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Tle1 (transducin-like enhancer of split 1) is a corepressor that interacts with a variety of DNA-binding transcription factors and has been implicated in many cellular functions; however, physiological studies are limited. Tle1-deficient (Tle1(Δ/Δ)) mice, although grossly normal at birth, exhibit skin defects, lung hypoplasia, severe runting, poor body condition, and early mortality. Tle1(Δ/Δ) mice display a chronic inflammatory phenotype with increased expression of inflammatory cytokines and chemokines in the skin, lung, and intestine and increased circulatory IL-6 and G-CSF, along with a hematopoietic shift toward granulocyte macrophage progenitor and myeloid cells. Tle1(Δ/Δ) macrophages produce increased inflammatory cytokines in response to Toll-like receptor (TLR) agonists and lipopolysaccharides (LPS), and Tle1(Δ/Δ) mice display an enhanced inflammatory response to ear skin 12-O-tetradecanoylphorbol-13-acetate treatment. Loss of Tle1 not only results in increased phosphorylation and activation of proinflammatory NF-κB but also results in decreased Hes1 (hairy and enhancer of split-1), a negative regulator of inflammation in macrophages. Furthermore, Tle1(Δ/Δ) mice exhibit accelerated growth of B6-F10 melanoma xenografts. Our work provides the first in vivo evidence, to our knowledge, that TLE1 is a major counterregulator of inflammation with potential roles in a variety of inflammatory diseases and in cancer progression.

Abstract 1197 TLE1 belongs to the Groucho/TLE family of co-repressors that act as master regulators during development affecting segmentation, neurogenesis, myogenesis, and multiple cell fate decisions. TLE1 modulate several major signaling pathways including Wnt and Notch, and specifically interacts with multiple transcription factors involved in hematopoiesis such TCF/LEF, HES1, RUNX1/AML. TLE1 has also been implicated in Crohn's disease via its interaction with NOD2, a regulator of NFkB. Our laboratory identified TLE1 as a likely AML tumor suppressor gene, commonly deleted in subgroups of AML, and others have shown its role as a tumor suppressor gene in myeloid and other hematopoietic malignancies. To better understand the role of TLE1 in hematopoiesis and leukemogenesis we created a line of Tle1 null mice. Tle1 null mice are born normally, but become progressively growth retarded by 3 days of life, with only 50% survival by 4 weeks as compared to heterozygous and wild type littermates. Abnormalities are observed in several organs systems including the hematopoietic system. We characterized the hematopoietic system in Tle1 knock out mice between two and 12 weeks of age. The bone marrow cellularity in the Tle1 knock out mice is comparable to the wild type mice at all time points examined. However, frequency of granulocyte macrophage progenitors in bone marrow mononuclear cells is significantly higher in the Tle1 knockout bone marrow compared to heterozygous and wild type mice. The proportion and number of myeloid cells as evidenced by Gr1, Mac1 expression are significantly higher in the bone marrow, spleen and blood of these knockout mice. There were significantly lower B-cells (B220+cells) in the Tle1 knockout mice compared to heterozygous and wild type. In colony forming assays there was a trend towards higher number of CFU-GM (7.66 vs 5), p=0.07) and CFU-M (27.16 vs 12.5, p=0.05) colonies from Tle1 null bone marrow as compared to wild type bone marrow. The spleens from four week and 17 months old Tle1 knockout mice had higher frequency of Gr1-negative, Mac1-positive and F4/80 positive macrophages. We also observed a significantly higher production of the inflammatory cytokines IL6 and TNFafrom peritoneal macrophages harvested from Tle1 null mice as compared to those from wild type mice in response to TLR ligand stimulation. To investigate the potential mechanism of this inhibitory effect of TLE1 on inflammation we demonstrated that TLE1 expression is able to block the nuclear translocation of NFkB in THP1 cells in response to LPS-K12 (p In summary this work demonstrates that the lack of Tle1 expression biases hematopoiesis towards myeloid differentiation, a finding of potential relevance given the inactivation of TLE1 seen in subsets of myeloid malignancies. We further show that inactivation of Tle1 leads to an increase in macrophages primed to release increased inflammatory cytokines. This is notable given the recent observation that TLE1 may modulate the effects of NOD2 in the pathogenesis of Crohn's disease. These Tle1 null mice will allow the investigation of the potential role of TLE1 as a modulator of a variety of other inflammatory diseases. Disclosures: No relevant conflicts of interest to declare.

Transducin-like enhancer of split-1 (TLE1) is a novel tumor suppressor gene that is deleted or methylated in subgroups of acute myeloid leukemia (AML) and other hematological malignancies. TLE1 belongs to a family of Gro/TLE proteins that function as corepressors that can bind and potentially inhibit numerous transcription factors known to be involved in development and carcinogenesis including myc, beta-catenin, NF-kB, and TGF-B. In Drosophila Groucho is recognized as a master regulatory gene in development. Our studies and those from other groups have shown that over-expression of TLE1 in leukemia cells slows cell cycle progression, colony formation and tumor growth in xenografts, while silencing results in increased cell proliferation. To further evaluate the role of TLE1 in development and oncogenesis we have knocked out TLE1 in mice. These mice are viable and do not spontaneously develop malignancies in the absence of additional cooperating oncogenes. In Drosophila the TLE homologue Groucho binds and represses Drosophila Myc expression of target genes. N-Myc is important for hematopoietic stem cell proliferation, survival and differentiation and is over-expressed in most AML samples. We postulated that TLE 1 might be an important regulator of N-Myc activity in leukemia and the loss of TLE may accelerate the leukemia development in cooperation with N-Myc. We demonstrate here that the loss of Tle1 cooperates with N-Myc to dramatically increase the serial replating and proliferation of hematopoietic progenitor cells from fetal livers. Mice transplanted with N-Myc transduced hematopoietic cells from the fetal liver of Tle1 knockout mice developed leukemia with a much shorter latency as compared with fetal liver cells from wild type mice. We are currently investigating the mechanisms of this cooperation. This is the first demonstration that Tle1 is a potent modulator of Myc induced tumorigenesis and may have implications in a broader spectrum of malignancies. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3113.