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
. 2010 Aug 15;70(16):6467-76.
doi: 10.1158/0008-5472.CAN-09-4058. Epub 2010 Aug 3.

Immunologic consequences of signal transducers and activators of transcription 3 activation in human squamous cell carcinoma

Emilia Albesiano  1 , Meghan Davis, Alfred P See, James E Han, Michael Lim, Drew M Pardoll, Young Kim
Affiliations

Immunologic consequences of signal transducers and activators of transcription 3 activation in human squamous cell carcinoma

Emilia Albesiano et al. Cancer Res. .

Abstract

Paracrine cross-talk between tumor cells and immune cells within the tumor microenvironment underlies local mechanisms of immune evasion. Signal transducer and activator of transcription 3 (STAT3), which is constitutively activated in diverse cancer types, is a key regulator of cytokine and chemokine expression in murine tumors, resulting in suppression of both innate and adaptive antitumor immunity. However, the immunologic effects of STAT3 activation in human cancers have not been studied in detail. To investigate how STAT3 activity in human head and neck squamous cell carcinoma (HNSCC) might alter the tumor microenvironment to enable immune escape, we used small interfering RNA and small-molecule inhibitors to suppress STAT3 activity. STAT3 inhibition in multiple primary and established human squamous carcinoma lines resulted in enhanced expression and secretion of both proinflammatory cytokines and chemokines. Although conditioned medium containing supernatants from human HNSCC inhibited lipopolysaccharide-induced dendritic cell activation in vitro, supernatants from STAT3-silenced tumor cells reversed this immune evasion mechanism. Moreover, supernatants from STAT3-silenced tumor cells were able to stimulate the migratory behavior of lymphocytes from human peripheral blood in vitro. These results show the importance of STAT3 activation in regulating the immunomodulatory mediators by human tumors and further validate STAT3 as a promising target for therapeutic intervention.

PubMed Disclaimer

Figures

Figure 1
Figure 1
STAT3 suppression in HNSCC cell lines. qRT-PCR demonstrated down-regulation of STAT3 mRNA by (a) siRNA treatment, as compared to no treatment or controls (scramble siRNA), or by (b) B7 shRNA lentivirus (LV) treatment, as compared to no treatment or control lentivirus. Histograms represent the means of at least three experiments normalized to mRNA expression of GAPDH. Data are presented as fold difference relative to control-untreated cell lines. * p <0.05; ** p <0.01. (c) STAT3 activation in tumor cells, as detected by EMSA. Stat-3/3, Stat3 homodimer; Stat-3/1, Stat3-Stat1 heterodimer, Stat-1/1, Stat1 homodimer. The STAT3 homodimer band in HN29, HN11and Cal27 cell lines decreased after treatment with STAT3 siRNA, compared to control. U937 stimulated with IL-6 represents the positive control (lane 7). Pre-incubation with an anti-STAT3 antibody shifts the STAT3 homodimer band (supershift band; lane 8).
Figure 2
Figure 2
STAT-3 suppression in tumor cells increases expression of proinflammatory mediators. qRT-PCR analysis showed up-regulation of proinflammatory cytokine and chemokines mRNA by siRNA treatment only, as compared to no treatment and to controls in (a) Cal27, (b) HN11, and (c) HN29 cell lines. Histograms represent the means of at least three experiments normalized to GAPDH. Data are presented as fold difference relative to control-untreated cell lines. * p <0.05; ** p <0.01.
Figure 3
Figure 3
Proinflammatory cytokines and chemokines can be detected in STAT3 siRNA treated HNSCC cell lines. ELISAs were performed from treated or control (a) Cal27, (b) HN11, and (c) HN29 cell line culture supernatants. Results are presented as the mean ± s.e.m., n=3, for each ELISA. * p <0.05; ** p <0.01.
Figure 4
Figure 4
Inhibition of DC maturation by tumor derived factors is abrogated upon STAT3 knockdown. Flow cytometry plots demonstrate surface expression of HLA-DR vs. CD86 of CD11c+ gated DCs. Upper row: immature monocyte-derived DCs (middle panel) and LPS-matured DCs (right panel). Left panel showing isotype controls on LPS-matured DCs. Middle row: LPS-matured, DCs cultured in presence of condition medium from untreated (left panel), control (middle panel), and STAT3 siRNA treated Cal27 supernatant (right panel). Lower row: LPS-matured, DCs cultured in presence of condition medium from control lentivirus (LV) vector transduced HN11 supernatant (left panel), STAT3 B7 and B8 shRNA LV vector transduced HN11 supernatant (middle and left panels, respectively).
Figure 5
Figure 5
Supernatant from STAT3 siRNA suppressed tumor cells enhances leukocytes migration. Chemotaxis assay demonstrated enhanced PBMC migration in the presence of CM containing siRNA treated and control Cal27 culture supernatants. PBMCs were incubated in presence of serum free medium and 100% serum as negative and positive controls, respectively. Results are presented as the mean ± s.e.m., n=3, for each ELISA. * p <0.05; ** p <0.01.
Figure 6
Figure 6
Stattic treatment decreases STAT3 activation and induces proinflammatory mediator expression. (a) STAT3 activation in tumor cells, as detected by EMSA. Stat-3/3, Stat3 homodimer; Stat-3/1, Stat3/Stat1 heterodimer, Stat-1/1, Stat1 homodimer. The STAT3 homodimer band in both Cal27 and HN11 cell lines was decreased after treatment with 10μM Stattic for 48h, compared to untreated and to 1-0.1 μM Stattic treatments. U937 stimulated with IL-6 is the positive control (lane 5). Pre-incubation with an anti-STAT3 antibody shifted the STAT3 homodimer band (supershift band; lane 6). (b) qRT-PCR analysis showed up-regulation of proinflammatory cytokine and chemokines mRNA by 10mM Stattic for 48h, compared to untreated, DMSO only (drug diluent), and to 1 μM Stattic treatment. Histograms represent the means of at least three experiments normalized to GAPDH mRNA expression (internal control). Data are presented as fold difference relative to untreated cell lines. * p <0.05.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Yu H, Jove R. The STATs of cancer--new molecular targets come of age. Nat Rev Cancer. 2004;4(2):97–105. - PubMed
    1. Darnell JE. Validating Stat3 in cancer therapy. Nat Med. 2005;11(6):595–6. - PubMed
    1. Nagpal JK, Mishra R, Das BR. Activation of stat-3 as one of the early events in tobacco chewing-mediated oral carcinogenesis. Cancer. 2002;94(9):2393–400. - PubMed
    1. Sen M, Tosca PJ, Zwayer C, et al. Lack of toxicity of a STAT3 decoy oligonucleotide. Cancer Chemother Pharmacol. 2008 - PMC - PubMed
    1. Schust J, Sperl B, Hollis A, Mayer TU, Berg T. Stattic: A small-molecule inhibitor of STAT3 activation and dimerization. Chem Biol. 2006;13(11):1235–42. - PubMed

Publication types

MeSH terms