Evaluation of engraftment and growth dynamics of orthotopic and heterotopic in vivo models of human breast cancer

Purpose of the study. This work was to assess the engraftment and growth dynamics of breast cancer xenografts during orthotopic and subcutaneous injection using various types of biological material, as well as to develop an adequate model of breast cancer for further research.

Materials and methods. We used a disaggregated fragment of a tumor obtained from the patient, a certified breast cancer cell line VT20 – human breast carcinoma; a primary human breast carcinoma cell line. Female immunodeficient mice of the Balb/c Nude line in the amount of 36 animals were used as recipient animals. The subcutaneous and orthotopic models of breast cancer were developed in this project. Tumor growth was observed for 28 days from the moment of injection and tumor nodes were measured 2 times a week until the end of the experiment. Results were assessed using medians and percentiles. The nonparametric Mann-Whitney test was used to assess the significance of differences.

Results. The dynamics of the growth of tumor cells when injected into various sites was determined in the process of this work. The most successful in terms of a subcutaneous injection was the injection of tumor cells of the certified VT20 line. By the end of the experiment, the median tumor node of this group was 100.32 mm³. The analysis revealed tumor dynamics with orthotopic injection of tumor material, and the median volume of the tumor node in the group with the passport culture cell VT20 and the primary culture cell reached the same value – 149.22 and 148.25. mm³. It was found that both the cell line and the cell suspension were injected into tumor nodes that reached a significantly larger volume when injected orthotopically.

Conclusion. We have obtained a tumor model of breast cancer using various methods of material implantation and with the possibility of further use in testing new pharmacological substances.

1. Shakhzadova AO, Starinsky VV, Lisichnikova IV. Cancer care to the population of Russia in 2022. Siberian journal of oncology. 2023;22(5):5–13. (In Russ.). https://doi.org/10.21294/1814-4861-2023-22-5-5-13, EDN: PESHHL

2. Zeng L, Li W, Chen CS. Breast cancer animal models and applications. Zool Res. 2020 Sep 18;41(5):477–494. https://doi.org/10.24272/j.issn.2095-8137.2020.095

3. Lee RS, Sad K, Fawwal DV, Spangle JM. Emerging Role of Epigenetic Modifiers in Breast Cancer Pathogenesis and Therapeutic Response. Cancers (Basel). 2023 Aug 7;15(15):4005. https://doi.org/10.3390/cancers15154005

4. Rajagopal K, Kalusalingam A, Bharathidasan AR, Sivaprakash A, Shanmugam K, Sundaramoorthy M, et al. In Silico Drug Design of Anti-Breast Cancer Agents. Molecules. 2023 May 18;28(10):4175. https://doi.org/10.3390/molecules28104175

5. Chan J, Wang X, Turner JA, Baldwin NE, Gu J. Breaking the paradigm: Dr Insight empowers signature-free, enhanced drug repurposing. Bioinformatics. 2019 Aug 15;35(16):2818–2826. https://doi.org/10.1093/bioinformatics/btz006

6. Ma J, Wang J, Ghoraie LS, Men X, Haibe-Kains B, Dai P. A Comparative Study of Cluster Detection Algorithms in Protein-Protein Interaction for Drug Target Discovery and Drug Repurposing. Front Pharmacol. 2019;10:109. https://doi.org/10.3389/fphar.2019.00109

7. Zhou H, Liu H, Yu Y, Yuan X, Xiao L. Informatics on Drug Repurposing for Breast Cancer. Drug Des Devel Ther. 2023;17:1933– 1943. https://doi.org/10.2147/DDDT.S417563

8. Centonze M, Di Conza G, Lahn M, Fabregat I, Dituri F, Gigante I, et al. Autotaxin inhibitor IOA-289 reduces gastrointestinal cancer progression in preclinical models. J Exp Clin Cancer Res. 2023 Aug 8;42(1):197. https://doi.org/10.1186/s13046-023-02780-4

9. Gao X, Yourick JJ, Sprando RL. Toxicological applications of human induced pluripotent stem cell-derived hepatocyte-like cells: an updated review. J Toxicol Sci. 2023;48(8):441–456. https://doi.org/10.2131/jts.48.441

10. Pellegrino B, Herencia-Ropero A, Llop-Guevara A, Pedretti F, Moles-Fernández A, Viaplana C, et al. Preclinical In Vivo Validation of the RAD51 Test for Identification of Homologous Recombination-Deficient Tumors and Patient Stratification. Cancer Res. 2022 Apr 15;82(8):1646–1657. https://doi.org/10.1158/0008-5472.CAN-21-2409

11. Singhal SS, Garg R, Mohanty A, Garg P, Ramisetty SK, Mirzapoiazova T, et al. Recent Advancement in Breast Cancer Research: Insights from Model Organisms-Mouse Models to Zebrafish. Cancers (Basel). 2023 May 29;15(11):2961. https://doi.org/10.3390/cancers15112961

12. Kit SO, Maksimov AYu, Goncharova AS, Kolesnikov EN, Sanamyants SV, Katsieva TB, et al. Growth features of patient-derived subcutaneous and orthotopic xenografts of human cardioesophageal cancer in immunodeficient mice. Modern Problems of Science and Education. 2020;(2):90. (In Russ.). https://doi.org/10.17513/spno.29573, EDN: OWIDSM

13. Zibirov RF, Mozerov SA. Characterization of the tumor cell microenvironment. Oncology. P. A. Herzen Journal. 2018;7(2):67– 72. (In Russ.). https://doi.org/10.17116/onkolog20187267-72

14. Chen S, Du Y, Guan XY, Yan Q. The current status of tumor microenvironment and cancer stem cells in sorafenib resistance of hepatocellular carcinoma. Front Oncol. 2023;13:1204513. https://doi.org/10.3389/fonc.2023.1204513

15. Zhang Y, Lv N, Li M, Liu M, Wu C. Cancer-associated fibroblasts: tumor defenders in radiation therapy. Cell Death Dis. 2023 Aug 22;14(8):541. doi.org/10.1038/s41419-023-06060-z

16. Pastushenko I, Brisebarre A, Sifrim A, Fioramonti M, Revenco T, Boumahdi S, et al. Identification of the tumour transition states occurring during EMT. Nature. 2018 Apr;556(7702):463–438. https://doi.org/10.1038/s41586-018-0040-3

17. Kuburich NA, Sabapathy T, Demestichas BR, Maddela JJ, den Hollander P, Mani SA. Proactive and reactive roles of TGF-β in cancer. Semin Cancer Biol. 2023 Oct;95:120–139. https://doi.org/10.1016/j.semcancer.2023.08.002

18. Qin Y, Peng F, Ai L, Mu S, Li Y, Yang C, et al. Tumor-infiltrating B cells as a favorable prognostic biomarker in breast cancer: a systematic review and meta-analysis. Cancer Cell Int. 2021 Jun 12;21(1):310. https://doi.org/10.1186/s12935-021-02004-9

19. Lindner S, Dahlke K, Sontheimer K, Hagn M, Kaltenmeier C, Barth TFE, et al. Interleukin 21-induced granzyme B-expressing B cells infiltrate tumors and regulate T cells. Cancer Res. 2013 Apr 15;73(8):2468–2479. https://doi.org/10.1158/0008-5472.CAN-12-3450

20. Liu X, Li X, Wei H, Liu Y, Li N. Mast cells in colorectal cancer tumour progression, angiogenesis, and lymphangiogenesis. Front Immunol. 2023;14:1209056. https://doi.org/10.3389/fimmu.2023.1209056

21. Nail HM, Chiu CC, Leung CH, Ahmed MMM, Wang HMD. Exosomal miRNA-mediated intercellular communications and immunomodulatory effects in tumor microenvironments. J Biomed Sci. 2023 Aug 21;30(1):69. https://doi.org/10.1186/s12929-023-00964-w

22. Krbal L, Soukup J, Stanislav J, Hanusova V. Derivation and basic characterization of colorectal carcinoma primary cell lines. Biomed Pap Med Fac Univ Palacky Olomouc Czech Repub. 2017 Dec;161(4):360–368. https://doi.org/10.5507/bp.2017.040

23. Timofeeva SV, Filippova SYu, Sitkovskaya AO, Gnennaya NV, Mezhevova IV, Shamova TV, et al. Bioresource collection of cell lines and primary tumors of the National Medical Research Center of Oncology of the Ministry of Health of Russia. Cardiovascular Therapy and Prevention. 2022;21(11):3397. (In Russ.). https://doi.org/10.15829/1728-8800-2022-3397

24. Janik K, Popeda M, Peciak J, Rosiak K, Smolarz M, Treda C, et al. Efficient and simple approach to in vitro culture of primary epithelial cancer cells. Biosci Rep. 2016 Dec;36(6):e00423. https://doi.org/10.1042/BSR20160208

25. Nishikata T, Ishikawa M, Matsuyama T, Takamatsu K, Fukuhara T, Konishi Y. Primary culture of breast cancer: a model system for epithelial-mesenchymal transition and cancer stem cells. Anticancer Res. 2013 Jul;33(7):2867–2873.

26. Charni-Natan M, Goldstein I. Protocol for Primary Mouse Hepatocyte Isolation. STAR Protoc. 2020 Sep 18;1(2):100086. https://doi.org/10.1016/j.xpro.2020.100086

27. Damm G, Schicht G, Zimmermann A, Rennert C, Fischer N, Kießig M, et al. Effect of glucose and insulin supplementation on the isolation of primary human hepatocytes. EXCLI J. 2019;18:1071–1091. https://doi.org/10.17179/excli2019-1782

28. Skarkova V, Krupova M, Vitovcova B, Skarka A, Kasparova P, Krupa P, et al. The Evaluation of Glioblastoma Cell Dissociation and Its Influence on Its Behavior. Int J Mol Sci. 2019 Sep 18;20(18):4630. https://doi.org/10.3390/ijms20184630

29. Kapałczyńska M, Kolenda T, Przybyła W, Zajączkowska M, Teresiak A, Filas V, et al. 2D and 3D cell cultures – a comparison of different types of cancer cell cultures. Arch Med Sci. 2018 Jun;14(4):910–919. https://doi.org/10.5114/aoms.2016.63743

30. Long Y, Xie B, Shen HC, Wen D. Translation Potential and Challenges of In Vitro and Murine Models in Cancer Clinic. Cells. 2022 Nov 30;11(23):3868. https://doi.org/10.3390/cells11233868