Image-based detection and targeting of therapy resistance in pancreatic adenocarcinoma.

Nature.

2016 Jun 06

Fox RG, Lytle NK, Jaquish DV, Park FD, Ito T, Bajaj J, Koechlein CS, Zimdahl B, Yano M, Kopp JL, Kritzik M, Sicklick JK, Sander M, Grandgenett PM, Hollingsworth MA, Shibata S, Pizzo D, Valasek MA, Sasik R, Scadeng M, Okano H, Kim Y, MacLeod AR, Lowy AM, R
PMID: 27281208 | DOI: 10.1038/nature17988.

Pancreatic intraepithelial neoplasia is a pre-malignant lesion that can progress to pancreatic ductal adenocarcinoma, a highly lethal malignancy marked by its late stage at clinical presentation and profound drug resistance. The genomic alterations that commonly occur in pancreatic cancer include activation of KRAS2 and inactivation of p53 and SMAD4 (refs 2, 3, 4). So far, however, it has been challenging to target these pathways therapeutically; thus the search for other key mediators of pancreatic cancer growth remains an important endeavour. Here we show that the stem cell determinant Musashi (Msi) is a critical element of pancreatic cancer progression both in genetic models and in patient-derived xenografts. Specifically, we developed Msi reporter mice that allowed image-based tracking of stem cell signals within cancers, revealing that Msi expression rises as pancreatic intraepithelial neoplasia progresses to adenocarcinoma, and that Msi-expressing cells are key drivers of pancreatic cancer: they preferentially harbour the capacity to propagate adenocarcinoma, are enriched in circulating tumour cells, and are markedly drug resistant. This population could be effectively targeted by deletion of either Msi1 or Msi2, which led to a striking defect in the progression of pancreatic intraepithelial neoplasia to adenocarcinoma and an improvement in overall survival. Msi inhibition also blocked the growth of primary patient-derived tumours, suggesting that this signal is required for human disease. To define the translational potential of this work we developed antisense oligonucleotides against Msi; these showed reliable tumour penetration, uptake and target inhibition, and effectively blocked pancreatic cancer growth. Collectively, these studies highlight Msi reporters as a unique tool to identify therapy resistance, and define Msi signalling as a central regulator of pancreatic cancer.

Expression of NR5A2, NUP153, HNF4A, USP15 and FNDC3B is consistent with their use as novel biomarkers for bovine mammary stem/progenitor cells

Journal of molecular histology

2021 Jan 05

Choudhary, RK;Capuco, AV;
PMID: 33400051 | DOI: 10.1007/s10735-020-09948-8

Mammary stem cells (MaSC) are essential for growth and maintenance of mammary epithelium. Previous studies have utilized morphological characteristics or retention of bromodeoxyuridine (BrdU) label to identify MaSC and progenitor cells, these approaches may not be feasible or may not identify all resident stem cells. Alternatively, these special cells may be identified by assessing protein and mRNA expression of appropriate markers. The focus of this study was to assess the staining patterns and in situ quantification of novel candidate markers for bovine MaSC/progenitor cells. The candidate markers for MaSC/progenitor cells for immunohistochemical analysis were: NR5A2, NUP153, HNF4A, USP15 and FNDC3B and for in situ transcripts quantification were HNF4A and NUP153. We also evaluated protein expression pattern of presumptive MaSC markers known from the literature namely, ALDH1, MSI1 and Notch3. We found that NR5A2, NUP153, HNF4A and USP15-labeled cells represented 2.5-6% of epithelial cells prepubertally and were distributed in a fashion consistent with the location and abundance of MaSC/progenitor cells. A transient increase (10-37%) in expression of these markers was observed at peak lactation. FNDC3B was localized mainly in the nucleus prepubertally and in the cytoplasm of myoepithelial cells and nuclei of a limited number of alveolar cells during lactation. Abundant expression (~ 48%) and luminal localization of ALDH1 precludes its use as a bovine MaSC marker but may include transamplifying progenitor cells. MSI1 staining was consistent with MaSC localization. Onset of lumen formation in mammary ducts of prepubertal gland was associated with Notch 3 expression in the apical surface of luminal cells. RNAscope analysis of HNF4A and NUP153 transcripts in calf mammary gland showed very low copy numbers in a few epithelial cells, supporting the idea that these markers are expressed by fewer cells of epithelial origin. This study suggests that NR5A2, NUP153, HNF4A, USP15 and FNDC3B are likely markers for bovine MaSC/progenitor cells. Quantification of RNA transcripts of HNF4A and NUP153 in bovine MEC as potential MaSC markers are novel. Further studies to correlate protein expression of these markers with their transcripts level using single cell analysis in larger samples in lactating cow at different physiological stages are warranted.

Expression profile of intestinal stem cell markers in colitis-associated carcinogenesis

Scientific Reports

2017 Jul 26

Kim HS, Lee C, Kim WH, Maeng YH, Jang BG.
PMID: 28747693 | DOI: 10.1038/s41598-017-06900-x

The intestinal epithelium has two distinct two stem cell populations, namely, crypt base columnar (CBC) cells and +4 cells. Several specific markers have been identified for each stem cell population. In this study, we examined the expression profiles of these markers in colitis-associated carcinogenesis (CAC) to investigate whether they can be used as biomarkers for the early detection of dysplasia. The expression of intestinal stem cell (ISC) markers was measured by real-time polymerase chain reaction during CAC that was induced by azoxymethane and dextran sodium sulfate treatment. CBC stem cell markers increased continuously with tumor development, whereas a +4 cell expression profile was not present. CBC stem cell population was suppressed in the acute colitis and then expanded to repopulate the crypts during the regeneration period. Notably, RNA in situ hybridization revealed that all dysplasia and cancer samples showed increased expression of CBC stem cell markers in more than one-third of the tumor height, whereas regenerative glands had CBC stem cell markers confined to the lower one-third of the crypt. These results suggest that CBC stem cell markers could be a useful tool for the early detection of colitis-induced tumors.

	Description
sense Example: Hs-LAG3-sense	Standard probes for RNA detection are in antisense. Sense probe is reverse complent to the corresponding antisense probe.
Intron# Example: Mm-Htt-intron2	Probe targets the indicated intron in the target gene, commonly used for pre-mRNA detection
Pool/Pan Example: Hs-CD3-pool (Hs-CD3D, Hs-CD3E, Hs-CD3G)	A mixture of multiple probe sets targeting multiple genes or transcripts
No-XSp Example: Hs-PDGFB-No-XMm	Does not cross detect with the species (Sp)
XSp Example: Rn-Pde9a-XMm	designed to cross detect with the species (Sp)
O# Example: Mm-Islr-O1	Alternative design targeting different regions of the same transcript or isoforms
CDS Example: Hs-SLC31A-CDS	Probe targets the protein-coding sequence only
EnEm	Probe targets exons n and m
En-Em	Probe targets region from exon n to exon m
Retired Nomenclature
tvn Example: Hs-LEPR-tv1	Designed to target transcript variant n
ORF Example: Hs-ACVRL1-ORF	Probe targets open reading frame
UTR Example: Hs-HTT-UTR-C3	Probe targets the untranslated region (non-protein-coding region) only
5UTR Example: Hs-GNRHR-5UTR	Probe targets the 5' untranslated region only
3UTR Example: Rn-Npy1r-3UTR	Probe targets the 3' untranslated region only
Pan Example: Pool	A mixture of multiple probe sets targeting multiple genes or transcripts

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