells in the 3D culture systems were created using scaffolds from healthy liver or colorectal cancer liver metastasis tissue, which showed overall resistance compared to 2D-cultured HT-29 cells. 3D-HT-29 cells on CRLM scaffolds were resistant to 5-fluorouracil. PBMC: Peripheral blood mononuclear cell; CRLM: colorectal cancer liver metastasis; HL: healthy liver; 5-FU: 5-fluorouracil; TGF-β: transforming growth factor-β; IL-10: interleukin-10; Treg: regulatory T cell.

Figure 8

Cancer stem cells within a colorectal cancer tissue and drug resistance or drug sensitive phenotype. Tumorigenic cells in colon cancer existed as a high-density CD133⁺ population. Small population of CD133⁺ cells that are tumorigenic and drug resistant when subcutaneously implanted in immunodeficient mice reproduce the original tumor properties. In contrast, CD133^- cells do not develop tumors and show drug sensitive phenotype. CSC: Cancer stem cell.