Actinic keratosis is an early in situ squamous cell carcinoma: a proposal for reclassification.

The term actinic keratosis (AK) describes a sun-induced, clinical erythematous lesion covered with scale, but does not provide an understanding of the biology or histopathology of the lesion. Consequently, several classification systems for AK have been suggested, but as yet no consensus has been reached. These systems strive to correlate the pathological and clinical features to better provide physicians with the most accurate information to enable correct decisions to be made regarding treatments, Prognosis and metastatic potential. AK is a clinical description that has a histological diagnosis consistent with squamous cell carcinoma (SCC) in situ. We recommend an AK classification system that describes these lesions as squamous cell carcinomas (SCCs), using the terminology 'early in situ SCC Type AK I', 'early in situ SCC type AK II' and 'in situ SCC Type AK III', there by giving clinicians better guidance for diagnosis and specific treatment recommendations.

Apoptosis pathways as promising targets for skin cancer therapy.

Apoptosis pathways provide efficient safeguard mechanisms against cancer that are mediated via cell-intrinsic responses and immune-mediated extrinsic signals. Intrinsic pro-apoptotic pathways are largely controlled by p53 and Bcl-2 proteins, whereas the extrinsic induction of apoptosis is initiated by death ligands, such as tumour necrosis factor-alpha (TNF-alpha), CD95L/FasL and TNF-related apoptosis-inducing ligand (TRAIL), or by granzyme B. Initiation of these pathways results in the induction of a caspase cascade leading to cell death. The inactivation of pro-apoptotic pathways is elementary for tumourigenesis and may be responsible for therapy resistance. Thus, apoptosis-based strategies represent important tools for the development of effective tumour therapies. The aim of these therapies is to restore p53 activity, downregulate anti-apoptotic Bcl-2 proteins or NF-kappaB activity, and to upregulate extrinsic, death receptor-mediated pathways. The initial results of apoptosis-based strategies are proving promising. Also, topical treatments for actinic keratosis (AK), such as cyclo-oxygenase-2 inhibitors (e.g. diclofenac 3% gel), have been shown to trigger pro-apoptotic pathways. There is hope that pro-apoptotic strategies will lead to pronounced therapeutic success against skin cancer. Importantly, the involvement of the different pro-apoptotic pathways in specific tumour types needs to be unravelled and understood in order to evaluate drug effectiveness, as well as to modify and optimise therapeutic approaches.

The role of apoptosis in therapy and prophylaxis of epithelial tumours by nonsteroidal anti-inflammatory drugs (NSAIDs).

In addition to having anti-inflammatory activities, nonsteroidal anti-inflammatory drugs (NSAIDs) also inhibit neoplastic cell proliferation by inducing apoptosis. Diclofenac is the anti-neoplastic compound in diclofenac 3% gel (Solaraze) used for topical treatment of actinic keratosis (AK). Main target of NSAIDs seems to be the inhibition of cyclo-oxygenase-2 (COX-2), which is overexpressed in several epithelial tumours and catalyses the synthesis of prostaglandins. The precise mechanism of action of diclofenac in cutaneous cells is still unclear, but induction of apoptosis is a key effect of anti-neoplastic drugs, including NSAIDs. In this paper we give an overview of the anti-tumoural activities of NSAIDs with emphasis on induction of apoptosis. Cyclo-oxygenase-2-mediated synthesis of prostaglandin E(2) (PGE(2)) leads to activation of mitogen-activated protein kinase (MAPK), as well as phosphatidylinositol 3-kinase (PI3K)/Akt pathways. Induction of the anti-apoptotic Bcl-2 and Mcl-1, as well as activation of the caspase-8 inhibitor cFLIP have been reported. In addition, altered lipid concentrations in the cytoplasmic membrane may modulate death receptor activities. Downregulation of both the intrinsic mitochondrial and the extrinsic pathways have been reported. Our data demonstrate induced apoptosis and activation of the caspase cascade in three of four cutaneous squamous cell carcinoma (SCC) cell lines, after treatment with diclofenac plus hyaluronic acid and diclofenac alone; one cell line remained nonresponsive. The effects were less pronounced in normal keratinocytes and cytotoxic effects were not seen. Detailed analysis of apoptosis pathways employed by diclofenac in these cells may help to improve therapeutic strategies and to overcome possible mechanisms that are involved in nonresponsiveness.

Low prevalence of p53, p16(INK4a) and Ha-ras tumour-specific mutations in low-graded actinic keratosis.

BACKGROUND: Ultraviolet radiation induces DNA damage and is the major risk factor for the development of non-melanoma skin cancer (NMSC). Different mutation rates of p53, p16(INK4a) and Ha-ras in cutaneous squamous cell carcinoma (SCC) and the earlier stage actinic keratosis (AK) have been reported. OBJECTIVES: To assess the presence of missense mutations in hotspot exons of p53, p16(INK4a) and Ha-ras in low-graded AK. PATIENTS/METHODS: Cryo-biopsies of 75 sun-exposed AK lesions and 75 sun-shielded areas of normal skin from 75 AK patients were analysed to identify mutations in p53 (exons 7 and 8), p16(INK4a) (exon 2) and Ha-ras (exon 1) using polymerase chain reaction (PCR) followed by direct sequencing. As a representative subset of the specimens, ten mutation-negative AK were also micro-dissected in order to exclude the possibility that additional mutations were undetected. RESULTS: Eight missense and one nonsense point mutations were found in the 75 AK lesions examined (12%), of which seven (9%) were tumour-specific (i.e. present in AK lesions only) and two (3%) were p16(INK4a) mutations (i.e. also detected in normal skin). Three of the tumour-specific mutations (42%) were cytosine (C) to thymine (T) transitions at pyrimidine-rich sequences. Tumour-specific mutations were identified in 1% of p16(INK4a) (exon 2), 1% of Ha-ras (exon 1) and at a higher rate of 7% in p53 (exons 7 and 8), including one nonsense mutation. CONCLUSIONS: The evaluation of a large number of AK specimens in this study have found a low gene mutation rate in low-graded AK lesions. p53 mutations rather than p16(INK4a) and/or Ha-ras mutations may be an early event in the development of AK to cutaneous SCC.

Treatment of multiple actinic keratoses with topical diclofenac 3% gel in organ transplant recipients: a series of six cases.

BACKGROUND: Non-melanoma skin cancer (NMSC) represents a significant cause of morbidity in organ transplant patients; the relative risk of squamous cell carcinoma and actinic keratosis (AK) is 100 and 250 times higher, respectively, compared with immunocompetent patients. OBJECTIVES: The aim of this study was to investigate the effects of 3% diclofenac gel on the clearance rates of multiple AKs in organ transplant recipients (OTRs). PATIENTS/METHODS: An open-label study was conducted in six patients (three kidney, one liver and two heart transplant patients) with histories of multiple NMSCs and extensive actinic keratoses (AKs). Patients were treated with diclofenac 3% gel, twice daily for 16 weeks. Complete and partial clearance rates of AKs were assessed after 16 weeks and biopsies were performed 4 weeks post-therapy on clinically typical AKs identified prior to the start of treatment. RESULTS: Three out of six patients showed complete clinical and histological clearance after 16 weeks of treatment with diclofenac 3% gel. Two further patients showed a marked (> or = 75% lesion reduction) improvement in their overall AK lesion count in the treatment area. One patient showed a 30% lesion reduction in the area treated. Local adverse events at the site of application were very mild and included mild erythema and marginal erosion. No systemic side effects were reported. CONCLUSIONS: Results suggest that diclofenac 3% gel may be useful in the treatment and control of multiple AK lesions in OTRs. Further studies are needed to investigate the efficacy and safety of this therapy for the local treatment of AK in greater numbers of immunosuppressed patients.

Management of actinic cheilitis using diclofenac 3% gel: a report of six cases.

BACKGROUND: Actinic cheilitis is a frequent manifestation of actinic dysplasia and requires early therapy to prevent its progression into invasive squamous cell carcinoma (SCC). Several therapies are used, ranging from unspecific lesion-adapted destructive techniques (i.e. laser) to ambitious surgical field-management (vermillionectomy). There is increasing awareness of the effectiveness of field adapted, non destructive therapies, such as photodynamic therapy or 5% imiquimod. Diclofenac 3% gel is used in the treatment of actinic keratosis (AK), but it has not been evaluated for the treatment of actinic cheilitis. OBJECTIVES: This non-blinded, uncontrolled case series study evaluated the effects of diclofenac 3% gel in the treatment of actinic cheilitis. PATIENTS/METHODS: Six patients with histologically verified actinic cheilitis were treated with diclofenac 3% gel, twice daily for 6 weeks. Clinical assessment was performed 2-4 weeks after the end of treatment. RESULTS: Four out of six patients showed clinical clearing of actinic cheilitis 2-4 weeks after the end of treatment. Biopsies were taken from the treated areas at the final visit to verify clinical clearance. Side effects in most of the patients included mild erythema and mild to moderate swelling of the lips. CONCLUSIONS: Topical therapy with diclofenac 3% gel may be an efficient, cosmetically more appealing alternative treatment for actinic cheilitis than currently used destructive therapies. However, future studies and long-term follow-up of patients will be needed to compare its efficacy with established forms of therapy.

Differentiation between actinic keratoses and disseminated superficial actinic porokeratoses with reflectance confocal microscopy.

BACKGROUND: Clinical differentiation between actinic keratosis (AK) and disseminated superficial actinic porokeratosis (DSAP) may pose a significant challenge, and histological evaluation is often also required for diagnosis. Distinct morphological features can be distinguished upon histopathological examination, but the use of non-invasive tools, such as reflectance confocal microscopy (RCM), may be an eligible alternative for confirmation of diagnosis. OBJECTIVES: The aim of this study was to determine the relevant RCM criteria for the identification of disseminated superficial actinic porokeratoses (DSAPs) and to define distinguishing criteria for DSAPs compared with actinic keratosis (AKs). PATIENTS/METHODS: A total of 20 patients with a clinical diagnosis of AK or DSAP were included in this study. All lesions were evaluated by clinical examination, and RCM and one clinically identified lesion was biopsied for histological confirmation. RESULTS: Cellular and nuclear atypia, inflammation, spongiosis, parakeratosis and changes in epidermal architecture were present in both lesion types (i.e. AKs and DSAPs). However, these features were more pronounced in AKs. Whereas AKs exhibited more disseminated parakeratotic changes, parakeratosis was found focally present on the border of DSAP lesions. Most characteristically, a distinct border corresponding to cornoid lamella in RCM can be identified in DSAPs. CONCLUSIONS: Distinguishing features of DSAPs, such as cornoid lamella, sharp demarcation of the lesion and focal keratinocyte atypia are easily identifiable using RCM, and correlate well with histopathology. Whilst RCM has previously been used in the evaluation of AKs, it has not yet been used to investigate DSAPs. The findings in this study suggest the feasibility of non-invasive tools, such as RCM for the differentiation of AKs and DSAPs. However, further studies are warranted to assess the sensitivity and specificity of RCM in the diagnosis of DSAP.

Genetically determined susceptibility to COX-2 inhibitors: a report of exaggerated responders to diclofenac 3% gel in the treatment of actinic keratoses.

Diclofenac 3% gel is an effective treatment for actinic keratoses (AKs) and is reported to be generally well tolerated with only mild local reactions. However, there is a subset of patients that seem to be susceptible to developing severe local reactions following application of diclofenac 3% gel. Although some of these reactions can be explained as being allergic contact dermatitis and/or photoallergic contact dermatitis, others cannot. We report a series of 10 patients who all developed severe local reactions following application of diclofenac 3% gel, despite negative diclofenac patch testing. This raises the question as to whether there is a subset of patients with skin cancer or AK lesions that are highly/more susceptible to local reactions caused by cyclo-oxygenase-2 (COX-2) inhibitors and peroxisome proliferator-activated receptor (PPAR) agonists? We speculate that underlying molecular differences exist in these patients that make the skin more susceptible to COX-2 inhibitors.

Tenascin-C patterns and splice variants in actinic keratosis and cutaneous squamous cell carcinoma.

BACKGROUND: Tenascin-C (Tn-C) is an extracellular matrix protein with multiple functions that is present at low levels in normal tissues, but which is highly present in various tumours. The mRNA expression and protein level of Tn-C including its various isoforms have not been investigated comprehensively so far in cutaneous squamous cell carcinoma (SCC) and the precursor lesion actinic keratosis (AK). OBJECTIVES: To assess the dysregulated expression and splice variants of Tn-C in cutaneous squamous cell dysplasia and carcinoma. METHODS: Biopsies from 66 patients (or representative subsets) that comprised 25 specimens from normal skin, 19 AK and 22 cutaneous SCC were analysed for Tn-C splice variants using splice-specific primers. The amount of Tn-C mRNA was investigated by quantitative real-time reverse transcription-polymerase chain reaction. In addition, the presence of Tn-C protein was analysed in sections of paraffin-embedded tissues using immunohistochemistry. RESULTS: The large Tn-C splice variant was present in only 5% of normal skin samples, in comparison with 63% of AK (P < 0.001) and 88% of SCC (P < 0.001). Tn-C mRNA expression was significantly increased in AK and SCC compared with normal skin (P < 0.001). The corresponding proteins were rarely detected in cells of the vascular epithelial layers and perifollicular layers of some normal skin specimens, and their spatial localization expanded into the papillary dermis of AK. The largest amount and the widest distribution were found in samples of SCC, in which Tn-C was located in the basal cells at the tumour invasion front and additionally in the papillary dermis and reticular dermis. CONCLUSIONS: Tn-C is present in the dermis, its expression is increased during skin cancer development, and the large splice variant is characteristic for AK and SCC, which may prove useful for diagnostic approaches in cutaneous SCC.

E6/E7 expression of human papillomavirus types in cutaneous squamous cell dysplasia and carcinoma in immunosuppressed organ transplant recipients.

BACKGROUND: DNA of cutaneous human papillomavirus (HPV) types is frequently found in nonmelanoma skin cancer, and their E6 and E7 proteins can have transforming properties. OBJECTIVES: To assess the biological activity of HPV types found in tumour tissues we examined HPV E6/E7 RNA expression and the antibody response to E6, E7 and L1 proteins. METHODS: Thirty-one snap-frozen biopsies from six immunosuppressed organ transplant recipients representing seven squamous cell carcinomas (SCCs), one basal cell carcinoma, four actinic keratoses (AKs), seven normal skin and 12 verrucae vulgaris (Vv) were analysed for 24 cutaneous HPV types by an L1 DNA polymerase chain reaction (PCR)-based method. The presence of E6/E7 transcripts of HPV 5, 8, 9, 15 and 20 was investigated by real-time reverse transcription-PCR. HPV DNA load was determined for HPV 8, 9 and 15 in 11 biopsies. Antibody response was measured by enzyme-linked immunosorbent assay using affinity-purified, bacterially expressed complete viral proteins fused to glutathione S-transferase as antigens. RESULTS: HPV DNA was detected in 25 of 31 tissue samples, indicating eight single and 17 multiple HPV infections. E6/E7 transcripts of HPV 8, 9 and 15 were found in low copy numbers in one SCC and three AKs, but not in normal skin or Vv. All four patients examined showed antibodies to cutaneous HPV antigens, but the antibody response did not correlate with E6/E7 expression detected in the tumour. CONCLUSIONS: Transcriptional activity of the E6/E7 oncogenes in AK and SCC suggests an active role of HPV in the lesion.