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  • [ Recombinant Protein ] The Impact of Cytokines on NK Cells: Shaping Their Activity and Function

[ Recombinant Protein ] The Impact of Cytokines on NK Cells: Shaping Their Activity and Function

2023-06-05

 


A class of signaling molecules known as cytokines has gained prominence as an essential player in the field of natural killer (NK) cell treatment. These small proteins modulate NK cell activity and function in a variety of ways, with significant impacts. By understanding the specific roles of cytokines, we gain valuable insights into how they can be harnessed to enhance the therapeutic potential of NK cells in immunotherapies and cancer treatments.
 

  • IL-2

a. Enhancement of Expansion
IL-2 directly stimulates NK cell proliferation through binding to the IL-2 receptor, promoting cell division and expansion. This cytokine also synergizes with other factors and contributes to the autocrine loop of NK cell activation-induced proliferation, ultimately leading to the generation of a larger and functionally competent NK cell population[1].
 
b. Enhancement of Activation and Cytotoxicity
IL-2-activated NK cells exhibit potent tumor cell lysis capabilities, as evidenced by the significant ability of their granules, such as perforin and granzymes, to induce tumor cell death[2].
 
c. Enhancement of IFN-γ Production
In the study, it was found that when activated with IL-2, NK-92 cells produced and secreted more IFN-γ. IFNs have been demonstrated to have great therapeutic efficacy against several diseases, in contrast to the different commercially available enzymes, vaccinations, antibodies, and antibiotics[3].
 
d. Synergy with IL-15

  • IL-12

a. Enhancement of Activation and Cytotoxicity
NK cells are successfully activated by IL-12 in a low dose. By upregulating cytotoxic chemicals like perforin and granzymes, it increases the cytotoxicity of NK cells against target cells, enhancing their ability to kill[2].
 
b. Differentiation
The effects of IL-12 on NK cell differentiation are mediated through the regulation of specific transcription factors. Notably, IL-12 leads to increased expression of ID3, while downregulating Gata3 and TOX. These transcription factors are involved in controlling NK cell development and function[4].
 
c. Enhancement of Cytokines Production
When triggered by IL-12 and HER2-overexpressing tumor cells covered in Herceptin, NK cells release potent immune-stimulating molecules such as IFN-γ, TNF-α, MIP-1, and GM-CSF. According to the study, the production of this specific group of cytokines is strongly synergized by two distinct signals that pass from the cell surface to the nucleus via FcRIII and IL-12R[5].
 
d. Cytokine-Induced Memory-Like (ML or CIML) NK cells Formation
Human NK cells, when activated with a combination of IL-12, IL-15, and IL-18, undergo differentiation into memory-like cells. Upon subsequent restimulation with cytokines (IL-12 + IL-15 or IL-12 + IL-18) or exposure to K562 leukemia targets, these previously stimulated NK cells exhibit a significant increase in the production of IFN-γ[6]. 
 

  • IL-15

a. Homeostasis and Survival
IL-15 acts on NK cells to reduce the levels of Bim, a protein involved in cell death regulation. It achieves this through two signaling pathways: phosphorylation of Bim by Erk1/2, leading to its degradation via the proteasome, and activation of the PI(3)K-Akt pathway, which phosphorylates Foxo3a and prevents it from increasing Bim production[7].
 
b. Enhancement of Expansion and Activation
IL-15 shares similar biological effects with IL-2 and exhibits a synergistic effect when combined with IL-2 in promoting the proliferation and activation of different subsets of NK cells[8], especially a high dose of IL-15 with a low dose of IL-2 on CD56bright NK cells.
 
c. Enhancement of Cytotoxicity and Cytokines Production
IL-15 enhances the production of cytokines and chemokines, including IFN-γ, by NK cells, leading to an enhanced immune response. It also improves the ability of NK cells to ADCC against target cells. IL-15 therapy increases the expression of receptors involved in NK cell activation, such as NKG2D, NKp30, NKp46, and CD16[9].
 
d. Cytokine-Induced Memory-Like (ML or CIML) NK cells Formation
IL-15 plays a vital role in the formation of memory NK cells, which undergo proliferation in vivo but return to a resting state one week after adoptive transfer. These ML NK cells, exhibit enhanced IFN-γ response upon restimulation compared to control and host NK cells. Importantly, this increased IFN-γ production is inherent to the ML NK cells themselves, independent of their proliferative capacity, and persists even after cell division[10].
 
e. Synergy with IL-2, IL-12, IL-18[11], TNF-α[12]

→ Croyez GMP ® IL-15 (Interleukin-15), Human
→ Brochure


 

  • IL-18

a. Enhancement of Expansion and Activation
Based on the research, the IL-2/ IL-18 raises the expression of APC-associated receptors and ligands on NK cells that are engaged in immune effector function and destroy target cells, such as CD80, CD86, HLA-DR, HLA-DQ, ICOS, and CD25[13].
 
b. Enhancement of Cytotoxicity
NK cells are stimulated by IL-18, which increases their cytotoxicity toward cancerous or infected cells. The combination of IFN-alpha and IL-18 effectively induces the expression of IFN-gamma in both primary, nonactivated NK cells and the NK-92 cell line. Additionally, ex vivo expanded NK cells stimulated with IL-18/IL-15/IL-12 display high levels of IFN-γ production, potent cytotoxicity, and sustained IFN-γ production even after stimulation is removed[14].
 
c. Synergy with IL-2, IL-12[13], IL-15, IFN-α

→ Croyez GMP ® IL-18 (Interleukin-18), Human
→ Brochure


 

  • IL-21

a. Development and Survival
The growth and survival of NK cells are influenced by IL-21. In the bone marrow, it encourages the maturation and differentiation of NK cell precursors, which helps to produce functional NK cells. Additionally, mature NK cells are maintained and supported by IL-21 in peripheral tissues[15].
 
b. Enhancement of Activation and Cytotoxicity
IL-21, a recently identified cytokine, induces the maturation of NK cells into a specialized phenotype with heightened effector function, characterized by changes in cell size, granularity, receptor expression, and cytotoxic capacity, such as IL-10 and IFN-γ production. In vivo treatment with IL-21 enhances NK cell-mediated anti-tumor immunity, suggesting its role in promoting the terminal differentiation of NK cells and bolstering their cytotoxicity and immune surveillance against tumors[16].
 
c. Cytokine-Induced Memory-Like (ML or CIML) NK cells Formation
Both in humans and mice, it has been shown that IL-21 is crucial for the formation of memory-like NK cells, and in people with latent tuberculosis infection, the growth of these cells is linked to a protective immune response[17].
 
d. Synergy with IL-15, IL-18

→ Croyez GMP ® IL-21 (Interleukin-21), Human


 

  • Type I IFN family

a. Enhancement of Activation and Cytotoxicity
Natural killer (NK) cells are activated by type I interferons (IFNs), which increases their anticancer properties. Lack of IFNAR1 or IFN expression, as well as downstream type I IFN pathway members such TYK2 or STAT1, causes NK cells to have lower cytotoxicity and poor tumor surveillance. Treatment with IL-12 or IL-15 can partially restore NK cell activity, but even enhanced activity is insufficient for tumor rejection, emphasizing the crucial role of IFNs and STAT1 in promoting NK cell cytotoxicity and innate antitumor immunity[18].
 
b. Cytokine-Induced Memory-Like (ML or CIML) NK cells Formation
Antigen-dependent or antigen-independent mechanisms can lead to immunological memory development in NK cells and the development of memory NK cells. This process of producing memory NK cells is greatly aided by type I IFNs. According to the study, type I IFNs help memory NK cells that are dependent on antigens to differentiate. They promote the production of Zbtb32, a transcription factor that helps these NK cells proliferate and perform their protective role. Memory NK cells may be manipulated to specifically target tumor cells in cancer treatments[19].
 
c. Modulation of Phenotype
NK cell-mediated killing of antigen-activated cytotoxic T lymphocytes (CTLs) is inhibited by type I IFNs through altering the expression of NK cell receptor ligands and inhibitory molecules on CTLs. The production of molecules like TRAIL on NK cells can also be induced by type I IFNs, which increases the cytotoxic activity of these cells against tumor cells[19].
 

  • TGF-β

a. Suppression[20] of Activation and Cytotoxicity
NK cells are immunosuppressed by TGF-β via a number of methods. The T-bet-IFN pathway is inhibited, cell metabolism is changed, and activating receptors and chemokine receptors are down-regulated. Furthermore, TGF-β stimulates the transformation of NK cells into less cytotoxic ILC1s, perhaps through the action of lncRNAs. 
 
b. Impact on Development
TGF-β affects the development and subset formation of NK cells. It inhibits the differentiation of CD16+ NK cells and promotes the conversion of a minor fraction of CD56brightCD16+ cells into CD56brightCD16? cells. The findings suggest that TGF-β plays a role in influencing the genesis of NK cell subsets in human peripheral blood[21].
 
c. Tumor Immune Evasion
TGF-β hampers NK cell function, allowing tumor cells to evade immune surveillance. Tumors exploit this by creating a TGF-β-rich microenvironment, suppressing NK cell and immune cell activity, thereby facilitating immune evasion. In the study, manipulating TGF-β signaling to enhance mTOR activity in NK cells shows promise in enhancing immune-mediated tumor suppression and limiting metastases[22].
 
 
In conclusion, the complex interactions between cytokines and NK cells suggest intriguing approaches for advancing NK cell therapy. Increased NK cell proliferation, activation, cytotoxicity, and cytokine production can be achieved by adjusting the amounts and combinations of cytokines. This will increase the efficiency of the NK cells in destroying tumor cells and triggering immunological responses. The area of immunotherapy can be advanced and patient outcomes can be improved in the fight against cancer and other diseases with further study and clinical trials aimed at utilizing the potency of cytokines in NK cell therapy.
Keep Me Updated
 
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  2. Lehmann C, Zeis M, Uharek L. Activation of natural killer cells with interleukin 2 (IL-2) and IL-12 increases perforin binding and subsequent lysis of tumour cells. Br J Haematol. 2001 Sep;114(3):660-5.
  3. Cui F, Qu D, Sun R, Zhang M, Nan K. NK cell-produced IFN-γ regulates cell growth and apoptosis of colorectal cancer by regulating IL-15. Exp Ther Med. 2020 Feb;19(2):1400-1406.
  4. Lehmann D, Spanholtz J, Sturtzel C, Tordoir M, Schlechta B, Groenewegen D, Hofer E. IL-12 directs further maturation of ex vivo differentiated NK cells with improved therapeutic potential. PLoS One. 2014 Jan 31;9(1):e87131.
  5. Parihar R, Dierksheide J, Hu Y, Carson WE. IL-12 enhances the natural killer cell cytokine response to Ab-coated tumor cells. J Clin Invest. 2002 Oct;110(7):983-92.
  6. Romee R, Schneider SE, Leong JW, Chase JM, Keppel CR, Sullivan RP, Cooper MA, Fehniger TA. Cytokine activation induces human memory-like NK cells. Blood. 2012 Dec 6;120(24):4751-60.
  7. Huntington ND, Puthalakath H, Gunn P, Naik E, Michalak EM, Smyth MJ, Tabarias H, Degli-Esposti MA, Dewson G, Willis SN, Motoyama N, Huang DC, Nutt SL, Tarlinton DM, Strasser A. Interleukin 15-mediated survival of natural killer cells is determined by interactions among Bim, Noxa and Mcl-1. Nat Immunol. 2007 Aug;8(8):856-63.
  8. Carson WE, Giri JG, Lindemann MJ, Linett ML, Ahdieh M, Paxton R, Anderson D, Eisenmann J, Grabstein K, Caligiuri MA. Interleukin (IL) 15 is a novel cytokine that activates human natural killer cells via components of the IL-2 receptor. J Exp Med. 1994 Oct 1;180(4):1395-403.
  9. Zhang M, Wen B, Anton OM, Yao Z, Dubois S, Ju W, Sato N, DiLillo DJ, Bamford RN, Ravetch JV, Waldmann TA. IL-15 enhanced antibody-dependent cellular cytotoxicity mediated by NK cells and macrophages. Proc Natl Acad Sci U S A. 2018 Nov 13;115(46):E10915-E10924.
  10. Gang M, Wong P, Berrien-Elliott MM, Fehniger TA. Memory-like natural killer cells for cancer immunotherapy. Semin Hematol. 2020 Oct;57(4):185-193.
  11. Lusty E, Poznanski SM, Kwofie K, Mandur TS, Lee DA, Richards CD, Ashkar AA. IL-18/IL-15/IL-12 synergy induces elevated and prolonged IFN-γ production by ex vivo expanded NK cells which is not due to enhanced STAT4 activation. Mol Immunol. 2017 Aug;88:138-147.
  12. Lee J, Lee SH, Shin N, Jeong M, Kim MS, Kim MJ, Yoon SR, Chung JW, Kim TD, Choi I. Tumor necrosis factor-alpha enhances IL-15-induced natural killer cell differentiation. Biochem Biophys Res Commun. 2009 Sep 4;386(4):718-23.
  13. Senju H, Kumagai A, Nakamura Y, Yamaguchi H, Nakatomi K, Fukami S, Shiraishi K, Harada Y, Nakamura M, Okamura H, Tanaka Y, Mukae H. Effect of IL-18 on the Expansion and Phenotype of Human Natural Killer Cells: Application to Cancer Immunotherapy. Int J Biol Sci. 2018 Mar 9;14(3):331-340.
  14. Matikainen S, Paananen A, Miettinen M, Kurimoto M, Timonen T, Julkunen I, Sareneva T. IFN-alpha and IL-18 synergistically enhance IFN-gamma production in human NK cells: differential regulation of Stat4 activation and IFN-gamma gene expression by IFN-alpha and IL-12. Eur J Immunol. 2001 Jul;31(7):2236-45.
    Lusty E, Poznanski SM, Kwofie K, Mandur TS, Lee DA, Richards CD, Ashkar AA. IL-18/IL-15/IL-12 synergy induces elevated and prolonged IFN-γ production by ex vivo expanded NK cells which is not due to enhanced STAT4 activation. Mol Immunol. 2017 Aug;88:138-147.
  15. Parrish-Novak J, Dillon SR, Nelson A, Hammond A, Sprecher C, Gross JA, Johnston J, Madden K, Xu W, West J, Schrader S, Burkhead S, Heipel M, Brandt C, Kuijper JL, Kramer J, Conklin D, Presnell SR, Berry J, Shiota F, Bort S, Hambly K, Mudri S, Clegg C, Moore M, Grant FJ, Lofton-Day C, Gilbert T, Rayond F, Ching A, Yao L, Smith D, Webster P, Whitmore T, Maurer M, Kaushansky K, Holly RD, Foster D. Interleukin 21 and its receptor are involved in NK cell expansion and regulation of lymphocyte function. Nature. 2000 Nov 2;408(6808):57-63.
  16. Brady J, Hayakawa Y, Smyth MJ, Nutt SL. IL-21 induces the functional maturation of murine NK cells. J Immunol. 2004 Feb 15;172(4):2048-58. doi: 10.4049/jimmunol.172.4.2048. Erratum in: J Immunol. 2006 Mar 15;176(6):3840.
  17. Venkatasubramanian S, Cheekatla S, Paidipally P, Tripathi D, Welch E, Tvinnereim AR, Nurieva R, Vankayalapati R. IL-21-dependent expansion of memory-like NK cells enhances protective immune responses against Mycobacterium tuberculosis. Mucosal Immunol. 2017 Jul;10(4):1031-1042.
  18. Lee CK, Rao DT, Gertner R, Gimeno R, Frey AB, Levy DE. Distinct requirements for IFNs and STAT1 in NK cell function. J Immunol. 2000 Oct 1;165(7):3571-7.
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  22. Regis S, Dondero A, Caliendo F, Bottino C, Castriconi R. NK Cell Function Regulation by TGF-β-Induced Epigenetic Mechanisms. Front Immunol. 2020 Feb 25;11:311.
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