Constitutive STAT3 activation in epidermal keratinocytes enhances cell clonogenicity and favours spontaneous immortalization by opposing differentiation and senescence checkpoints.

STAT3, a pleiotropic transcription factor acting downstream of cytokines and growth factors, is known to enhance proliferation, migration, invasion and aerobic glycolysis in tumors upon aberrant activation. In the murine epidermis, STAT3 is necessary for experimentally induced carcinogenesis. Skin tumorigenesis is conversely enhanced by overexpression in keratinocytes of the constitutively active STAT3C mutant, which also induces robust, psoriasis-like epidermal hyperplasia. We show here that STAT3C expression at physiological levels in knock-in mice leads to mild epidermal hyperplasia and attenuated expression of terminal differentiation markers. Altered differentiation is confirmed in isolated primary epidermal keratinocytes in vitro, correlating with enhanced proliferative and clonogenic potential, attenuated senescence and, strikingly, high-frequency spontaneous immortalization. These results suggest that moderate levels of continuous STAT3 activation, which closely resemble those triggered by chronic inflammation or persistent growth factor stimulation, may establish a preneoplastic state in part by promoting the escape of epidermal progenitor cells from differentiation and senescence checkpoints.

STAT3ß controls inflammatory responses and early tumor onset in skin and colon experimental cancer models.

Chronic inflammation is a well-recognized pathogenic factor in tumor initiation and progression. Mice lacking the pro-oncogenic transcription factor STAT3 were shown to be protected from both colitis-associated and epidermal cancers induced by the AOM/DSS and DMBA/TPA protocols, respectively. However, these murine models did not distinguish between the two STAT3 isoforms, the full-length STAT3α, believed to exert most pro-oncogenic functions attributed to STAT3, and the shorter STAT3ß, often referred to as a dominant-negative, but possessing specific transcriptional activities. Here we assessed the contribution of STAT3ß to inflammation-driven tumorigenesis making use of mice lacking this isoform, but still expressing STAT3α (STAT3(Δß/Δß)). We show that the lack of STAT3ß leads to exacerbated acute responses to both TPA and DSS, thus confirming its anti-inflammatory role. Enhanced inflammation correlates with earlier tumor onset in both the epidermis and the intestine in STAT3(Δß/Δß) mice. In contrast, overall tumor development and final tumor burden were unaffected. These results suggest that STAT3ß, by limiting inflammation during the initial phases of tumorigenesis, contributes to tissue homeostasis and counteracts malignant transformation and initial tumor growth. Accordingly, the balance between the two STAT3 isoforms, likely determined by the complex signaling networks shaping the tumor microenvironment and driving tumor transformation and progression, is apparently crucial to determine the initial tumor transformation rates in inflammation-associated cancers.

STAT1 and STAT3 in tumorigenesis: A matter of balance.

The transcription factors STAT1 and STAT3 appear to play opposite roles in tumorigenesis. While STAT3 promotes cell survival/proliferation, motility and immune tolerance and is considered as an oncogene, STAT1 mostly triggers anti-proliferative and pro-apoptotic responses while enhancing anti-tumor immunity. Despite being activated downstream of common cytokine and growth factor receptors, their activation is reciprocally regulated and perturbation in their balanced expression or phosphorylation levels may re-direct cytokine/growth factor signals from proliferative to apoptotic, or from inflammatory to anti-inflammatory. Here we review the functional canonical and non-canonical effects of STAT1 and STAT3 activation in tumorigenesis and their potential cross-regulation mechanisms.

Ups and downs: the STAT1:STAT3 seesaw of Interferon and gp130 receptor signalling.

Downstream of cytokine or growth factor receptors, STAT3 counteracts inflammation and promotes cell survival/proliferation and immune tolerance while STAT1 inhibits proliferation and favours innate and adaptive immune responses. STAT1 and STAT3 activation are reciprocally regulated and perturbation in their balanced expression or phosphorylation levels may re-direct cytokine/growth factor signals from proliferative to apoptotic, or from inflammatory to anti-inflammatory. Here we review the functional canonical and non-canonical effects of STAT1/3 activation and discuss the hypothesis that perturbation of their expression and/or activation levels may provide novel therapeutic strategies in different clinical settings and particularly in cancer.

In the absence of IGF-1 signaling, IFN-gamma suppresses human malignant T-cell growth.

Several approaches to target insulin-like growth factor-1 (IGF-1) signaling have resulted in the inhibition of the growth of a broad range of tumor cells. Malignant T cells are insensitive to the antiproliferative effects of the interferon-gamma (IFN-gamma)/signal transducer and activator of transcription 1 (STAT1) pathway because of the IGF-1-dependent internalization of the IFN-gammaR2 signaling chain. Here we show that human malignant T cells are also resistant to the growth inhibitory effect of both the IGF-1 receptor-specific inhibitor picropodophyllin (PPP) and retrovirus-mediated gene transfer of a dominant negative IGF-1 receptor. However, blockade of IGF-1 receptor perturbs IFN-gammaR2 internalization and induces its cell surface accumulation in malignant T cells. This allows the reinstatement of the IFN-gamma-induced STAT1 activation, a high expression of proapoptotic molecules, and the suppression of malignant T-cell growth both in vitro and in vivo in a severe combined immunodeficiency (SCID) mouse model. These data indicate that the inhibition of IGF-1 signaling combined with IFN-gamma administration could be a promising approach to suppress the growth of neoplastic T cells resistant to each treatment on its own.

IFNgammaR2 trafficking tunes IFNgamma-STAT1 signaling in T lymphocytes.

Ligand-dependent downregulation of the interferon gamma receptor signaling chain (IFNgammaR2) has always been seen as a key mechanism for shielding T lymphocytes from the antiproliferative effects of the IFNgamma-signal transducer and activator of transcription 1 (STAT1) pathway. Now, however, a ligand-independent mechanism of IFNgammaR2 internalization is emerging as a more general way of limiting IFNgamma-STAT1 signaling in T cells, with insulin-like growth factor-1 (IGF-1) and iron as the main players. Here, we review the array of immunomodulatory effects exerted by these two factors on different cell types involved in the immune response; these effects suggest that an inflammatory environment generates signals that favor IFNgammaR2 cell-surface accumulation and IFNgamma-induced apoptosis in T cells, whereas an anti-inflammatory environment promotes IFNgammaR2 internalization and induces T cell unresponsiveness to IFNgamma signaling.

Iron regulates T-lymphocyte sensitivity to the IFN-gamma/STAT1 signaling pathway in vitro and in vivo.

The refractoriness of T cells to the interferon-gamma (IFN-gamma)/signal transducer and activator of transcription 1 (STAT1) pathway, which shields them from the antiproliferative effect of IFN-gamma, is attributed mainly to down-regulation of the IFN-gammaR2 signaling chain. However, the mechanisms responsible for this down-regulation are unclear. Here we show that iron uptake mediated by the transferrin receptor (TfR) delivers a signal that leads to IFN-gammaR2 internalization and thus plays an essential role in attenuating activation of the IFN-gamma/STAT1 pathway in human T lymphocytes. The effect of iron on IFN-gammaR2 internalization was specific as it did not affect expression of the IFN-gammaR1 binding chain. Deferoxamine (DFO), an iron-chelating agent, up-regulated IFN-gammaR2 surface expression and reinstated IFN-gamma/STAT1 activation in proliferating T lymphocytes. Resistance of malignant T cells to the antiproliferative effect of IFN-gamma in vitro was abrogated by addition of DFO. Conversely, iron inhibited IFN-gamma-induced apoptosis in malignant T cells in serum-free conditions. In combination but not individually, DFO and IFN-gamma strongly inhibited growth of human malignant T cells in an in vivo severe combined immunodeficient (SCID) mouse model. These data provide valuable insights for novel therapeutic approaches aimed at reinstating the IFN-gamma/STAT1 apoptotic signaling pathway in autoreactive or neoplastic T cells by means of iron chelation.

IGF-1 down-regulates IFN-gamma R2 chain surface expression and desensitizes IFN-gamma/STAT-1 signaling in human T lymphocytes.

The ability of insulin-like growth factor-1 (IGF-1) to regulate surface expression of the interferon-gamma receptor 2 (IFN-gamma R2) transducing chain and activation of IFN-gamma-induced signal transducer and activator of transcription-1 (STAT-1) in human T cells was analyzed. We show that, especially in the absence of serum (which contains IGF-1), IGF-1 down-regulated surface expression of the IFN-gamma R2 chain and inhibited both IFN-gamma-dependent STAT-1 activation and apoptosis in T-cell lines ST4, Jurkat, and Molt-4. IFN-gamma R2 down-regulation resulted from its enhanced internalization since IGF-1 completely restored the uptake of anti-IFN-gamma R2 monoclonal antibody (mAb) in serum-deprived T-cell lines. When the interaction between IGF-1 and its receptor was blocked by anti-IGF-1R mAb, enhancement of IFN-gamma R2 surface expression, STAT-1 activation, and reinstatement of IFN-gamma-induced apoptosis were observed. Enhanced expression of IFN-gamma R2 was also observed in phytohemagglutinin (PHA)-activated T lymphoblasts cultured in the presence of anti-IGF-1R mAb, whereas IGF-1 or anti-IGF-1R mAb did not modify the high IFN-gamma R2 expression in B and myeloid cell lines. Both IGF-1 and anti-IGF-1R mAb did not modify the constitutive expression of IFN-gamma R2 mRNA in T cells as well as the high IFN-gamma R1 binding chain surface expression in T, B, and myeloid cells. These data indicate that IGF-1 plays a critical role in the desensitization of IFN-gamma/STAT-1 signaling in T lymphocytes by delivering a signal for IFN-gamma R2 internalization.