RepSox

RepSox is a potent and selective inhibitor of the TGFβR-1 with IC50 of 23 nM [1].

TGFβR-1 belongs to the TGFβ receptor family, which is a serine/threonine kinase receptor and bind ligand TGFβ. It gets involved in tumor transformation, cell differentiation and proliferation.

RepSox was found to replace the function of Sox2 to induce reprogramming of iPS cells, via the mechanism of inhibiting TGFβR-1 [1]. MEF cells were treated with RepSox for 3 days, and then it was removed at the time of transduction with Oct4, Klf4, and cMyc. The result showed the TGFβR-1 -associated expression of L-Myc was increased by 5-fold after RepSox treatment [1]. A partially reprogrammed cell line (OKMS 6) was treated with RepSox and performed global gene expression analysis at 10, 24, and 48 hours following the initiation of treatment. The result showed the Id1, Id2 and Id3 gene, repressed by TGFβR-1 signaling, were released from that repression and the expression level was increased. Additionally, other genes responsive for TGFβR-1 signaling were inhibited. It suggested RepSox was able to inhibit TGFβR-1 and suppress the TGFβR-1 downstream signaling [1].

In mouse model, iPS cells reprogrammed by RepSox were labeled with Tomato-red protein and then they were injected mouse blastocysts to produce mosaic embryo. The result showed that both embryo and adult mouse were with significant contribution from RepSox induced iPS cells. It suggested that the inhibitory activity of RepSox for TGFβR-1 was active in vivo [1].

Reference:
[1] Ichida J K et al. , A small molecule inhibitor of Tgaf-β signaling replaces Sox2 in reprogramming by inducing Nanog. Cell Stem Cell, 2009, 5(5): 491-503.

RepSox slows decay of CD34+ acute myeloid leukemia cells and decreases T cell immunoglobulin mucin-3 expression.[Pubmed:24855276]

Stem Cells Transl Med. 2014 Jul;3(7):836-48.

Despite initial response to therapy, most acute myeloid leukemia (AML) patients relapse. To eliminate relapse-causing leukemic stem/progenitor cells (LPCs), patient-specific immune therapies may be required. In vitro cellular engineering may require increasing the "stemness" or immunogenicity of tumor cells and activating or restoring cancer-impaired immune-effector and antigen-presenting cells. Leukapheresis samples provide the cells needed to engineer therapies: LPCs to be targeted, normal hematopoietic stem cells to be spared, and cancer-impaired immune cells to be repaired and activated. This study sought to advance development of LPC-targeted therapies by exploring nongenetic ways to slow the decay and to increase the immunogenicity of primary CD34(+) AML cells. CD34(+) AML cells generally displayed more colony-forming and aldehyde dehydrogenase activity than CD34(-) AML cells. Along with exposure to bone marrow stromal cells and low (1%-5%) oxygen, culture with RepSox (a reprogramming tool and inhibitor of transforming growth factor-beta receptor 1) consistently slowed decline of CD34(+) AML and myelodysplastic syndrome (MDS) cells. RepSox-treated AML cells displayed higher CD34, CXCL12, and MYC mRNA levels than dimethyl sulfoxide-treated controls. RepSox also accelerated loss of T cell immunoglobulin mucin-3 (Tim-3), an immune checkpoint receptor that impairs antitumor immunity, from the surface of AML and MDS cells. Our results suggest RepSox may reduce Tim-3 expression by inhibiting transforming growth factor-beta signaling and slow decay of CD34(+) AML cells by increasing CXCL12 and MYC, two factors that inhibit AML cell differentiation. By prolonging survival of CD34(+) AML cells and reducing Tim-3, RepSox may promote in vitro immune cell activation and advance development of LPC-targeted therapies.

RepSox improves viability and regulates gene expression in rhesus monkey-pig interspecies cloned embryos.[Pubmed:28247195]

Biotechnol Lett. 2017 May;39(5):775-783.

OBJECTIVE: To investigate the effect of the small molecule, RepSox, on the expression of developmentally important genes and the pre-implantation development of rhesus monkey-pig interspecies somatic cell nuclear transfer (iSCNT) embryos. RESULTS: Rhesus monkey cells expressing the monomeric red fluorescent protein 1 which have a normal (42) chromosome complement, were used as donor cells to generate iSCNT embryos. RepSox increased the expression levels of the pluripotency-related genes, Oct4 and Nanog (p < 0.05), but not of Sox2 compared with untreated embryos at the 2-4-cell stage. Expression of the anti-apoptotic gene, Bcl2, and the pro-apoptotic gene Bax was also affected at the 2-4-cell stage. RepSox treatment also increased the immunostaining intensity of Oct4 at the blastocyst stage (p < 0.05). Although the blastocyst developmental rate was higher in the group treated with 25 microM RepSox for 24 h than in the untreated control group (2.4 vs. 1.2%, p > 0.05), this was not significant. CONCLUSION: RepSox can improve the developmental potential of rhesus monkey-pig iSCNT embryos by regulating the expression of pluripotency-related genes.

A small-molecule inhibitor of tgf-Beta signaling replaces sox2 in reprogramming by inducing nanog.[Pubmed:19818703]

Cell Stem Cell. 2009 Nov 6;5(5):491-503.

The combined activity of three transcription factors can reprogram adult cells into induced pluripotent stem cells (iPSCs). However, the transgenic methods used for delivering reprogramming factors have raised concerns regarding the future utility of the resulting stem cells. These uncertainties could be overcome if each transgenic factor were replaced with a small molecule that either directly activated its expression from the somatic genome or in some way compensated for its activity. To this end, we have used high-content chemical screening to identify small molecules that can replace Sox2 in reprogramming. We show that one of these molecules functions in reprogramming by inhibiting Tgf-beta signaling in a stable and trapped intermediate cell type that forms during the process. We find that this inhibition promotes the completion of reprogramming through induction of the transcription factor Nanog.

Identification of 1,5-naphthyridine derivatives as a novel series of potent and selective TGF-beta type I receptor inhibitors.[Pubmed:15317461]

J Med Chem. 2004 Aug 26;47(18):4494-506.

Optimization of the screening hit 1 led to the identification of novel 1,5-naphthyridine aminothiazole and pyrazole derivatives, which are potent and selective inhibitors of the transforming growth factor-beta type I receptor, ALK5. Compounds 15 and 19, which inhibited ALK5 autophosphorylation with IC50 = 6 and 4 nM, respectively, showed potent activities in both binding and cellular assays and exhibited selectivity over p38 mitogen-activated protein kinase. The X-ray crystal structure of 19 in complex with human ALK5 is described, confirming the binding mode proposed from docking studies.