Chester Lai

Repeated low-dose skin exposure is an effective sensitizing stimulus, a factor to be taken into account in predicting sensitization risk

Background  Contact sensitization by ingredients in personal products is an important clinical problem. It is not clear how sensitization is induced by the generally low concentrations at which they occur but it might be the result of repeated exposure.

Variants of the melanocortin-1 receptor: do they matter clinically?

The melanocortin 1 receptor (MC1R) gene encodes for a seven‐pass transmembrane receptor primarily expressed on melanocytes and melanoma cells. Single nucleotide polymorphisms (SNPs, also termed variants) in MC1R frequently cause red hair, fair skin and are associated with melanoma and keratinocyte‐derived skin cancer development. Activation of wild‐type (WT) MC1R in skin assists cutaneous photoprotection whereas reduced MC1R signalling, seen with MC1R variants, impairs ultraviolet radiation (UVR)‐protective responses. As ancestral humans migrated out of Africa, the evolutionary advantage of MC1R variants may have related to improved cutaneous vitamin D synthesis and higher birthweight reported with certain MC1R variants. Reduced photoprotection secondary to MC1R dysfunction involves pigmentary and non‐pigmentary mechanisms (reduced DNA repair, effects on cell proliferation and possibly immunological parameters), leading to clonal expansion of mutated cells within skin and subsequent carcinogenesis. Recent investigations suggest an association between MC1R genotype and vitiligo, with preliminary evidence that a MC1R agonist, [Nle4‐D‐Phe7]‐alpha‐MSH, in combination with UVB, assists repigmentation. Future development of compounds to correct defective MC1R responses secondary to MC1R variants could result in photoprotective benefits for fair‐skinned individuals and reduce their skin cancer risk.

OX40+ Regulatory T Cells in Cutaneous Squamous Cell Carcinoma Suppress Effector T-Cell Responses and Associate with Metastatic Potential.

Purpose: Cutaneous squamous cell carcinoma (cSCC) is the most common human cancer with metastatic potential. Despite T cells accumulating around cSCCs, these tumors continue to grow and persist. To investigate reasons for failure of T cells to mount a protective response in cSCC, we focused on regulatory T cells (Tregs) as this suppressive population is well represented among the infiltrating lymphocytes. Experimental Design: Flow cytometry was conducted on cSCC lymphocytes and in vitro functional assays were performed using sorted tumoral T cells. Lymphocyte subsets in primary cSCCs were quantified immunohistochemically. Results: FOXP3+ Tregs were more frequent in cSCCs than in peripheral blood (P < 0.0001, n = 86 tumors). Tumoral Tregs suppressed proliferation of tumoral effector CD4+ (P = 0.005, n = 10 tumors) and CD8+ T cells (P = 0.043, n = 9 tumors) and inhibited IFNγ secretion by tumoral effector T cells (P = 0.0186, n = 11 tumors). The costimulatory molecule OX40 was expressed predominantly on tumoral Tregs (P < 0.0001, n = 15 tumors) and triggering OX40 with an agonist anti-OX40 antibody overcame the suppression exerted by Tregs, leading to increased tumoral effector CD4+ lymphocyte proliferation (P = 0.0098, n = 10 tumors). Tregs and OX40+ lymphocytes were more abundant in primary cSCCs that metastasized than in primary cSCCs that had not metastasized (n = 48 and n = 49 tumors, respectively). Conclusions: Tregs in cSCCs suppress effector T-cell responses and are associated with subsequent metastasis, suggesting a key role for Tregs in cSCC development and progression. OX40 agonism reversed the suppressive effects of Tregs in vitro, suggesting that targeting OX40 could benefit the subset of cSCC patients at high risk of metastasis. Clin Cancer Res; 22(16); 4236–48. ©2016 AACR.

Characteristics of immunosuppressive regulatory T cells in cutaneous squamous cell carcinomas and role in metastasis.

BACKGROUND Non-melanoma skin cancer is the most common cancer worldwide, and cutaneous squamous cell carcinomas (SCCs) account for substantial morbidity and mortality because of their potential for metastasis. SCCs are surrounded by an immune cell infiltrate containing regulatory T cells (Tregs). The aim of this study was to characterise Tregs in SCCs and investigate whether increased Treg numbers in primary skin SCCs are associated with subsequent metastasis. METHODS Lymphocytes were extracted from freshly excised skin SCC tumours and corresponding peripheral blood and normal skin. Flow cytometry was used for T-cell analysis and cell sorting. Tritiated thymidine based lymphocyte proliferation assays and interferon γ (IFNγ) ELISPOT assays were used to assess peritumoral lymphocyte function in vitro. Immunohistochemistry was performed on primary cutaneous SCC sections from tumours that subsequently metastasised and from those that did not after 5-year follow-up. FINDINGS Increased frequencies of CD3+CD4+CD25hiCD127loFOXP3+ Tregs were found in SCCs (21·5% of CD4+ immune infiltrate, n=60 tumours) compared with corresponding peripheral blood (5·4%) and normal skin (7·6%). SCC Tregs expressed significantly higher levels of the co-stimulatory molecules OX40 (37·2% of FOXP3+ cell population, n=10 tumours) and 4-1BB (12·6%, n=9) than peritumoral non-regulatory T cells and Tregs from peripheral blood and normal skin (p=0·0005). The inhibitory receptor CTLA4 and the transcription factor Helios were expressed at high levels in peritumoral Tregs. SCC Tregs significantly suppressed phytohaemagglutinin-stimulated peritumoral CD4+ T-cell proliferation (p=0·005, n=10), peritumoral CD8+ T-cell proliferation (p=0·015, n=9), and IFNγ secretion by CD4+ effector T cells (p=0·026, n=10). Increased in-vitro proliferation of phytohaemagglutinin-stimulated peritumoral CD4+ T cells was shown after the addition of anti-OX40 antibodies (p=0·0078, n=9 tumours) and anti-4-1BB antibodies (p=0·0039, n=9). Immunohistochemistry showed fewer CD8+ T cells in SCCs that metastasised (n=29) than in non-metastatic SCCs (n=26) (28·5% of immune infiltrate vs 44·6%%, p<0·0001) and more FOX3+ Tregs (28·5% of immune infiltrate vs 49·3%, p<0·0001). INTERPRETATION Our study shows that immunosuppressive Tregs are present in the immune infiltrate of cutaneous SCCs, and contribute to ineffective anti-tumour immune responses, thereby permitting SCC development and promoting metastasis. FUNDING Wellcome Trust, National Institute for Health Research.

Epithelial damage and tissue γδ T cells promote a unique tumor-protective IgE response

IgE is an ancient and conserved immunoglobulin isotype with potent immunological function. Nevertheless, the regulation of IgE responses remains an enigma, and evidence of a role for IgE in host defense is limited. Here we report that topical exposure to a common environmental DNA-damaging xenobiotic initiated stress surveillance by γδTCR+ intraepithelial lymphocytes that resulted in class switching to IgE in B cells and the accumulation of autoreactive IgE. High-throughput antibody sequencing revealed that γδ T cells shaped the IgE repertoire by supporting specific variable-diversity-joining (VDJ) rearrangements with unique characteristics of the complementarity-determining region CDRH3. This endogenous IgE response, via the IgE receptor FcεRI, provided protection against epithelial carcinogenesis, and expression of the gene encoding FcεRI in human squamous-cell carcinoma correlated with good disease prognosis. These data indicate a joint role for immunosurveillance by T cells and by B cells in epithelial tissues and suggest that IgE is part of the host defense against epithelial damage and tumor development.Evidence of protective and homeostatic roles for IgE is relatively limited. Strid and colleagues demonstrate that γδ T cells induce switching to IgE that provides protection against experimentally induced epithelial cancer.

Melanoma in situ affecting the penis of a naturist.

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Subclonal Evolution of Cancer-Related Gene Mutations in p53 Immunopositive Patches in Human Skin

Normal sun-exposed skin contains numerous epidermal patches that stain positive for p53 protein (p53 immunopositive patches, PIPs), which are considered potential early precursors of skin cancer. Although the TP53 gene is mutated in many PIPs, it is unclear whether PIPs contain any other cancer-related mutations. Here we report that PIPs, predominantly <3,000 p53 immunopositive cells in size, within normal chronically exposed skin contain mutations in multiple genes that are mutated in cutaneous squamous cell cancers. These mutations in the PIPs were not detected within the non-PIP epidermis of corresponding normal chronically exposed skin. Although some of these genetic alterations are clonal in the PIPs, many of the mutations are subclonal within these lesions. Similar mutations are seen in later precancers (actinic keratoses and Bowens disease). Our results demonstrate that PIPs in chronically exposed skin contain multiple mutations in cancer-related genes. In addition, the results indicate that the clonal evolution of mutations that are seen within later precancerous lesions and in established malignancy can also occur in PIPs within normal human skin.

In vitro human T cell responses to diphencyprone

The experimental contact sensitizer Diphencyprone (DPCP) is used both for topical immunotherapy and investigation of the human immune system. Analysis of mechanisms underlying altered immune responses requires in vitro systems. We have developed a method for detecting human T cell responses to DPCP in vitro. Twenty subjects were sensitized with DPCP (30?g/cm2) and 4 weeks later, were challenged with doses from 0.48 to 3.125?g. Responses at 48h showed all 20 were allergic. Peripheral blood mononuclear cells were cultured with a range of concentrations of DPCP for 6 days and proliferation quantified (3H thymidine uptake). The subjects were exposed repeatedly to DPCP (topical immunotherapy for warts or alopecia areata) or as repeated patch test challenges. Eight subjects gave positive lymphocyte proliferation responses (LPR), maximal responses being elicited with 16 or 32?M DPCP. Positive LPR became detectable from 20 weeks after sensitization and progressively more people become responsive over time to 40 weeks. The different time courses for manifestation of allergic reactivity in skin and blood probably reflects absence of memory T cells in blood due to their recruitment to skin as resident memory T cells. This assay can now be developed for analysis of components of the immune response.

High incidence of skin cancer in the Channel Islands.

Background.  Previous studies looking at rates of malignant melanoma (MM) and nonmelanoma skin cancer (NMSC) in the UK have documented one of the highest rates in the southwest of England; however, the incidence of these tumours in Guernsey and Jersey, two of the Channel Islands, has not previously been reported.

Nanoscale dysregulation of collagen structure-function disrupts mechano-homeostasis and mediates pulmonary fibrosis

Matrix stiffening with downstream activation of mechanosensitive pathways is strongly implicated in progressive fibrosis; however, pathologic changes in extracellular matrix (ECM) that initiate mechano-homeostasis dysregulation are not defined in human disease. By integrated multiscale biomechanical and biological analyses of idiopathic pulmonary fibrosis lung tissue, we identify that increased tissue stiffness is a function of dysregulated post-translational collagen cross-linking rather than any collagen concentration increase whilst at the nanometre-scale collagen fibrils are structurally and functionally abnormal with increased stiffness, reduced swelling ratio, and reduced diameter. In ex vivo and animal models of lung fibrosis, dual inhibition of lysyl oxidase-like (LOXL) 2 and LOXL3 was sufficient to normalise collagen fibrillogenesis, reduce tissue stiffness, and improve lung function in vivo. Thus, in human fibrosis, altered collagen architecture is a key determinant of abnormal ECM structure-function, and inhibition of pyridinoline cross-linking can maintain mechano-homeostasis to limit the self-sustaining effects of ECM on progressive fibrosis.