The patients

The risk of squamous cell carcinoma is dramatically raised in immune compromised patients, such as organ transplant recipients. People with kidney transplants who develop skin lesions form a special focus group in our clinic: they are closely monitored in order to detect possible skin carcinomas at an early stage for adequate and minimized invasive treatment. The dominant risk factor here is apparently the lifelong immunosuppressive treatment to prevent the rejection of the transplant. However, environmental factors, like solar UV exposure and HPV infection, and genetic factors, like a fair skin and polymorphisms in crucial genes, are also of great importance to carcinoma risk in organ transplant recipients.  

Research

Our department investigates the role of genetic and environmental factors, including medication, in the genesis of skin carcinomas, especially in relation to the elevated risk in renal transplant patients. In these studies we collaborate closely with the departments of Nephrology, Immuno-Hematology. Medical Microbiology, Pathology, Clinical Epidemiology, Medical Genetics, Toxico-Genetics and Medical Statistics. Progression of skin carcinoma is studied by a genome-wide analysis of gene expression in different stages of tumor development (comparable to studies in the “Lymphoma” line of research), the growth of tumors in cultured 3D skin models (see research line “Human Skin Models”) and in mouse models. In vitro organotypic models of human actinic keratosis and squamous cell carcinoma may prove excellent experimental vehicles in developing well targeted anti-tumor medication without any restriction and with direct human relevance, thus superseding current animal models.

Figure 1: Shown are Left) Human skin model generated with a SCC biopsy; Middle) Cross section of the skin model  after 4 weeks of culture (invasive phenotype is presents) and Right) Hand of patient with an SCC.

Aim

The ultimate aim of this line of research is to identify the tumor-specific changes in the relevant skin (stem) cells and in signal transduction pathways that are characteristic of and essential  to the development squamous cell carcinoma (SCC) and its precursor, actinic keratosis (AK). This information may enable us to single out crucial molecular targets for clinical interventions specifically affecting the tumor or its early development without collateral damage to healthy tissue.

Studies in Patients and with human cells

In renal transplant recipients (RTRs) we study risk factors for the development of SCCs to identify patients that run an increased risk, and to identify possible pathogenic mechanisms which contribute to SCC development. In the past years we have focused on the role of cutaneous HPV infections, but in vivo studies proved to be hampered by the sparseness of HPV in the non-neoplastic skin in combination with a lack of techniques to identify the infected skin cells.
In vitro studies with human skin cells transfected with HPV gene constructs have revealed widely different effects from different types of HPV (e.g. increased survival or suppression of apoptotic responses upon UV exposure), lending support to the concept of “high-risk” versus “low-risk” types.
SCCs, AKs and normal skin from RTRs have been analyzed fro gen expressions and chromosomal aberrations. A great many genes proved to be differentially expressed between these three groups of samples and these differences were difficult to match unambiguously to any crucial or dominant change in signaling pathway. An activation of the RAS-pathway from AK to SCC appeared to be the most prominent result. Chromosomal aberrations in SCC from our RTRs appear to be less than reported earlier for SCC.from immunocompetent people. This may reflect an (in part) different pathogenesis of SCC in RTRs.
More recently, our attention has been drawn to immune reactions directed at the transplant, ranging from rejection to tolerance, and the strength of the immune challenge, e.g., increased by simultaneous transplantation of kidney and pancreas when compared to transplant of kidney only. The corresponding variations in immune reactions appear to affect the SCC risk.

Studies in mice

In mice we study early oncogenic changes induced by UV irradiation and the role of stem cells in tumor formation and maintenance. The effects of various immunosuppressive treatments (with azathioprine, ciclosporin, tacrolimus, mycophenolate mofetil and rapamycin) on UV carcinogenesis are assessed in a mouse model, and compared to observations on carcinoma risk in RTRs), and with short term effects in human skin models (“human skin equivalents”, HSEs). Surprisingly and in contrast to the increased risk of skin carcinomas among RTRs, we did not find an obligatory acceleration of UV carcinogenesis from treatment with the various immunosuppressive drugs. In contrast to earlier experiments, we found a lack of correspondence between the effect of a immunosuppressive drug on the number of early clones with overexpressed mutant-p53 and ensuing tumor development (i.e. increases in clones did not necessarily translate into enhanced tumor development and vv decreases in clones did not translate into delayed tumor onset). Following the lack of an enhanced UV carcinogenesis by immunosuppressive drugs, we want to investigate the effect of a full organ transplant on UV carcinogenesis.

Figure 2: Shown are mutant-p53 posittive cells present in Left) interfolicular foci of irradiated mice skin and Right) SCC.

Subsequent to our earlier study on the role of quiescent stem cells in the development of carcinomas from chronic low level UV exposure of the skin, we will investigate the role of newly proliferating stem cells in UV carcinogenesis; we will do this in collaboration with prof. Hans Clevers (Hubrecht Lab, Ultrecht, The Netherlands) who – together with the group of prof Tofgard (Karolinska Inst., Stockholm,Sweden) – discovered these (Lgr-5 and -6 positive) stem cells in the hair follicles of the skin.

Figure 3: Shown is a cross sections of an epidermis with CPD accumulation in BrdU-retaining cells/stem cells (arrow).

UV and vitamin D

In collaboration with the department of  Human Genetics we investigate the effect of UV exposure of the skin, and the effect of vitamin D3 on intestinal cancer in a transgenic APC-mutant mouse model. Further studies on the photosynthesis of vitamin D3 in the skin are important because the net production under various conditions (solar spectra) is not adequaltely known (the interplay between the 6 photochemical reactions involved need to be studied in details especially for wavelengths over 310 nm). This information is required to inform people better on adequate exposures for vitamin D synthesis with minimal increase in the risk of skin carcinomas. Vitamin D3 active metabolite (1,25 dihydroxyvitmain D3) also ligates to the vitamin D receptor in skin cells, and is likely to exert an inhibitory effect on the growth of skin carcinomas.

Principal investigators

• F.R. de Gruil
• J.N. Bouwes Bavinck

Skin carcinomas are the most frequent malignancies in white populations and exposure to ultraviolet radiation from the sun appears to be a major etiological factor.  UV radiation damages DNA which causes mutations in genes which can render skin cells malignant. Variations in genetic background, viz. in pigmentation and repair of damaged DNA, determine susceptibility to the formation of skin carcinomas. Certain types of human papilloma virus (HPV) could raise the susceptibility. Aside of this direct carcinogenic impact, UV irradiation affects the immune system, most notably the cellular immunity against skin carcinomas can be suppressed. Such immune modulatory effects are utilized in the dermatological clinic in phototherapy of skin diseases, most specifically psoriasis, but these immunological effects can also cause “sun allergy” of the skin (“polymorphic light eruption”, PLE). Of general benefit to our health is the UV-driven synthesis of vitamin D3 in the skin, which among other can lead to inhibition of tumor growth.