**5. Conclusions**

In this study, we unveiled the cytotoxicity of CPT in GSC loss and two-cell differentiation defects, which could provide information for its therapeutic application. CPT could induce ectopic expression of Bam in GSCs via top1, and such a phenotype could be enhanced by overexpression of CycA, which might contribute to the observed GSC loss. In addition, CPT can cause DNA damage in the early germline cell by regulating Bam expression at both transcript and protein level, thus leading to cell arrest at G1/S and two-cell accumulation. Collectively, the results in this study provided convincing results that CPT may have therapeutic potential as an anticancer agent in germ cells. Further study is needed to evaluate the safety of CPT in advanced models to confirm the mechanism in germline cells of other organisms.

**Supplementary Materials:** The following supporting information can be downloaded at: https: //www.mdpi.com/article/10.3390/ijms24021617/s1.

**Author Contributions:** Conceptualization, J.Z. and Z.S.; methodology, J.Z., Z.S., S.Z. and Y.C.; validation, G.Z.; formal analysis, J.Z., G.Z. and X.Y.; investigation, J.Z., Z.S. and Y.C.; data curation, J.Z., Y.C. and X.Y.; writing—original draft, J.Z. and X.Y.; writing—review and editing, J.Z.; supervision, G.Z. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research was funded by National natural science foundation of China (No. 31572335) and National natural science foundation of Guangdong Province (No. 2021A1515110795).

**Institutional Review Board Statement:** Not applicable.

**Data Availability Statement:** All data generated or analyzed during this study are included in this published article and its Supplementary Materials. Reagents used in this publication will be provided upon request.

**Conflicts of Interest:** The authors declare no conflict of interest.
