Limits...
Spontaneous tumour regression in keratoacanthomas is driven by Wnt/retinoic acid signalling cross-talk.

Zito G, Saotome I, Liu Z, Ferro EG, Sun TY, Nguyen DX, Bilguvar K, Ko CJ, Greco V - Nat Commun (2014)

Bottom Line: A fundamental goal in cancer biology is to identify the cells and signalling pathways that are keys to induce tumour regression.Furthermore, we demonstrate that developmental programs utilized for skin hair follicle regeneration, such as Wnt, are hijacked to sustain tumour growth and that the retinoic acid (RA) signalling pathway promotes tumour regression by inhibiting Wnt signalling.Finally, we find that RA signalling can induce regression of malignant tumours that do not normally spontaneously regress, such as squamous cell carcinomas.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, Yale Stem Cell Center, Yale Cancer Center, Yale School of Medicine, New Haven, Connecticut 06510, USA.

ABSTRACT
A fundamental goal in cancer biology is to identify the cells and signalling pathways that are keys to induce tumour regression. Here we use a spontaneously self-regressing tumour, cutaneous keratoacanthoma (KAs), to identify physiological mechanisms that drive tumour regression. By using a mouse model system that recapitulates the behaviour of human KAs, we show that self-regressing tumours shift their balance to a differentiation programme during regression. Furthermore, we demonstrate that developmental programs utilized for skin hair follicle regeneration, such as Wnt, are hijacked to sustain tumour growth and that the retinoic acid (RA) signalling pathway promotes tumour regression by inhibiting Wnt signalling. Finally, we find that RA signalling can induce regression of malignant tumours that do not normally spontaneously regress, such as squamous cell carcinomas. These findings provide new insights into the physiological mechanisms of tumour regression and suggest therapeutic strategies to induce tumour regression.

Show MeSH

Related in: MedlinePlus

RA downregulates Wnt and promote KA tumour regression.(a) RNA sequencing heatmap showing a subset of genes differentially expressed between tumour in growth (0 weeks post DMBA) and early regression (1 week post DMBA). The colour coded values are mean centred log10(CPM+1). CPM (count per million): number of reads mapped to a gene for each million reads mapped to any genes. (b) qRT–PCR analysis of RA and Wnt signalling pathway on 0, 1, 3 and 6 weeks post-DMBA treatment. All the values are normalized to the 6-week post-DMBA time point. Data are represented as mean±s.d. (n=3 for each time point) (**<0.01, *<0.05, obtained by unpaired t-test statistical analysis). (c) Schematic representation that represents RA/Wnt cross-talk in regressing KAs. (d) Immunohistochemistry analysis for nuclear β-catenin+ cells in RA and mock-injected KAs (scale bar, 50 μm). (e) qRT–PCR analysis on RA and mock-injected KA tumours at 2 days and 4 weeks after last injection (n=5). (f) qRT–PCR analysis of proliferation and differentiation genes in RA and vehicle-injected KA tumours. RA versus mock-injected tumours comparison has been conducted by using the ΔΔCT method and normalized to glyceraldehyde-3-phosphate dehydrogenase transcript. Data are represented as mean±s.d. (n=5). Dotted line represents the normalized expression level of each gene analysed for the experiment (*<0.05, **<0.01, ****<0.001, obtained by unpaired t-test statistical analysis). (g) Hematoxylin and eosin staining performed on RA and mock-injected tumours at 4 weeks post injection (scale bar, 200 μm for the pictures on the left and 50 μm for the pictures on the right). Dotted lines delineate the tumour/stroma interface.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3974217&req=5

f6: RA downregulates Wnt and promote KA tumour regression.(a) RNA sequencing heatmap showing a subset of genes differentially expressed between tumour in growth (0 weeks post DMBA) and early regression (1 week post DMBA). The colour coded values are mean centred log10(CPM+1). CPM (count per million): number of reads mapped to a gene for each million reads mapped to any genes. (b) qRT–PCR analysis of RA and Wnt signalling pathway on 0, 1, 3 and 6 weeks post-DMBA treatment. All the values are normalized to the 6-week post-DMBA time point. Data are represented as mean±s.d. (n=3 for each time point) (**<0.01, *<0.05, obtained by unpaired t-test statistical analysis). (c) Schematic representation that represents RA/Wnt cross-talk in regressing KAs. (d) Immunohistochemistry analysis for nuclear β-catenin+ cells in RA and mock-injected KAs (scale bar, 50 μm). (e) qRT–PCR analysis on RA and mock-injected KA tumours at 2 days and 4 weeks after last injection (n=5). (f) qRT–PCR analysis of proliferation and differentiation genes in RA and vehicle-injected KA tumours. RA versus mock-injected tumours comparison has been conducted by using the ΔΔCT method and normalized to glyceraldehyde-3-phosphate dehydrogenase transcript. Data are represented as mean±s.d. (n=5). Dotted line represents the normalized expression level of each gene analysed for the experiment (*<0.05, **<0.01, ****<0.001, obtained by unpaired t-test statistical analysis). (g) Hematoxylin and eosin staining performed on RA and mock-injected tumours at 4 weeks post injection (scale bar, 200 μm for the pictures on the left and 50 μm for the pictures on the right). Dotted lines delineate the tumour/stroma interface.

Mentions: Wnt downregulation was observed in advanced stages of tumour regression at 6 weeks post-DMBA treatment. Thus, we asked which signalling pathways could act initially and be responsible for Wnt downregulation. In order to address this question, we used an unbiased approach and performed RNA sequencing to compare the transcriptional profile of epithelial cells of KA tumours at 0 and 1 week post-DMBA treatment. We found that a total number of 107 genes were differentially regulated between the time points analysed (Supplementary Table 1). Among others, we found that the RA signalling pathway stood out as RA-related transcripts Crabp1, Fam5C (Brinp3) and Cyp26b1 were upregulated at 1 week post DMBA (Fig. 6a in red). During mouse development and hair follicle regeneration, the RA pathway has been shown to inhibit Wnt pathway1738. qRT–PCR of RA-related transcripts both in the epithelial population only as well as in whole tumour (Supplementary Fig. 6a) shows that RA activation is an early event during KA tumour regression. If RA acts during the regression phase by downregulating Wnt, then RA upregulation ought to occur prior to Wnt downregulation during KA regression. To test this hypothesis, we first treated wild-type keratinocytes with RA in vitro. Consistent with our model, we found that RA-treated keratinocytes induced not only upregulation of Wnt inhibitors Sfrp2 and Sfrp4 but also promoted downregulation of Wnt target genes by qRT–PCR (Supplementary Fig. 6b). This finding establishes a link between RA upregulation and Wnt downregulation in skin epithelial cells.


Spontaneous tumour regression in keratoacanthomas is driven by Wnt/retinoic acid signalling cross-talk.

Zito G, Saotome I, Liu Z, Ferro EG, Sun TY, Nguyen DX, Bilguvar K, Ko CJ, Greco V - Nat Commun (2014)

RA downregulates Wnt and promote KA tumour regression.(a) RNA sequencing heatmap showing a subset of genes differentially expressed between tumour in growth (0 weeks post DMBA) and early regression (1 week post DMBA). The colour coded values are mean centred log10(CPM+1). CPM (count per million): number of reads mapped to a gene for each million reads mapped to any genes. (b) qRT–PCR analysis of RA and Wnt signalling pathway on 0, 1, 3 and 6 weeks post-DMBA treatment. All the values are normalized to the 6-week post-DMBA time point. Data are represented as mean±s.d. (n=3 for each time point) (**<0.01, *<0.05, obtained by unpaired t-test statistical analysis). (c) Schematic representation that represents RA/Wnt cross-talk in regressing KAs. (d) Immunohistochemistry analysis for nuclear β-catenin+ cells in RA and mock-injected KAs (scale bar, 50 μm). (e) qRT–PCR analysis on RA and mock-injected KA tumours at 2 days and 4 weeks after last injection (n=5). (f) qRT–PCR analysis of proliferation and differentiation genes in RA and vehicle-injected KA tumours. RA versus mock-injected tumours comparison has been conducted by using the ΔΔCT method and normalized to glyceraldehyde-3-phosphate dehydrogenase transcript. Data are represented as mean±s.d. (n=5). Dotted line represents the normalized expression level of each gene analysed for the experiment (*<0.05, **<0.01, ****<0.001, obtained by unpaired t-test statistical analysis). (g) Hematoxylin and eosin staining performed on RA and mock-injected tumours at 4 weeks post injection (scale bar, 200 μm for the pictures on the left and 50 μm for the pictures on the right). Dotted lines delineate the tumour/stroma interface.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3974217&req=5

f6: RA downregulates Wnt and promote KA tumour regression.(a) RNA sequencing heatmap showing a subset of genes differentially expressed between tumour in growth (0 weeks post DMBA) and early regression (1 week post DMBA). The colour coded values are mean centred log10(CPM+1). CPM (count per million): number of reads mapped to a gene for each million reads mapped to any genes. (b) qRT–PCR analysis of RA and Wnt signalling pathway on 0, 1, 3 and 6 weeks post-DMBA treatment. All the values are normalized to the 6-week post-DMBA time point. Data are represented as mean±s.d. (n=3 for each time point) (**<0.01, *<0.05, obtained by unpaired t-test statistical analysis). (c) Schematic representation that represents RA/Wnt cross-talk in regressing KAs. (d) Immunohistochemistry analysis for nuclear β-catenin+ cells in RA and mock-injected KAs (scale bar, 50 μm). (e) qRT–PCR analysis on RA and mock-injected KA tumours at 2 days and 4 weeks after last injection (n=5). (f) qRT–PCR analysis of proliferation and differentiation genes in RA and vehicle-injected KA tumours. RA versus mock-injected tumours comparison has been conducted by using the ΔΔCT method and normalized to glyceraldehyde-3-phosphate dehydrogenase transcript. Data are represented as mean±s.d. (n=5). Dotted line represents the normalized expression level of each gene analysed for the experiment (*<0.05, **<0.01, ****<0.001, obtained by unpaired t-test statistical analysis). (g) Hematoxylin and eosin staining performed on RA and mock-injected tumours at 4 weeks post injection (scale bar, 200 μm for the pictures on the left and 50 μm for the pictures on the right). Dotted lines delineate the tumour/stroma interface.
Mentions: Wnt downregulation was observed in advanced stages of tumour regression at 6 weeks post-DMBA treatment. Thus, we asked which signalling pathways could act initially and be responsible for Wnt downregulation. In order to address this question, we used an unbiased approach and performed RNA sequencing to compare the transcriptional profile of epithelial cells of KA tumours at 0 and 1 week post-DMBA treatment. We found that a total number of 107 genes were differentially regulated between the time points analysed (Supplementary Table 1). Among others, we found that the RA signalling pathway stood out as RA-related transcripts Crabp1, Fam5C (Brinp3) and Cyp26b1 were upregulated at 1 week post DMBA (Fig. 6a in red). During mouse development and hair follicle regeneration, the RA pathway has been shown to inhibit Wnt pathway1738. qRT–PCR of RA-related transcripts both in the epithelial population only as well as in whole tumour (Supplementary Fig. 6a) shows that RA activation is an early event during KA tumour regression. If RA acts during the regression phase by downregulating Wnt, then RA upregulation ought to occur prior to Wnt downregulation during KA regression. To test this hypothesis, we first treated wild-type keratinocytes with RA in vitro. Consistent with our model, we found that RA-treated keratinocytes induced not only upregulation of Wnt inhibitors Sfrp2 and Sfrp4 but also promoted downregulation of Wnt target genes by qRT–PCR (Supplementary Fig. 6b). This finding establishes a link between RA upregulation and Wnt downregulation in skin epithelial cells.

Bottom Line: A fundamental goal in cancer biology is to identify the cells and signalling pathways that are keys to induce tumour regression.Furthermore, we demonstrate that developmental programs utilized for skin hair follicle regeneration, such as Wnt, are hijacked to sustain tumour growth and that the retinoic acid (RA) signalling pathway promotes tumour regression by inhibiting Wnt signalling.Finally, we find that RA signalling can induce regression of malignant tumours that do not normally spontaneously regress, such as squamous cell carcinomas.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, Yale Stem Cell Center, Yale Cancer Center, Yale School of Medicine, New Haven, Connecticut 06510, USA.

ABSTRACT
A fundamental goal in cancer biology is to identify the cells and signalling pathways that are keys to induce tumour regression. Here we use a spontaneously self-regressing tumour, cutaneous keratoacanthoma (KAs), to identify physiological mechanisms that drive tumour regression. By using a mouse model system that recapitulates the behaviour of human KAs, we show that self-regressing tumours shift their balance to a differentiation programme during regression. Furthermore, we demonstrate that developmental programs utilized for skin hair follicle regeneration, such as Wnt, are hijacked to sustain tumour growth and that the retinoic acid (RA) signalling pathway promotes tumour regression by inhibiting Wnt signalling. Finally, we find that RA signalling can induce regression of malignant tumours that do not normally spontaneously regress, such as squamous cell carcinomas. These findings provide new insights into the physiological mechanisms of tumour regression and suggest therapeutic strategies to induce tumour regression.

Show MeSH
Related in: MedlinePlus