Expression of apoptosis-inducing factor (AIF) in keratoacanthomas and squamous cell carcinomas of the skin

The complex biological trait 'susceptibility to apoptosis' is a nosological feature distinguishing squamous cell carcinomas (SCC) from keratoacanthomas (KA). The purpose of this study was to compare the expression of apoptosis-inducing factor (AIF), a major effector of the caspase-independent apoptosis pathway, in formalin-fixed SCC (N = 23) and KA (N = 29) resection specimens. SCC express statistically significant more AIF than KA both as proportion of AIF+ cells by immunohistochemistry (median: 54% vs 33%; P < 0.01) and as total AIF protein content by western blot quantification (six-fold increased; P < 0.01). However, the contribution of AIF to apoptosis, measured as fraction of apoptotic nuclei with overt DNA fragmentation by the TUNEL method that co-express AIF translocated to nucleus, is significantly less prevalent among SCC (median: 19% vs 48% in KA; P < 0.01). These findings indicate to a distinctive involvement of AIF in the progression of certain epithelial skin tumors that might be exploited as a promising treatment target.     

Expression of stromelysin 3 in keratoarcanthoma and squamous cell carcinoma

Stromelysin 3 (ST3) is a member of the metalloproteinase family, which is expressed in the skin during wound healing and in the stroma of basal cell carcinoma. A high level of expression of ST3 has been observed in carcinomas of poor prognosis. In benign tumors, though, ST3 is not expressed or is at a low level. We have immunohistochemically studied the expression of ST3 in 89 randomly selected cases of squamous cell carcinomas (SCCs), 104 of keratoacanthomas (KA), and 23 cases of metastatic SCC. Stromelysin 3 was expressed only by fibroblasts surrounding the tumors and not by epithelial cells. The proportion of tumors positively stained was 22% of KA, 47% of randomly selected SCC, and 70% of metastatic SCC. In areas of poorly demarcated neoplastic cells, a reinforcement of the staining was observed in the stroma. The intensity and dispersion of staining were used to determine the level of expression. There were significantly more SCC in the groups of high expression levels, and both parameters were significantly higher in SCC than in KA. Expression of ST3 in benign tumors is unusual. Its expression in the the stroma of keratoacanthomas can be related to the high tissue remodeling activity observed in these tumors. It also could be interpreted as in favor of the neoplastic nature of KA. Nevertheless, the level of expression was higher in SCC than in KA and seemed to be related to the prognosis of these tumors. These results correlate well with those obtained in breast cancers and in noncutaneous SCC.     

Differential diagnosis of keratoacanthomas and squamous cell carcinomas: diagnostic value of DNA image cytometry and p53 expression

DNA-cytometry and immunohistochemistry with the anti-p53 antibody DO-I was performed in 24 keratoacanthomas (KA) and 21 squamous cell carcinomas (SCC) (13 well-differentiated, 8 moderately differentiated) to establish the possible value of these methods for the differential diagnosis of both epithelial tumors. Aneuploidy was detected in 1 (4%) KA and 12 (57%) SCC ( p < 0.05). In the latter tumors, histologic grade was associated with an abnormal DNA-content. Six (46%) well and 6 (75%) moderately differentiated SCC were shown to be aneuploid. Over-expression of p⁵³ protein was found in 16 (76%) SCC and 14(66%) KA ( p >0.05; not significant). However, quantification of p⁵³ expression by evaluating both the intensity of immunostaining and the number of cells with over-expression by means of an immunoreactivity score (IRS) showed significant differences ( p <0.05). There was no correlation of p⁵³ over-expression and aneuploidy in the tumors examined. The analysis of ploidy and immunostaining with anti-p⁵³ antibodies may give useful additional information regarding the differential diagnosis of SCC and KA, if only aneuploidy or a high IRS are considered.
Pilch H, Weiss J, Heubner C, Heine M. Differential diagnosis of keratoacanthomas and squamous cell carcinomas: diagnostic value of DNA image cytometry and p53 expression.     

P16 is overexpressed in cutaneous carcinomas located on sun-exposed areas

BACKGROUND: Recently, an increased expression of P16, a cell cycle regulatory tumor suppressor protein, has been demonstrated in cervical squamous neoplasms as a marker of malignancy. In contrast, studies performed in skin carcinomas led to contradictory results. OBJECTIVES: Our first aim was to evaluate P16 expression in different types of non-melanoma skin cancers compared with normal skin and benign tumors. The second aim was to evaluate the relationship between P16 expression and the location of skin tumors (i.e. exposed versus non exposed sites). Finally, we also studied Ki67 expression in skin carcinomas and control biopsies. METHODS: Skin biopsy specimens with typical histologic features of squamous cell carcinoma (SCC; n = 30), Bowen's disease (BD; n = 17), basal cell carcinoma (BCC; n = 10), seborrheic keratosis (SK; n = 10) and normal human skin (NHS; n = 9) were obtained from 76 patients seen at our institution between 2001 and 2003. In all cases, P16 and Ki67 expression were evaluated by immunohistochemistry and image analysis. RESULTS: P16 overexpression was observed in 58% of cutaneous carcinomas (SCC: 60%; BD: 58%; BCC: 50%) versus 0% of SK or NHS (0%) (p = 0.006). Ki67 expression in over 5% of tumour cells was observed in 69% of cutaneous carcinomas (SCC: 54%; BD: 76%; BCC: 80%) versus 16% in the group including SK (30%) and NHS (0%) (p = 0.04). Overexpression of P16 was associated with a high rate of Ki67 positive tumour cells in 23/57 malignant skin tumors (40%). Both P16 was associated and Ki-67 were negative in 7/57 cases (12%). Sixty-eight percent of tumors located on sun-exposed areas versus 23% of those located on non sun-exposed areas overexpressed P16 (p = 0.02). CONCLUSION: Our study demonstrated that the expression of P16 and Ki67 is associated with skin carcinomas. No difference was observed according to histological types of carcinomas, suggesting that P16 and Ki67 expression did not correlate with the degree of proliferation and malignancy. Within cutaneous carcinoma specimens, P16 overexpression was significantly associated with the location on sun-exposed areas, suggesting a possible induction of P16 overexpression by UV radiation.     

Reduced ultraviolet irradiation delays subsequent squamous cell carcinomas in hairless mice

Background: Ultraviolet (UV) radiation induces non-melanoma skin cancer (NMSC), and UV prophylaxis is essential to prevent skin cancer. It is unclear whether patients diagnosed with squamous cell carcinomas (SCC) may benefit from reduced UV exposures in terms of delaying the development of new tumors. The objective was to evaluate the significance of discontinued or reduced UV exposure for the development of subsequent skin tumors.
Methods: Seven groups of mice ( n =175) were irradiated with UV doses of 2 and 4 standard erythema doses (SED) that were continued, reduced or discontinued at the time of appearance of the first skin tumor.
Results: The development of new tumors was delayed, corresponding to the degree of reductions in UV dose in an inversely linear manner. Discontinuation of UV doses delayed the median times to the second tumor by 24 days (2 SED, P =0.0549) and 33.5 days (4 SED, P <0.0001), and when reduced to 1 SED, the median delays were 18 days (2 SED, P =0.0469) and 33 days (4 SED, P <0.0001). The median delay to the third tumor was after UV reduction 47 days (4 SED, P <0.0001) and 35 days (2 SED, P =0.151), and after UV discontinuation 49 days (4 SED, P <0.0001) and 44 days (2 SED, P =0.111).
Conclusion: This suggests that patients with SCC may benefit from reduced UV exposure.     

Cancer/testis antigen MAGE-A4 expression pattern differs in epithelial skin tumors of organ-transplant recipients and immunocompetent patients

BACKGROUND: Lifetime risk for squamous cell carcinoma (SCC) of the skin is 1:30. Risk in organ-transplant recipients (OTR) is increased over 60-fold through long-term drug-induced immunosuppression. MAGE family-derived peptides are cancer/testis antigens recognized by specific CD8(+) T cells and employed for immunotherapy. We were interested in the frequency and distribution of MAGE-A4 in epithelial skin tumors of OTR and immunocompetent patients. METHODS: mAb 57B predominantly recognizing MAGE-A4 was used to stain 119 formalin-fixed, paraffin-embedded epithelial skin tumors (actinic keratosis, bowenoid actinic keratosis, Bowen's disease, and SCC; n = 17, 25, 61, 16, respectively) in immunocompetent patients (n = 84) and OTR (n = 35). RESULTS: All four epithelial skin tumors showed comparable immunoreactivity ranging from (25-71%, p = 0.361). Scattered immunoexpression pattern was more frequent in OTR (p = 0.025). SCC showed polarized immunoreactivity basally (p = 0.002). CONCLUSION: MAGE-A4 was expressed in a large part of epithelial skin tumors with predominantly scattered immunoexpression pattern in OTR. The difference in immunoexpression pattern for immune status was limited, suggesting important non-immunosuppressor-mediated mechanisms for increased skin carcinogenesis in OTR. mAb 57B may be a helpful tool for immunohistochemistry and micrographic surgery using formalin-fixed paraffin-embedded tissue.     

Expression of vascular endothelial growth factor in basal cell carcinoma and cutaneous squamous cell carcinoma of the head and neck

BACKGROUND: Basal cell carcinomas (BCC) and squamous cell carcinomas (SCC) can both arise from any cutaneous epithelial surface. BCC are slow growing and rarely metastasise, whereas SCC are usually more aggressive. It is likely that the angiogenic process plays a key role in determining rate of growth and propensity for dissemination. Angiogenesis is a complex process requiring many factors and a pivotal group of proteins involved in this process is vascular endothelial growth factor (VEGF). METHODS: Immunohistochemical expression of VEGF was assessed in 44 cases of BCC and 41 cases of cutaneous SCC from the head and neck region. RESULTS: VEGF was expressed by blood vessel endothelial cells in both adjacent skin and tumour, and in the basal keratinocyte layer of epidermis. In BCC, VEGF was expressed by tumour epithelial cells, predominantly at the invasive tumour front, in 24/44 cases and its expression was significantly greater than in adjacent skin (p = 0.038). More widespread VEGF expression was found in 32/41 cases of SCC, and it was significantly associated with the degree of tumour differentiation (p < 0.001). CONCLUSIONS: The patterns of VEGF expression in BCC and SCC may help to explain the different behaviour that is usually seen with these tumours.     

EGFR and MYC gene copy number aberrations are more common in squamous cell carcinoma than keratoacanthoma: a FISH study

Epidermal growth factor receptor (EGFR) and MYC genomic aberrations have been described in cutaneous squamous cell carcinoma (SCC) but have not been widely investigated in keratoacanthoma (KA). EGFR and MYC were evaluated by fluorescence in situ hybridization and immunohistochemistry in 8 verrucae, 19 involuting KA (IKA), 23 classic KA (CKA), 6 atypical KA (AKA) and 19 SCC. Increased EGFR gene copy number was seen in 9 of 23 CKA and 14 of 19 SCC (p = 0.03). Increased MYC gene copy number was observed in 7 of 23 CKA and 17 of 19 SCC (p = 0.0001). MYC gene amplification was more common in SCC than CKA (p = 0.005), while EGFR gene amplification was rare and not significant. MYC protein overexpression was identified in 6 of 23 CKA and 14 of 19 SCC (p = 0.005). There was no statistical difference in EGFR protein overexpression in SCC and CKA (p = 0.06). EGFR and MYC aberrations were rare in IKA. AKA showed EGFR and MYC anomalies at an incidence intermediate between CKA and SCC. EGFR and MYC gene copy number aberrations are more common in SCC than KA. The incidence of aberrations parallels the degree of cytologic atypia in KA.     

某些表皮肿瘤中CDK4、P16表达及其与临床病理特征的关系

目的:探讨CDK4、P16在某些表皮肿瘤中的表达及其与临床病理特征的关系。方法:采用ABC免疫组化技术观察48例皮肤鳞状细胞癌、41例基底细胞癌和15例角化棘皮瘤中CDK4、P16的表达。结果:鳞状细胞癌中CDK4表达明显高于基底细胞癌及角化棘皮瘤(P<0.01,P<0.05),P16表达则相反(P<0.05)。二者的表达在基底细胞癌与角化棘皮瘤之间无显著性差异(P>0.05)。CDK4在高分化鳞状细胞癌中的表达明显低于低分化者(P<0.05),在低分化及伴有淋巴结转移的鳞状细胞癌中P16表达明显下调(P<0.05),二者的表达与患者的性别、年龄、发病部位及病程无关(P>0.05)。结论:P16表达减少和CDK4表达增加可能参与皮肤癌的发生、发展和转移。     

Effects of biopsy-induced wound healing on residual basal cell and squamous cell carcinomas: rate of tumor regression in excisional specimens

BACKGROUND: Wound healing following a partial biopsy of basal cell (BCC) and squamous cell carcinomas (SCC) may induce tumor regression. METHODS: Nonmelanoma skin cancer (NMSC) biopsy and re-excision specimens from 1994 to 2001 were reviewed for histologic evidence of scar vs. presence of residual tumor in excision specimens. Regressed and non-regressed tumors were analyzed to assess the influence of anatomic location, biopsy technique (punch vs. shave), histologic subtype of BCC or SCC, time interval between biopsy and excision, and patient age. RESULTS: Nine hundred and ten excisions were performed for transected BCC or SCC, 217 (24%) of which showed scar with no residual tumor. Logistic regression analysis revealed significant differences in the regressed vs. non-regressed subsets. SCCs were more likely to regress than BCCs (40% vs. 20%, respectively, p < 0.00001). Independent of the NMSC type, tumors regressed more often following shave rather than punch biopsy (34% vs. 15%, respectively, p < 0.00001), as did tumors on the trunk and extremities compared with head and neck cases (31% vs. 21%, respectively, p < 0.01). CONCLUSIONS: In our series, 24% of NMSCs transected on the initial biopsy showed no residual tumor in the excision specimens, implying that some event in the interval between biopsy and excision may lead to the eradication of residual tumor. The exact mechanism is unclear, but wound healing likely plays an important role.     

Trisomy 7 in keratoacanthoma and squamous cell carcinoma detected by fluorescence in-situ hybridization

Keratoacanthoma (KA) is generally considered to be a clinically and histologically distinct entity, but it often remains difficult to separate from well-differentiated squamous cell carcinoma (WDSCC). Recently, trisomy 7 has been identified in squamous cell carcinoma of the skin. In this study, we examined classical KA (n=6), WDSCC (n=7) and squamous cell carcinoma with KA-like features (SCC-KA) (n=8) for trisomy 7 by fluorescence in-situ hybridization (FISH) to determine if this chromosomal abnormality is unique to squamous lesions diagnosed as WDSCC, or shared by both KA and SCC. In addition, the pertinent clinical-histopathologic findings were summarized. Trisomy 7 was identified in one KA, one SCC-KA and two WDSCC. This study demonstrates that there is a chromosomal abnormality shared by KA and SCC, providing further evidence that KA is most likely a form of SCC. Further studies are required to determine if trisomy 7 in these lesions is of prognostic significance.     

Some further features for differential diagnosis between squamous cell carcinomas and basal cell epitheliomas

Two further methods for the characterization of epidermal skin tumors are described: the antinuclear antibody (ANA) immunofluorescent test, which consists of indirect immunofluorescence with known high titer sera containing homogenous ANAs on epidermal skin tumors, and the ammoniacal-silver cytochemical method, which specifically stains nuclear histones. Squamous cell carcinomas (SCCs), basal cell epitheliomas (BCEs) as well as control specimens from normal skin and benign epidermal hyperplasias were studied. The ANA immunofluorescent test was positive for most SCCs, mixed SCC and basal cell carcinomas and metatypical BCEs. The ammoniacal-silver method gave a characteristic staining pattern shared among SCCs, mixed carcinomas and metatypical BCEs. BCEs, besides metatypical ones, were always negative by the ANA immunofluorescent test and the same applied for the control specimens. The ammoniacal-silver method gave a characteristic staining pattern for BCEs and control sections quite different from the staining pattern of the more aggressive forms of epidermal tumors. The two methods usually yielded parallel results.     

Comparative analysis of the expression of ERBIN and Erb-B2 in normal human skin and cutaneous carcinomas

BACKGROUND: ERBIN is a binding partner of Erb-B2, an orphan receptor within the Erb-B family critically involved in the regulation of cell growth and differentiation. Although its function remains unclear, ERBIN is thought to affect the polarity of epithelial cells and cell growth via the Ras signalling pathway. OBJECTIVES: To examine and compare the tissue distribution and the expression levels of ERBIN and Erb-B2 in normal skin and in cutaneous carcinomas. METHODS: Fifteen cases of basal cell carcinoma (BCC), 12 cases of squamous cell carcinoma (SCC) and five cases of keratoacanthoma (KA) were analysed by immunohistochemistry on paraffin-embedded sections using anti-ERBIN and anti-Erb-B2 antibodies. RESULTS: ERBIN and Erb-B2 had a similar distribution in normal human skin. They were primarily localized at the plasma membrane in differentiated keratinocytes and in duct cells from eccrine glands, whereas they were localized diffusely in the cytoplasma of basal keratinocytes. In both SCC and KA the subcellular distribution of ERBIN and Erb-B2 remained unchanged, whereas both proteins were redistributed from the plasma membrane into cytosolic aggregates in BCC. CONCLUSIONS: The subcellular localization of ERBIN in normal human skin is similar to that of Erb-B2 and varies with cell differentiation. Based on our findings and on the biological activities of Erb-B2, it is conceivable that disturbed expression or functioning of ERBIN and Erb-B2 is implicated in the development of the malignant phenotype of BCC.     

Stromal CD10 expression, as well as increased dermal macrophages and decreased Langerhans cells, are associated with malignant transformation of keratinocytes

Background:  It has become evident that resident stromal cells, such as fibroblasts and inflammatory cells, are involved in the metastatic process, including proliferation or migration of malignant neoplasms. We analyzed CD10+ stromal cells, dermal macrophages and Langerhans cells (LCs) in skin tumors.
Methods:  Immunohistological staining was performed with markers for macrophages (CD68), LC (CD1a), stromal fibroblasts (CD10) and cell proliferation (Ki67) in 12 normal skins (NSs) and 15 cases each of seborrheic keratosis (SK), actinic keratosis (AK), keratoacanthoma (KA), Bowen's disease (BD) and squamous cell carcinoma (SCC).
Results:  All SCCs showed weak to strong stromal CD10 expression, while all NS, SK and AK were negative. Weak CD10 expression was observed in only 2 of 15 samples in both BD and KA. The number of CD68+ cells and Ki67 labeling index in SCC and BD were significantly higher than that in KA, AK and SK. In contrast, the number of LC was lower in SCC and BD. The stromal CD10 expression was significantly correlated with the Ki67 labeling indices and CD68+ cells and negatively correlated with decreased LC.
Conclusions:  The stromal CD10 expression is associated with malignant transformation of keratinocytes together with infiltration of dermal macrophages and loss of LC.     

角化棘皮瘤和皮肤鳞状细胞癌bcl—2和Nm23癌基因蛋白表达的比较研究

目的：通过研究分析bcl-2和Nm23蛋白在角化棘皮瘤（KA）和皮肤鳞状细胞癌（SCC）中的表达和临床意义。探寻两病的鉴别标志。方法：应用免疫组化技术（SABC）对11例KA，28例SCC进行了检测。结果：bcl-2和Nm23蛋白在KA和SCC中的表达经统计学分析无显著性意义。结论：bcl-2和Nm23癌基因均参与了KA和SCC的发病过程，但都不能作为KA和SCC的鉴别标志。     

Widespread cutaneous carcinomas associated with human papillomaviruses 5, 14 and 20 after introduction of methotrexate in two long-term PUVA-treated patients

BACKGROUND: PUVA treatment for patients with severe psoriasis has been demonstrated to be highly effective. However, an increased risk of nonmelanoma and melanoma skin cancers has been reported. It is generally accepted that the risk of squamous-cell carcinoma (SCC) is significantly increased in patients with long-term PUVA therapy. The role of methotrexate (MTX) and infection with oncogenic human papillomaviruses which may act as cocarcinogens is poorly documented. CASE REPORTS: Two cases of multiple SCCs associated with numerous PUVA keratoses and PUVA freckles after long-term PUVA therapy and subsequent treatment with MTX are presented. In 1 case, the tumor progressed to metastatic SCC. Tumors and scrapings of psoriatic skin lesions were analyzed for the presence of oncogenic human papillomavirus (HPV) genotypes. The genotype of HPV-5, -14 and -20 was detected in scrapings and skin tumors using PCR amplification. CONCLUSION: These observations support the concept that long-term PUVA treatment is carcinogenic and rise questions concerning an additional influence of MTX in the development and progression of skin cancer. The risk of metastatic SCC seems to be underestimated in high-dose PUVA-treated patients due to longer latency for developing metastases and the small number of studies with long-term follow-up. Treatment with MTX should be considered cautiously in patients previously exposed to high doses of PUVA. The presence of oncogenic HPVs in carcinomas and psoriatic skin lesions detected only with the highly sensitive nested PCR method is not necessarily a proof of their implication in skin carcinogenesis.     

Cutaneous squamous cell carcinoma: a comprehensive clinicopathologic classification--part two

Cutaneous squamous cell carcinoma (SCC) includes many subtypes with widely varying clinical behaviors, ranging from indolent to aggressive tumors with significant metastatic potential. However, the tendency for pathologists and clinicians alike is to refer to all squamoid neoplasms as generic SCC. No definitive, comprehensive clinicopathological system dividing cutaneous SCCs into categories based upon their aggressiveness has yet been promulgated. Therefore, we have proposed the following based upon the malignant potential of SCC variants, separating them into categories of low (< or = 2% metastatic rate), intermediate (3-10%), high (greater than 10%), and indeterminate behavior. Low-risk SCCs include SCC arising in actinic keratosis, HPV-associated SCC, tricholemmal carcinoma, and spindle cell SCC (unassociated with radiation). Intermediate-risk SCCs include adenoid (acantholytic) SCC, intraepidermal epithelioma with invasion, and lymphoepithelioma-like carcinoma of the skin. High-risk subtypes include de novo SCC, SCC arising in association with predisposing factors (radiation, burn scars, and immunosuppression), invasive Bowen's disease, adenosquamous carcinoma, and malignant proliferating pilar tumors. The indeterminate category includes signet ring cell SCC, follicular SCC, papillary SCC, SCC arising in adnexal cysts, squamoid eccrine ductal carcinoma, and clear-cell SCC. Subclassification of SCC into these risk-based categories, along with enumeration of other factors including tumor size, differentiation, depth of invasion, and perineural invasion will provide prognostically relevant information and facilitate the most optimal treatment for patients.     

Human papillomavirus-DNA loads in actinic keratoses exceed those in non-melanoma skin cancers

Recent studies suggest a role of cutaneous human papillomaviruses (HPV) in non-melanoma skin cancer (NMSC) development. In this study viral DNA loads of six frequent HPV types were determined by quantitative, type-specific real-time-PCR (Q-PCR) in actinic keratoses (AK, n=26), NMSC (n=31), perilesional tissue (n=22), and metastases of squamous cell carcinomas (SCC) (n=8) which were previously shown to be positive for HPV5, 8, 15, 20, 24, or 36. HPV-DNA loads in AK, (partially microdissected) NMSC, and perilesional skin ranged between one HPV-DNA copy per 0.02 and 14,200 cell equivalents (median: 1 HPV-DNA copy per 344 cell equivalents; n=48). In 32 of the 79 HPV-positive skin biopsies and in seven of the eight metastases viral loads were even below the detection limit of Q-PCR. Low viral loads in NMSC were confirmed by in situ-hybridization showing only a few HPV-DNA-positive nuclei per section. Viral loads in SCC, basal cell carcinomas, and perilesional tissue were similar. But, viral loads found in AK were significantly higher than in SCC (p=0.035). Our data suggest that persistence of HPV is not necessary for the maintenance of the malignant phenotype of individual NMSC cells. Although a passenger state cannot be excluded, the data are compatible with a carcinogenic role of HPV in early steps of tumor development.