Massachusetts General Hospital

Self-renewal of human epithelial stem cells versus their commitment to differentiation are closely linked.  Understanding this process is of great potential impact for new therapeutic approaches to human tumors, which are mostly of epithelial origin. Our main interest is the interplay between intra- and inter-cellular signaling pathways involved in control of epithelial tissue homeostasis and tumor development. Our present research is focused on two main topics : (i) intrinsic control mechanisms underlying the balance between epithelial stem cell renewal and differentiation; (ii) role of underlying mesenchymal cells in control of epithelial cell fate determination and tumorigenesis. For a general overview of our research efforts and their impact, see the enclosed interview article

i) The gene regulatory network involved in intrinsic keratinocyte growth/differentiation control.     

 Keratinocytes and skin provide an attractive experimental system to study the connection between growth/differentiation potential of epithelial cells and transformation. Notch signalling is an important form of cell-cell communication with a function that is highly context-dependent. A substantial body of evidence (reviewed in¹) has confirmed and expanded our original findings that this pathway plays a key role in promoting epidermal cell differentiation and suppressing tumor development ^2,3. We showed that the Notch1 gene, which plays the most critical function in keratinocytes, is a direct target of p53 and that down-modulation of Notch1 expression in keratinocyte-derived tumors can be explained by mutation of p53 ⁴ or down-modulation of p53 expression by increased EGFR activation ⁵ or calcineurin inhibition ⁶. Notch activation, in turn, suppresses p63, a p53-related transcription factor with an essential role in maintenance of keratinocyte self renewing populations ⁷. The cross-talk between Notch, p53/EGFR and p63 is at the core of a genetic network with key role in keratinocyte growth/differentiation control and tumor development. Another components of this network with an essential role in epidermal development and differentiation is IRF6, a transcription factor of the IRF (Interferon Responsive Factors) family that we have recently established as Notch target ⁸. Importantly, deep sequencing analysis of oral, skin and lung squamous cell carcinomas (SCCs) by a number of other laboratories has revealed that one or more components of the Notch/p53/p63 network can be inactivated by mutations in human SCCs, confirming their key role in the disease ^9-12. We are currently further exploring regulation and function of this gene network in the skin.

Summary diagram of the Notch-connected network involved in keratinocyte differentiation and tumor suppression.  A set of representative genes with a known or likely role in keratinocyte differentiation and Notch signalling is indicated. For explanation and more complete specification of network elements and connections, see the text above and indicated references.

 ii) The dermal fibroblast gene regulatory network involved in skin field cancerization.

 While great attention has been paid to the role of Notch signalling in the epithelial compartment of the skin, we have started to explore the role of this pathway in the underlying mesenchyme. We have developed a genetic mouse model with loss of Notch signalling in the mesenchymal compartment of the skin, finding that this pathway is essential for cell fate maintenance of overlying hair follicle keratinocytes ¹³. More importantly, by combined analysis of the mouse model and clinically-derived skin samples, we have uncovered a novel essential role of mesenchymal Notch signalling in field cancerization in the skin ¹⁴. Epithelial tumours are usually thought to result from genetic changes in a discrete group of cells, originating from a single initial progenitor, and of changes in immediately surrounding normal cells. The field cancerization process refers instead to pro-tumourigenic changes, in both epithelial cells and surrounding stromal cells, that occur in larger areas, or "fields", of target organs like the skin, oral cavity, lung, prostate and breast. The widespread changes help explaining the frequent multifocality of epithelial tumors and the fact that, after removal, these tumors often recur. Field cancerization has been linked to the presence of genetic pro-tumourigenic changes in apparently normal ‘patches’ of epithelial cells that expand over time. Our recent findings indicate that compromised Notch signalling in the underlying dermal fibroblasts is likely to play an equally important primary role, resulting in tissue alterations (stromal atrophy and inflammation) that precede premalignant and malignant epithelial tumor development ¹⁴. Mechanistically, Notch signaling in dermal fibroblasts of both murine and human origin results in intrinsically increased expression of diffusible growth factors and inflammatory cytokines, cancer-associated matrix components and matrix remodeling enzymes, which are amenable to up-regulation of AP1 levels and activity ¹⁴. The findings are of likely clinical significance, as suppression of Notch/CSL signaling and associated gene expression events occur in stromal fields adjacent to cutaneous premalignant lesions (actinic keratosis), and can be induced by UVA exposure, a major cause of skin chronic and cancer-predisposing alterations ¹⁴. We have established an intensive research effort to further explore the molecular and cellular basis of field cancerization in the skin.

Summary diagram of the Notch-connected network involved in dermal fibroblast activation and skin field cancerization. The genes in the network are partially overlapping with those involved in the keratinocyte network illustrated above. The diagram is based on our recent findings that UVA exposure of human skin / dermal fibroblasts induces down-modulation of Notch expression and activity through specific mechanisms like increased DNA methylation of the Notch2 promoter 14. Notch suppression leads to up-regulated expression of AP1 family members and of AP1 target genes with a role in inflammation, promotion of cancer development and/or aging 14.

 Cited Publications

 Full list of publications 

 Honors and Awards

1982          Grant-in-Aid of Research of the Sigma Xi, The Scientific Research Society, New Haven, CT;

1984          Fellow; The Jane Coffin Childs Memorial Fund, New Haven CT

1987          Hull Cancer Research Award

1988          Swebilius Cancer Research Award

2001          Honorary Masters’ degree, Harvard University

2002-2003  NIH study section member (GMA-1)

2005/2007  Vice Chair/Chair Gordon Conference on Epithelial Differentiation

2010          Lecturing Professor, “Oncology at the Limits”, Cape Town, South Africa

2010           “Irwin Blank Symposium Lecture”, Society of Investigative Dermatology Meeting, Atlanta, Georgia

2011            Elected EMBO membership

2012           American Skin Association Lifetime Achievement Award

2013           European Research Council Panel Member, in “Physiology, Pathophysiology and Endocrynology”.

Present Group Members:

Bach Cuc Nguyen - bach-cuc.nguyen@cbrc2.mgh.harvard.edu

Yang Brooks - ybrooks@partners.org

Jun Dai - jdai@partners.org

Stefania Gastaldi - sgastaldi@partners.org

Seunghee Jo - sjo1@partners.org

Recent Publications:      Full List of Publications

1. Mandinova A, Lefort K, Tommasi di Vignano A, Stonely W, Ostano P, Chiorino G, Iwaki H, Nakanish J, and Dotto GP. The FoxO3a gene is a key negative target of canonical  Notch signaling in the keratinocyte UVB-response, EMBO J. 2008: 27, 1243-54.
2. Kolev V, Mandinova A, Guinea-Viniegra J, Hu B, Lefort K, Lambertini C, Neel V, Dummer R, Wagner EF, Dotto GP. EGFR signaling as a negative regulator of Notch1 gene transcription and function in proliferating keratinocytes and cancer, Nat Cell Biol. 2008: 10-902-11.
   
3. Dotto GP. Notch tumor suppressor function. Oncogene, 2008; 27: 5115-23.
   
4. Dotto GP. Crosstalk of Notch with p53 and p63 in cancer growth control. Nature Reviews Cancer 2009; 9: 587-595
   
5. Mandinova A, Kolev V, V Neel, B Hu, W Stonely, J Lieb, X Wu, C Colli, R Han, M Pazin, P Ostano, R Dummer, JL Brissette, and Dotto GP. An FGFR3/FOXN1 positive loop underlies benign skin keratosis versus squamous cell carcinoma formation. J Clin Invest. 2009: 119 : 3127-3137
   
6. Wu, X., Nguyen, B.C., Dziunycz, P., Chang, S., Brooks, Y., Lefort, K., Hofbauer, G.F.L., and Dotto, G.P. Calcineurin and ATF3: opposite role in keratinocyte cancer development versus senescence. Nature. 2010: 465 : 368-372.

Cutaneous Biology Research Center

Phone: 617-726-4354