Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2009 Nov;129(11):2676-85.
doi: 10.1038/jid.2009.151. Epub 2009 Jun 11.

Imiquimod enhances IFN-gamma production and effector function of T cells infiltrating human squamous cell carcinomas of the skin

Susan J Huang  1 , Dirkjan Hijnen, George F Murphy, Thomas S Kupper, Adam W Calarese, Ilse G Mollet, Carl F Schanbacher, Danielle M Miller, Chrysalyne D Schmults, Rachael A Clark
Affiliations

Imiquimod enhances IFN-gamma production and effector function of T cells infiltrating human squamous cell carcinomas of the skin

Susan J Huang et al. J Invest Dermatol. 2009 Nov.

Abstract

Squamous cell carcinomas (SCCs) are sun-induced skin cancers that are particularly numerous and aggressive in patients taking T-cell immunosuppressant medications. Imiquimod is a topical immune response modifier and Toll-like receptor 7 (TLR7) agonist that induces the immunological destruction of SCC and other skin cancers. TLR7 activation by imiquimod has pleiotropic effects on innate immune cells, but its effects on T cells remain largely uncharacterized. Because tumor destruction and formation of immunological memory are ultimately T-cell-mediated effects, we studied the effects of imiquimod therapy on effector T cells infiltrating human SCC. SCC treated with imiquimod before excision contained dense T-cell infiltrates associated with tumor cell apoptosis and histological evidence of tumor regression. Effector T cells from treated SCC produced more IFN-gamma, granzyme, and perforin and less IL-10 and transforming growth factor-beta (TGF-beta) than T cells from untreated tumors. Treatment of normal human skin with imiquimod induced activation of resident T cells and reduced IL-10 production but had no effect on IFN-gamma, perforin, or granzyme, suggesting that these latter effects arise from the recruitment of distinct populations of T cells into tumors. Thus, imiquimod stimulates tumor destruction by recruiting cutaneous effector T cells from blood and by inhibiting tonic anti-inflammatory signals within the tumor.

PubMed Disclaimer

Conflict of interest statement

CONFLICT OF INTEREST

The authors state no conflict of interest.

Figures

Figure 1
Figure 1. Imiquimod treated SCC are heavily infiltrated by CD8 T cells that are associated with tumor regression and apoptosis of tumor cells
(a) Cryosections of SCC treated with imiquimod were stained for CD8 (green) and Ki-67 (to identify proliferating tumor cells). Treated SCC had three clear histologic zones: (1) residual tumor, (2) a dense CD8 T cell infiltrate and (3) a deeper fibrotic zone of tumor regression. (b) A higher power view is shown of the interface between the zones of T cell infiltration and tumor regression. Residual tumor (arrow) is surrounded by a dense T cell infiltrate (T) beneath which is a zone of fibrotic regression (R). (c) An additional imiquimod treated tumor stained with Hoechst nuclear dye is shown. Residual tumor (arrow) can be identified by the large atypical keratinocyte nuclei. A zone of dense T cell infiltrate (T) and an area of fibrotic regression (R) are also evident. (d) CD8 T cells infiltrates are associated with tumor cell apoptosis. Hoescht nuclear stain delineates a tumor nodule (arrow) surrounded by T cells (T). TUNEL staining demonstrates widespread tumor cell apoptosis in areas of dense T cell infiltrates. Three additional treated tumors showed similar results.
Figure 2
Figure 2. Production of IFN-γ, perforin and granzyme is markedly enhanced in T cells from SCC treated with imiquimod
IFN-γ is produced by the majority of both (a) CD4+ and (b) CD8+ T cells in SCC treated with imiquimod, compared with lower production in untreated tumors. IL-17 production was low in both treated and untreated tumors. (c) CD4 and CD8 T cells from SCC had increased co-expression of perforin after imiquimod therapy; CD4 T cells also showed enhanced granzyme production. Study of additional treated tumors confirmed enhanced production of perforin by both (d) CD4 and (e) CD8 T cells and increased production of granzyme by (d) CD4 T cells.
Figure 3
Figure 3. CD8 T cells do not proliferate locally within imiquimod treated tumors
(a) Co-staining of cryosections of imiquimod treated SCC with CD8 and Ki-67 demonstrated proliferation of tumor cells but few CD8 T cells. (b) Higher power view of a second tumor with similar findings. (c) Quantification of CD8 T cell proliferation in three treated tumors.
Figure 4
Figure 4. Effector T cells from imiquimod treated SCC produce less IL-10 and TGF-β
(a) Non-regulatory T cells in untreated SCC are a significant source of IL-10. IL-10 is also produced by tumor FOXP3+ Treg (Treg). (b,c) IL-10 production by non-regulatory T cells is reduced in tumors treated with imiquimod. (d) Summarized data from treated and untreated tumors. (e) TGF-β production by non-regulatory T cells is also reduced after imiquimod therapy.
Figure 5
Figure 5. Human SCC tumor cells also produce IL-10
(a) Immunostaining of untreated SCC demonstrates that tumor cells, identified by their large, atypical nuclei on Hoechst stain (lower panels, blue), synthesize IL-10. The left panel (control) shows staining with secondary and tertiary antibodies alone and the right panel also includes the primary anti-IL-10 antibody. (b) The SCC cell line SCC13 produces IL-10. SCC13 tumor cell lines were analyzed for IL-10 production by intracellular cytokine analysis.
Figure 6
Figure 6. Treatment of skin effector T cells in vitro with imiquimod enhances activation and reduces IL-10 production but has no effect on IL-17 and IFN-γ
(a) T cells isolated from human skin treated for 1 week with imiquimod expressed increased CD69 and decreased CD25. Imiquimod treatment of purified skin T cells removed first from the skin microenvironment had no effect (not shown). (b) IL-10 is produced by both CD4 and CD8 non-regulatory T cells from normal human skin. (c) Non-regulatory T cells isolated from imiquimod treated skin produced less IL-10 but (d) production of IL-17 and IFN-γ was unaffected. For all parts of this figure, only non-regulatory T cells are shown; Treg were removed by gating out FOXP3+ T cells prior to analysis.
Figure 7
Figure 7. The effects of imiquimod on tumor associated T cells: consequences on T cell recruitment vs. local modulation of T cell function
A summary of the current studies and our prior work (Clark et al., 2008) is shown. (a) Untreated SCC lack vascular expression of E-selectin and are populated by non-cutaneous central memory T cells, 50% of which are FOXP3+ Treg. (b) Imiquimod treatment of SCC induces vascular E-selectin and the recruitment of tumor-specific CLA+ skin homing T cells. These cells dilute out Treg resident in the tumor and their activation within the tumor leads to production of IFN-γ, perforin, granzyme, and to tumor cell destruction. (c) Imiquimod also acts locally on tumor T cells, indirectly inducing the production of IL-6 by non-regulatory effector T cells (Teff), likely rendering them resistant to suppression. Imiquimod also reduces Teff production of IL-10 and TGF-β, thereby reducing tonic inhibitory signals within the tumor. Imiquimod also acts on to regulatory T cells (Treg) to reduce their ability to suppress via cytokine production (IL-10, TGF-β) and contact suppression (CD39, CD73).
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Ambach A, Bonnekoh B, Nguyen M, Schon MP, Gollnick H. Imiquimod, a Toll-like receptor-7 agonist, induces perforin in cytotoxic T lymphocytes in vitro. Mol Immunol. 2004;40:1307–1314. - PubMed
    1. Berg D, Otley CC. Skin cancer in organ transplant recipients: Epidemiology, pathogenesis, and management. Journal of the American Academy of Dermatology. 2002;47:1–17. - PubMed
    1. Brown VL, Atkins CL, Ghali L, Cerio R, Harwood CA, Proby CM. Safety and efficacy of 5% imiquimod cream for the treatment of skin dysplasia in high-risk renal transplant recipients: randomized, double-blind, placebo-controlled trial. Archives of dermatology. 2005;141:985–993. - PubMed
    1. Clark RA, Chong BF, Mirchandani N, Yamanaka K, Murphy GF, Dowgiert RK, et al. A novel method for the isolation of skin resident T cells from normal and diseased human skin. The Journal of investigative dermatology. 2006;126:1059–1070. - PubMed
    1. Clark RA, Huang SJ, Murphy GF, Mollet IG, Hijnen D, Muthukuru M, et al. Human squamous cell carcinomas evade the immune response by down-regulation of vascular E-selectin and recruitment of regulatory T cells. The Journal of experimental medicine. 2008;205:2221–2234. - PMC - PubMed

Publication types

MeSH terms