Dynamics of subpopulations of CD15⁺CD66b⁺ and CD62L⁺CD63⁺ circulating neutrophils in patients with benign neoplasms and progression of kidney cancer

Introduction. Due to the ambiguous role of neutrophils in carcinogenesis, it is relevant to study their phenotypic transformation and subpopulation composition that determines protumor (CD15⁺CD66b⁺) or antitumor (CD62L⁺CD63⁺) potential.

Aim. To assess the dynamics of CD15⁺CD66b⁺ and CD62L⁺CD6⁺ circulating neutrophil subpopulations in patients with benign neoplasms and during the progression of kidney cancer.

Materials and methods. The study focused on circulating neutrophils from patients with benign neoplasms and kidney cancer. The phenotype of monopopulations (CD15⁺, CD66b⁺, CD62L⁺, CD63⁺, CD95⁺) and subpopulations (CD15⁺CD66b⁺, CD62L⁺CD63⁺) of neutrophils was assessed using flow cytometry (BioSino, China). Statistical analysis was performed using Statistica 13 and Jamovi 2.3.28.

Results. A significant increase in the number of CD15⁺, CD62L⁺, and CD66b⁺ neutrophil monopopulations, as well as in the number of CD15⁺CD66b⁺ neutrophils, was observed in the groups of patients with benign kidney tumors and kidney cancer compared to the control group. In patients with benign renal tumors, the percentage of CD62L⁺CD63⁺  neutrophils was three times higher than in patients with stage I–II kidney cancer, and twice as high as in those with advanced kidney cancer. According to the Cox regression model, changes in the numbers of CD15⁺CD66b⁺ and CD62L⁺CD63⁺  neutrophils, alongside an increase in leukocyte count, serve as prognostic markers of kidney cancer in patients over 68 years of age.

Conclusion. The number of circulating neutrophils with a protumor phenotype (CD15⁺CD66b⁺) increases even at the stage of benign kidney neoplasms compared to controls and remains elevated throughout all stages of carcinogenesis. Meanwhile, the number of circulating neutrophils with an antitumor phenotype (CD62L⁺CD63⁺) significantly increases in benign renal neoplasms but decreases during cancer progression. Assessment of the circulating neutrophil phenotype may help predict the risk of kidney neoplasms.

1. Li P., Znaor A., Holcatova I. et al. Regional geographic variations in kidney cancer incidence rates in European countries. Eur Urol 2015;67(6):1134–41. DOI: 10.1016/j.eururo.2014.11.001

2. Motzer R.J., Jonasch E., Agarwal N. et al. Kidney cancer, version 3.2022, NCCN clinical practice guidelines in oncology. J Natl Compr Cancer Netw 2022;20:71–90. DOI: 10.6004/jnccn.2022.0001

3. Korotaeva A.A., Apanovich N.V., Braga E.A. et al. Current advances in kidney cancer immunotherapy. Onkourologiya = Cancer Urology 2019;15(4):30–8. (In Russ.). DOI: 10.17650/1726-9776-2019-15-4-30-38

4. De Palma M., Biziato D., Petrova T. V. Microenvironmental regulation of tumour angiogenesis. Nat Rev Cancer 2017;17(8):457–74. DOI: 10.1038/nrc.2017.51

5. Anker J., Miller J., Taylor N. et al. From bench to bedside: how the tumor microenvironment is impacting the future of immunotherapy for renal cell carcinoma. Cells 2021;10(11):3231. DOI: 10.3390/cells10113231

6. Savchenko A.A., Borisov A.G., Kudryavcev I.V. et al. The relationship between the phenotype and metabolism of blood neutrophils in patients with kidney cancer. Meditsinskaya immunologiya = Medical Immunology 2020;22(5):887–96. (In Russ.). DOI: 10.15789/1563-0625-IAM-2037

7. Danilova A.B., Baldueva I.A. Neutrophils as a component of the tumor microenvironment. Voprosy onkologii = Oncology Issues 2016;62(1):35–44. (In Russ.).

8. Mishalian I., Granot Z., Fridlender Z.G. The diversity of circulating neutrophils in cancer. Immunobiology 2017;222(1):82–8. DOI: 10.1016/j.imbio.2016.02.001

9. Silvestre-Roig C., Fridlender Z.G., Glogauer M., Scapini P. Neutrophil diversity in health and disease. Trends Immunol 2019;40(7):565–83. DOI: 10.1016/j.it.2019.04.012

10. Evrard M., Kwok I.W.H., Chong S.Z. et al. Developmental analysis of bone marrow neutrophils reveals populations specialized in expansion, trafficking, and effector functions. Immunity 2018; 48(2):364–79. DOI: 10.1016/j.immuni.2018.02.002

11. Lecot P., Sarabi M., Pereira Abrantes M. et al. Neutrophil heterogeneity in cancer: from biology to therapies. Front Immunol 2019;10:2155. DOI: 10.3389/fimmu.2019.02155

12. Savchenko A.A., Zdzitovetskiy D.E., Borisov A.G., Luzan N.A. Chemiluminescent and enzyme activity of neutrophils in patients with wides preadpurulent peritonitis depending on the outcome of disease. Vestnik RAMN = Annals of the Russian Academy of Medical Sciences 2014;5(6):23–8. (In Russ.).

13. Lin A., Lore K. Granulocytes: new members of the antigen presenting cell family. Front Immunol 2017;11:1781. DOI: 10.3389/fimmu.2017.01781

14. Nesterova I.V., Chudilova G.A., Kovaleva S.V. et al. Contradictory effect of recombinant interferon α2b on the non-transformed and transformed phenotypes of functionally significant subpopulations of neutrophilic granulocytes in vitro. Immunologiya = Immunology 2020; 41(2):124–34. (In Russ.). DOI: 10.33029/0206-4952-2020-41-2-124-134

15. Wang Z., Yang C., Li L. et al. CD62Ldim neutrophils specifically migrate to the lung and participate in the formation of the pre-metastatic niche of breast cancer. Front Oncol 2020;10:540484. DOI: 10.3389/fonc.2020.540484

16. Peng Z., Liu C., Victor A.R. et al. Tumors exploit CXCR4hiCD62Llo aged neutrophils to facilitate metastatic spread. Oncoimmunology 2021;10(1):1870811. DOI: 10.1080/2162402X.2020.1870811 17. Chudilova G.A., Nesterova I.V., Kovaleva S.V., Lomtatidze L.V. Regulatory cytokine effects in vitro on the phenotype of subpopulations CD62L+CD63–, CD62L+CD63+ and microbicidal activity of neutrophilic granulocytes in patients with colorectal cancer. Vestnik Rossiyskogo universiteta druzhby narodov = RUDN Journal of Medicine 2020;24(4):304–14. (In Russ.). DOI: 10.22363/2313-0245-2020-24-4-304-314

17. Wang Z., Yang C., Li L. et al. CD62Ldim neutrophils specifically migrate to the lung and participate in the formation of the pre-metastatic niche of breast cancer. Front Oncol 2020;10:540484. DOI: 10.3389/fonc.2020.540484

18. Rice C.M., Davies L.C., Subleski J.J. et al. Tumour-elicited neutrophils engage mitochondrial metabolism to circumvent nutrient limitations and maintain immune suppression. Nat Commun 2018;9(1):5099. DOI: 10.1038/s41467-018-07505-2

19. Blanco-Camarillo C., Aleman O.R., Rosales C. Low-density neutrophils in healthy individuals display a mature primed phenotype. Front Immunol 2021;12:672520. DOI: 10.3389/fimmu.2021.672520

20. Zhang J., Xu X., Shi M. et al. CD13hi neutrophil-like myeloid-derived suppressor cells exert immune suppression through Arginase 1 expression in pancreatic ductal adenocarcinoma. Oncoimmunology 2017;6(2):e1258504. DOI: 10.1080/2162402X.2016.1258504

21. Vanhaver C., Aboubakar Nana F. , Delhez N. et al. Immunosuppressive low-density neutrophils in the blood of cancer patients display a mature phenotype. Life Sci Alliance 2023;7(1):e202302332. DOI: 10.26508/lsa.202302332

22. Albiges L., Tannir N.M., Burotto M. et al. Nivolumab plus ipilimumab versus sunitinib for first-line treatment of advanced renal cell carcinoma: extended 4-year follow-up of the phase III CheckMate 214 trial. ESMO Open 2020;5(6):e001079. DOI: 10.1136/esmoopen-2020-001079

23. Tannir N.M., Signoretti S., Choueiri T.K. et al. Efficacy and safety of nivolumab plus ipilimumab versus sunitinib in first-line treatment of patients with advanced sarcomatoid renal cell carcinoma. Clin Cancer Res 2021;27(1):78–86. DOI: 10.1158/1078-0432.CCR-20-2063

24. Dumitru C.A., Lang S., Brandau S. Modulation of neutrophil granulocytes in the tumor microenvironment: mechanisms and consequences for tumor progression. Semin Cancer Biol 2013;23(3):141–8. DOI: 10.1016/j.semcancer.2013.02.005

25. Rosellini M., Marchetti A., Mollica V. et al. Prognostic and predictive biomarkers for immunotherapy in advanced renal cell carcinoma. Nat Rev Urol 2023;20(3):133–57. DOI: 10.1038/s41585-022-00676-0

26. Kraus R.F., Gruber M.A. Neutrophils-from bone marrow to first-line defense of the innate immune system. Front Immunol 2021;12:1–35. DOI: 10.3389/fimmu.2021.767175

27. Kanamaru R., Ohzawa H., Miyato H. et al. Low density neutrophils (LDN) in postoperative abdominal cavity assist the peritoneal recurrence through the production of neutrophil extracellular traps (NETs). Sci Rep 2018;8(1):632. DOI: 10.1038/s41598-017-19091-2

28. Risso V., Lafont E., LeGallo M. Therapeutic approaches targeting CD95L/CD95 signaling in cancer and autoimmune diseases. Cell Death Dis 2022;13(3):248. DOI: 10.1038/s41419-022-04688-x

29. Macher-Goeppinger S., Bermejo J.L., Wagener N. et al. Expression and prognostic relevance of the death receptor CD95 (Fas/APO1) in renal cell carcinomas. Cancer Lett 2011;301(2):203–11. DOI: 10.1016/j.canlet.2010.12.005

30. Nonomura N., Nishimura K., Ono Y. et al. Soluble Fas in serum from patients with renal cell carcinoma. Urology 2000;55(1):151–5. DOI: 10.1016/s0090-4295(99)00379-9

31. Dominguez-Luis M.J., Maria Jesus E. , Herrera-García A. et al. L-selectin expression is regulated by CXCL8-induced reactive oxygen species produced during human neutrophil rolling. Eur J Immunol 2019;49(3):386–97. DOI: 10.1002/eji.201847710

32. Chandrasekaran D., Sundaram S., Maheshkumar K. et al. Preoperative neutrophil-lymphocyte ratio/platelet-lymphocyte ratio: a potential and economical marker for renal cell carcinoma. J Cancer Res Ther 2022;18(6):1635–9. DOI: 10.4103/jcrt.JCRT_482_20

33. Sejima T., Iwamoto H., Morizane S. The significant immunological characteristics of peripheral blood neutrophil-to-lymphocyte ratio and Fas ligand expression incidence in nephrectomized tumor in late recurrence from renal cell carcinoma. Urol Oncol 2013;31(7):1343–9. DOI: 10.1016/j.urolonc.2011.09.008

34. Potapnev M.P., Gushina L.M., Moroz L.A. Phenotypic and functional heterogeneity of neutrophil subpopulations in normal and pathological conditions. Immunologiya = Immunology 2019; 40 (5):84–96. (In Russ.). DOI: 10.24411/0206-4952-2019-15009

35. Lai X., Gu Q., Zhou X. et al. Decreased expression of CD63 tetraspanin protein predicts elevated malignant potential in human esophageal cancer. Oncol Lett 2017;13(6):4245–51. DOI: 10.3892/ol.2017.6023