Project description:We isolated and selected intestinal adenoma organoids from Apc1638N/+ and Apc1638N/+; Kras mice. After the selection procedure, we maintained the cultures with or without TGF-beta for 18h. RNA was isolated to determine the effect of oncogenic Kras on the gene expression changes. Total RNA obtained from Apc1638N/+; Kras organoids were compared to Apc1638N/+ samples in the absence or presence of 3 ng/ml TGF-beta (18h).
Project description:Intestinal crypts isolated from Apcflox/flox; villin-CreERT mice were treated with Tamoxifen to induce the deletion of Apc. Tamoxifen-treated organoids were selected in the absence of Wnt agonists and then treated with TGF-beta. Total RNA obtained from Tamoxifen-treated, Apc-deleted intestinal organoids in the absence or presence of 3 ng/ml TGF-beta (18h).
Project description:We isolated and selected intestinal adenoma organoids from Apc1638N/+ and Apc1638N/+; Kras mice. After the selection procedure, we maintained the cultures with or without TGF-beta for 18h. RNA was isolated to determine the effect of oncogenic Kras on the gene expression changes.
Project description:Intestinal crypts isolated from Apcflox/flox; villin-CreERT mice were treated with Tamoxifen to induce the deletion of Apc. Tamoxifen-treated organoids were selected in the absence of Wnt agonists and then treated with TGF-beta.
Project description:The pro-tumourigenic role of epithelial TGFβ in colorectal cancer (CRC) has been controversial. Here we identify a cohort of aggressive ‘bad acting’ early-stage (T1) disseminating tumours characterised by high cell-intrinsic TGFβ signalling emanating from the epithelium, not stroma. To address its functional significance, we activated TGFβ signalling in the murine intestinal epithelium either alone or in concert with the common tumour suppressive and oncogenic mutations found in CRC, namely Apc and Kras. Consistent with previous studies, we found that activation of TGFβ rapidly induced apoptosis in Apc-mutant intestine and completely killed Apc-mutant organoids. However, in the presence of both Apc and Kras mutation, activation of TGFβ within the epithelium rampantly accelerates tumourigenesis. Importantly the transcriptional signatures derived from these mice overlapped with the “bad acting” T1 human tumours and this signalling could also predict recurrence in stage II CRC. Mechanistically, the activation of intrinsic TGFβ induced the expression of a growth-factor signalling module containing EGFR that synergised with Apc and Kras to drive marked activation of MAPK signalling. Importantly, inhibition of MEK and/or EGFR suppressed the acceleration conferred by TGFβ even in Kras-mutant cells, which are refractory to MEK/EGFR inhibition in the absence of epithelial TGFβ. Together, we identify both a determinant of early dissemination and a potential vulnerability for tumours with these born-to-be-bad traits.
Project description:Small intestinal tissue was collected for RNAseq analysis from mice with short term Apc inactivation or Apc inactivation with Kras mutation, in the presence or absence of Eif4a1 or Eif4a2 inactivaton.
Project description:We utilized bulk RNA sequencing to assess intrinsic differences between wildtype and autophagy-defective (Atg16l1 deleted) AKPS colorectal cancer organoids in both untreated and IFN gamma treated conditions. AKPS mutations consist of loss-of-function in tumor suppressors Apc, Trp53, Smad4, and gain-of-function in oncogenic Kras(G12D).
Project description:A profile of gene expression differences in mouse small intestine with short term induction of Apc inactivation, in the presence and absence of additional mutations in combinations of Kras, Pik3ca, Pten, and Slc7a5.
Project description:Constitutive Wnt activation upon loss of Adenoma polyposis coli (APC) acts as main driver of colorectal cancers (CRC). Targeting Wnt signaling has proven difficult because the pathway is crucial for homeostasis and stem cell renewal. To distinguish oncogenic from physiologic Wnt activity, we have performed transcriptome and proteome profiling in isogenic human colon organoids. Culture in the presence or absence of exogenous ligand allowed us to discriminate receptor-mediated signaling from the effects of CRISPR/Cas9 induced APC loss. We could catalogue two non-overlapping molecular signatures that were stable at distinct levels of stimulation. Newly identified markers for normal stem/progenitor cells and adenomas were validated by immunohistochemistry and flow cytometry. We found that oncogenic Wnt signals are associated with good prognosis in tumors of the consensus molecular subtype 2 (CMS2). In contrast, receptor-mediated signaling was linked to CMS4 tumors and poor prognosis. Together, our data represent a valuable resource for biomarkers that allow more precise stratification of Wnt responses in CRC.
