BRAFV600E drives dedifferentiation in small intestinal and colonic organoids and cooperates with mutant p53 and Apc loss in transformation

Abstract

BRAFV600E confers poor prognosis and is associated with a distinct subtype of colorectal cancer (CRC). Little is known, however, about the genetic events driving the initiation and progression of BRAFV600E mutant CRCs. Recent genetic analyses of CRCs indicate that BRAFV600E often coexists with alterations in the WNT- and p53 pathways, but their cooperation remains ill-defined. Therefore, we systematically compared small and large intestinal organoids from mice harboring conditional BraffloxV600E, Trp53LSL-R172H, and/or Apcflox/flox alleles. Using these isogenic models, we observe tissue-specific differences toward sudden BRAFV600E expression, which can be attributed to different ERK-pathway ground states in small and large intestinal crypts. BRAFV600E alone causes transient proliferation and suppresses epithelial organization, followed by organoid disintegration. Moreover, BRAFV600E induces a fetal-like dedifferentiation transcriptional program in colonic organoids, which resembles human BRAFV600E-driven CRC. Co-expression of p53R172H delays organoid disintegration, confers anchorage-independent growth, and induces invasive properties. Interestingly, p53R172H cooperates with BRAFV600E to modulate the abundance of transcripts linked to carcinogenesis, in particular within colonic organoids. Remarkably, WNT-pathway activation by Apc deletion fully protects organoids against BRAFV600E-induced disintegration and confers growth/niche factor independence. Still, Apc-deficient BRAFV600E-mutant organoids remain sensitive toward the MEK inhibitor trametinib, albeit p53R172H confers partial resistance against this clinically relevant compound. In summary, our systematic comparison of the response of small and large intestinal organoids to oncogenic alterations suggests colonic organoids to be better suited to model the human situation. In addition, our work on BRAF-, p53-, and WNT-pathway mutations provides new insights into their cooperation and for the design of targeted therapies.

Gender Differences in Urothelial Bladder Cancer. Effects of Natural Killer Lymphocyte Immunity

“…This suggests that AR sig-naling may help induce new breast cancers, but "dedifferentiated" primitive cancers may downregulate AR expression because they are driven by hormone-independent mechanisms. Similar phenomena have been observed in other cancers that become independent of the signaling molecules that characterize cells from the same tissue of origin [38][39][40].…”

Oncogenic BRAF, unrestrained by TGFβ-receptor signalling, drives right-sided colonic tumorigenesis

“…Aside from region-specific differences in basal proliferation rates and crypt dynamics, the signals and niche cell-types that determine cell-lineage allocation—i.e., the interconversion between adult and foetal-like stem cell populations and TA-progenitor cell states/fates—vary profoundly lengthways along the intestinal tract and up the vertical crypt-axis 56 . Accordingly, BRAF V600E -mutant small intestinal and colonic organoids differ profoundly in basal MAPK-pathway activity, with small intestinal organoids able to tolerate higher pathway activity upon BRAF V600E expression 57 . Notably, though, the transcriptomic profiles of BRAF V600E colonic organoids more closely resemble human BRAF V600E -mutant CRCs 57 .…”

“…Accordingly, BRAF V600E -mutant small intestinal and colonic organoids differ profoundly in basal MAPK-pathway activity, with small intestinal organoids able to tolerate higher pathway activity upon BRAF V600E expression 57 . Notably, though, the transcriptomic profiles of BRAF V600E colonic organoids more closely resemble human BRAF V600E -mutant CRCs 57 . Second, the BA model expresses physiological levels of a knock-in BRAF V600E mutant driven by the endogenous Braf promoter, whereas the transgenic model relies on the overexpression of a human BRAF V600K transgene, which attains expression of MAPK-target genes akin to levels in more advanced dysplastic tumours.…”

“…In strong support of our findings, while this paper was under review, Reischmann et al . reported that the expression of BRAF V600E in colonic organoids elicits a foetal-like dedifferentiation program, associated with a transcriptional signature enriched for Hippo-pathway targets that resembles human BRAF V600E -driven CRC 57 . Interestingly, activation of the ERK and YAP pathways has also been detected in an inflammation-driven model of serrated intestinal tumorigenesis arising through atypical-PKC deficiency, independently of Braf (and Kras ) mutations 62 , signifying that YAP dysregulation may commonly underlie distinct serrated neoplasia pathways independently of the driver mutation.…”

Subversion of Niche-Signalling Pathways in Colorectal Cancer: What Makes and Breaks the Intestinal Stem Cell

“…It further remains to be seen whether the foetal-like, YAP/TAZ-dependent, Lgr5 − regenerative state plays a role in the development of colonic tumours lacking overt Wnt-pathway mutations. In support of this notion, BRAF V600E -driven colonic organoids exhibit a foetal-like dedifferentiation program enriched for Hippo-pathway targets, which recapitulates the transcriptional profiles of human BRAF V600E -driven CRCs [ 162 ], although in vivo validation is currently lacking.…”