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. 2022 Nov 1;12(1):18423.
doi: 10.1038/s41598-022-21856-3.

Testis-expressed gene 11 inhibits cisplatin-induced DNA damage and contributes to chemoresistance in testicular germ cell tumor

Sachi Kitayama  1   2 , Kazuhiro Ikeda  1 , Wataru Sato  1 , Hideki Takeshita  2 , Satoru Kawakami  2 , Satoshi Inoue  3   4 , Kuniko Horie  5
Affiliations

Testis-expressed gene 11 inhibits cisplatin-induced DNA damage and contributes to chemoresistance in testicular germ cell tumor

Sachi Kitayama et al. Sci Rep. .

Abstract

Testicular germ cell tumor (TGCT) is a rare cancer but the most common tumor among adolescent and young adult males. Patients with advanced TGCT often exhibit a worse prognosis due to the acquisition of therapeutic resistance. Cisplatin-based chemotherapy is a standard treatment for advanced TGCTs initially sensitive to cisplatin, as exemplified by embryonal carcinoma. The acquisition of cisplatin resistance, however, could be a fatal obstacle for TGCT management. To identify cisplatin resistance-related genes, we performed transcriptome analysis for cisplatin-resistant TGCT cells compared to parental cells. In two types of cisplatin-resistant TGCT cell models that we established from patient-derived TGCT cells, and from the NEC8 cell line, we found that mRNA levels of the high-mobility-group nucleosome-binding gene HMGN5 and meiosis-related gene TEX11 were remarkably upregulated compared to those in the corresponding parental cells. We showed that either HMGN5 or TEX11 knockdown substantially reduced the viability of cisplatin-resistant TGCT cells in the presence of cisplatin. Notably, TEX11 silencing in cisplatin-resistant TGCT cells increased the level of cleaved PARP1 protein, and the percentage of double-strand break marker γH2AX-positive cells. We further demonstrated the therapeutic efficiency of TEX11-specific siRNA on in vivo xenograft models derived from cisplatin-resistant patient-derived TGCT cells. Taken together, the present study provides a potential insight into a mechanism of cisplatin resistance via TEX11-dependent pathways that inhibit apoptosis and DNA damage. We expect that our findings can be applied to the improvement of cisplatin-based chemotherapy for TGCT, particularly for TEX11-overexpressing tumor.

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Conflict of interest statement

The authors declare no competing interests.

Figures

Figure 1
Figure 1
Generation of cisplatin-resistant testicular germ cell tumor (TGCT) cells and identification of upregulated genes in cisplatin-resistant TGCT cells versus parental cells. (a) Inhibitory effects of cisplatin to the viability of parental and cisplatin-resistant TGCT patient-derived cells (TGCT-PDC). Cells were treated with cisplatin at indicated concentrations for 96 h and subjected to the quantitation of intracellular ATP content. CDDP, cisplatin. TGCT-PDC-R, cisplatin-resistant TGCT-PDC. Results are shown as mean fold change ± SE of relative luciferase activity (n = 3). (b) Inhibitory effects of cisplatin to the viability of parental and cisplatin-resistant NEC8 cells. Cells were treated with cisplatin at indicated concentrations for 72 h and subjected to WST-8 cell proliferation assay. Results are shown as mean fold change ± SE of relative absorbance at 450 nm wavelength (n = 5). NEC-R, cisplatin-resistant NEC8 cells. (c) Microarray analysis identified 334 overlapping genes commonly upregulated by ≥ 1.5 fold with a fluorescence signal ≥ 5 in cisplatin-resistant cells compared to the corresponding parental cells. (d) Top 5 signaling pathways enriched among the 334 commonly upregulated genes in cisplatin-resistant TGCT-PDC and NEC8 cells based on Gene Ontology Term. (e,f) Overexpression of HMGN5 (e) and TEX11 (f) in cisplatin-resistant TGCT cells. P, parental cells. R, cisplatin-resistant cells. Results are shown as mean ± SE (n = 3). **, P < 0.01.
Figure 2
Figure 2
Silencing of HMGN5 and TEX11 attenuates the proliferation of cisplatin-resistant TGCT-PDC and NEC8 cells in the presence of cisplatin. (a, b) Effects of control siRNA (siControl), HMGN5-specific siRNAs (siHMGN5 #1 and #2), or TEX11-specific siRNAs (siTEX11 #1 and #2) on the viability of parental TGCT-PDC (a) and cisplatin-resistant TGCT-PDC-R (b) cells analyzed by quantitation of intracellular ATP content in the absence of cisplatin. Results are shown as mean ± SE (n = 4). (c, d) HMGN5- and TEX11-specific siRNAs repress the viability of TGCT-PDC-R cells (d) but not of TGCT-PDC cells (c) treated with cisplatin (0.2 μM). (e, f) Effects of indicated siRNAs on the viability of parental NEC8 (e) and cisplatin-resistant NEC8-R (f) cells analyzed by WST-8 cell proliferation assay in the absence of cisplatin. (g, h) HMGN5- and TEX11-specific siRNAs repress the viability of NEC8-R cells (g) but not of NEC8 cells (h) treated with cisplatin (0.2 μM). *, P < 0.05; **, P < 0.01.
Figure 3
Figure 3
Effects of TEX11 expression on cisplatin-induced apoptosis and DNA double-strand break (DSB) marker γH2AX expression in TGCT cells. (a, b) Immunoblotting of cleaved PARP1 in cisplatin-resistant TGCT-PDC-R (a) and NEC8-R (b) cells transfected with control (siControl) or TEX11-specific siRNAs (siTEX11 #1 and #2), without or with cisplatin treatment 48 h after siRNA transfection. β-actin was used as a loading control. (c) Representative γH2AX immunohistochemical staining in TGCT-PDC-R cells transfected with indicated siRNAs, followed by cisplatin treatment 6 h after siRNA transfection. (d) Percentages of γH2AX-positive cells among the examined TGCT-PDC-R cells treated with the indicated siRNAs. The results are shown as mean percentage ± SE. (e) Representative γH2AX immunohistochemical staining in NEC8-R cells transfected with indicated siRNAs in the presence of cisplatin. (f) Percentages of γH2AX-positive cells among the examined NEC8-R cells treated with the indicated siRNAs. *, P < 0.05.
Figure 4
Figure 4
TEX11-specific siRNA injection significantly represses the development of TGCT-PDC-R-derived xenograft tumors in severely immunodeficient mice treated with cisplatin. (a) Volumes of tumors generated by TGCT-PDC-R cells in 7-week male NOD/SCID mice treated with intratumoral injection of control (siControl) or TEX11-specific (siTEX11) siRNAs plus intraperitoneal cisplatin administration. (b)Weights of tumors generated by TGCT-PDC-R in mice treated with injection of indicated siRNAs plus cisplatin administration. (c) Body weights of tumor-bearing mice treated with injection of indicated siRNAs plus cisplatin administration. Results are shown as mean ± SE. **, P < 0.01. (d) Images of dissected tumors at the endpoint. (e) Representative images of tumor-bearing mice at the endpoint. (f) Immunoblotting of TEX11 and cleaved PARP1 in dissected TGCT-PDC-R tumors. β-actin was used as a loading control. (g) Relative expression of TEX11 and cleaved PARP1 normalized to β-actin expression analyzed by densitometry. Results are shown as mean ± SE (n = 5). *, P < 0.05, **, P < 0.01.
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References

    1. Cheng L, et al. Testicular cancer. Nat. Rev. Dis. Primers. 2018;4:29. doi: 10.1038/s41572-018-0029-0. - DOI - PubMed
    1. McHugh DJ, Feldman DR. Conventional-dose versus high-dose chemotherapy for relapsed germ cell tumors. Adv. Urol. 2018;2018:7272541. doi: 10.1155/2018/7272541. - DOI - PMC - PubMed
    1. Galluzzi L, et al. Molecular mechanisms of cisplatin resistance. Oncogene. 2012;31:1869–1883. doi: 10.1038/onc.2011.384. - DOI - PubMed
    1. Eastman A. Reevaluation of interaction of cis-dichloro (ethylenediamine)platinum(II) with DNA. Biochemistry. 1986;25:3912–3915. doi: 10.1021/bi00361a026. - DOI - PubMed
    1. Huang JC, Zamble DB, Reardon JT, Lippard SJ, Sancar A. HMG-domain proteins specifically inhibit the repair of the major DNA adduct of the anticancer drug cisplatin by human excision nuclease. Proc. Natl. Acad. Sci. USA. 1994;91:10394–10398. doi: 10.1073/pnas.91.22.10394. - DOI - PMC - PubMed

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