Precision-engineered reporter cell lines reveal ABCG2 regulation in live lung cancer cells

Kovacsics, Daniella; Brózik, Anna; Tihanyi, Borbála; Matula, Zsolt; Borsy, Adrienn; Mészáros, Nikolett; Szabó, Edit; Németh, Eszter; Fóthi, Ábel; Zámbó, Boglarka; Szüts, Dávid; Várady, György; Orbán, Tamás I.; Apáti, Ágota; Sarkadi, Balázs

doi:10.1016/j.bcp.2020.113865

Cited by 13 publications

(10 citation statements)

References 82 publications

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“…Abcg2 −/− knockout and wild-type mice were widely used to study the effect of ABCG2 on the tissue distribution of potential substrates by analyzed their plasma, small intestine, colorectal, liver, kidneys, and testicles (28,29). Recently, Daniella et al established ABCG2-knockout and EGFP tagged ABCG2 reporter cell lines in human lung adenocarcinoma cells, which were useful to study the ABCG2 gene regulation and visualizing protein activity in live cells (30). In this study, we established an ABCG2-knockout human colorectal cancer cell line by CRISPR-Cas9 mediated genome editing technology which we have used previously (31,32).…”

Section: Discussionmentioning

confidence: 99%

AZ32 Reverses ABCG2-Mediated Multidrug Resistance in Colorectal Cancer

Liu

Chen

et al. 2021

Front. Oncol.

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Colorectal cancer is a common malignancy with the third highest incidence and second highest mortality rate among all cancers in the world. Chemotherapy resistance in colorectal cancer is an essential factor leading to the high mortality rate. The ATP-binding cassette (ABC) superfamily G member 2 (ABCG2) confers multidrug resistance (MDR) to a range of chemotherapeutic agents by decreasing their intracellular content. The development of novel ABCG2 inhibitors has emerged as a tractable strategy to circumvent drug resistance. In this study, an ABCG2-knockout colorectal cancer cell line was established to assist inhibitor screening. Additionally, we found that ataxia-telangiectasia mutated (ATM) kinase inhibitor AZ32 could sensitize ABCG2-overexpressing colorectal cancer cells to ABCG2 substrate chemotherapeutic drugs mitoxantrone and doxorubicin by retaining them inside cells. Western blot assay showed that AZ32 did not alter the expression of ABCG2. Moreover, molecule docking analysis predicted that AZ32 stably located in the transmembrane domain of ABCG2. In conclusion, our result demonstrated that AZ32 could potently reverse ABCG2-mediated MDR in colorectal cancer.

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Section: Discussionmentioning

confidence: 99%

AZ32 Reverses ABCG2-Mediated Multidrug Resistance in Colorectal Cancer

Liu

Chen

et al. 2021

Front. Oncol.

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“…Hence, there is an increasing demand for new drugs for CSC-targeted therapy. In this context, we propose that CSCtargeting drug screening platforms can be developed using reporters of CSCs, which have been recently published (38)(39)(40)(41)(42)(43)(44)(45). These fluorescent reporters can be easily adapted for large scale screening.…”

Section: Clinical Trials For Targeting Cscsmentioning

confidence: 99%

“…Hence, a reporter cell line is recently made using CRISPR-Cas9 gene editing coupled with homology-directed repair. They targeted the EGFP coding sequence to the translational start site of ABCG2, generating ABCG2 knock-out and in situ tagged ABCG2 reporter cells (42). This fluorescent reporter system allowed the detection of endogenous regulation of ABCG2 expression by different stress responses and offers a method to screen molecules that can inhibit drug resistant CSCs.…”

Section: Abcg2 Reportermentioning

confidence: 99%

Reporters of Cancer Stem Cells as a Tool for Drug Discovery

Mohan

et al. 2021

Front. Oncol.

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In view of the importance of cancer stem cells (CSCs) in chemoresistance, metastasis and recurrence, the biology of CSCs were explored in detail. Based on that, several modalities were proposed to target them. In spite of the several clinical trials, a successful CSC-targeting drug is yet to be identified. The number of molecules screened and entered for clinical trial for CSC-targeting is comparatively low, compared to other drugs. The bottle neck is the lack of a high-throughput adaptable screening strategy for CSCs. This review is aimed to identify suitable reporters for CSCs that can be used to identify the heterogeneous CSC populations, including quiescent CSCs, proliferative CSCs, drug resistant CSCs and metastatic CSCs. Analysis of the tumor microenvironment regulating CSCs revealed that the factors in CSC-niche activates effector molecules that function as CSC markers, including pluripotency markers, CD133, ABCG2 and ALDH1A1. Among these factors OCT4, SOX2, NANOG, ABCG2 and ALDH1A1 are ideal for making reporters for CSCs. The pluripotency molecules, like OCT4, SOX2 and NANOG, regulate self-renewal, chemoresistance and metastasis. ABCG2 is a known regulator of drug resistance while ALDH1A1 modulates self-renewal, chemoresistance and metastasis. Considering the heterogeneity of CSCs, including a quiescent population and a proliferative population with metastatic ability, we propose the use of a combination of reporters. A dual reporter consisting of a pluripotency marker and a marker like ALDH1A1 will be useful in screening drugs that target CSCs.

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“…There are convincing data showing that the presence of ABCG2 variants may directly affect clinical tumor cell resistance against sunitinib [175] and gefitinib [17], or increase the toxic side effects of certain anticancer drugs [63,142,176]. In addition, in the cancer cells ABCG2 expression may be significantly upregulated either indirectly by hormonal or stress responses [161,162,164], or following a treatment with anticancer drugs, whether these are ABCG2 substrates or not [168,177,178].…”

Section: The Role Of Alterations In Abcg2 Expression In Cancer Drug Rmentioning

confidence: 99%

“…To further explore both the transcriptional‐ and the protein‐level regulations of ABCG2, recently we have generated fluorescence‐based reporter tumor cell lines by using the CRISPR‐Cas9 engineering system [178]. In A549 lung carcinoma cells, we have targeted an eGFP‐coding sequence to the translational start site of ABCG2 .…”

Section: Diseases Connected To the Altered Expression And Function Ofmentioning

confidence: 99%

The ABCG2/BCRP transporter and its variants – from structure to pathology

2020

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The ABCG2 protein has a key role in the transport of a wide range of structurally dissimilar endo-and xenobiotics in the human body, especially in the tissue barriers and the metabolizing or secreting organs. The human ABCG2 gene harbors a high number of polymorphisms and mutations, which may significantly modulate its expression and function. Recent high-resolution structural data, complemented with molecular dynamic simulations, may significantly help to understand intramolecular movements and substrate handling, as well as the effects of mutations on the membrane transporter function of ABCG2. As reviewed here, structural alterations may result not only in direct alterations in drug binding and transporter activity, but also in improper folding or problems in the carefully regulated process of trafficking, including vesicular transport, endocytosis, recycling, and degradation. Here, we also review the clinical importance of altered ABCG2 expression and function in general drug metabolism, cancer multidrug resistance, and impaired uric acid excretion, leading to gout.

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Precision-engineered reporter cell lines reveal ABCG2 regulation in live lung cancer cells

Cited by 13 publications

References 82 publications

AZ32 Reverses ABCG2-Mediated Multidrug Resistance in Colorectal Cancer

AZ32 Reverses ABCG2-Mediated Multidrug Resistance in Colorectal Cancer

Reporters of Cancer Stem Cells as a Tool for Drug Discovery

The ABCG2/BCRP transporter and its variants – from structure to pathology

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