The Role of Human Papillomaviruses and Polyomaviruses in BRAF-Inhibitor Induced Cutaneous Squamous Cell Carcinoma and Benign Squamoproliferative Lesions

Background: Human papillomavirus (HPV) has long been proposed as a cofactor in the pathogenesis of cutaneous squamous cell carcinoma (cSCC). More recently, the striking clinico-pathological features of cSCCs that complicate treatment of metastatic melanoma with inhibitors targeting BRAF mutations (BRAFi) has prompted speculation concerning a pathogenic role for oncogenic viruses. Here, we investigate HPV and human polyomaviruses (HPyV) and correlate with clinical, histologic, and genetic features in BRAFi-associated cSCC. Materials and Methods: Patients receiving BRAFi treatment were recruited at Barts Health NHS Trust. HPV DNA was detected in microdissected frozen samples using reverse line probe technology and degenerate and nested PCR. HPV immunohistochemistry was performed in a subset of samples. Quantitative PCR was performed to determine the presence and viral load of HPyVs with affinity for the skin (HPyV6, HPyV7, HPyV9, MCPyV, and TSPyV). These data were correlated with previous genetic mutational analysis of H, K and NRAS, NOTCH1/2, TP53, CDKN2A, CARD11, CREBBP, TGFBR1/2. Chromosomal aberrations were profiled using single nucleotide polymorphism (SNP) arrays. Results: Forty-five skin lesions from seven patients treated with single agent vemurafenib in 2012–2013 were analyzed: 12 cSCC, 19 viral warts (VW), 2 actinic keratosis (AK), 5 verrucous keratosis/other squamoproliferative (VK/SP) lesions, one melanocytic lesion and 6 normal skin samples. Significant histologic features of viral infection were seen in 10/12 (83%) cSCC. HPV DNA was detected in 18/19 (95%) VW/SP, 9/12 (75%) cSCC, 4/5 (80%) SP, and 3/6 (50%) normal skin samples and in 1/12 cases assessed by immunohistochemistry. HPyV was co-detected in 22/30 (73%) of samples, usually at low viral load, with MCPyV and HPyV7 the most common. SNP arrays confirmed low levels of chromosomal abnormality and there was no significant correlation between HPV or HPyV detection and individual gene mutations or overall mutational burden. Conclusion: Despite supportive clinicopathologic evidence, the role for HPV and HPyV infection in the pathogenesis of BRAFi-induced squamoproliferative lesions remains uncertain. Synergistic oncogenic mechanisms are plausible although speculative. Nonetheless, with the prospect of a significant increase in the adjuvant use of these drugs, further research is justified and may provide insight into the pathogenesis of other BRAFi-associated malignancies

Neoliberalism with a community face?:A critical analysis of asset-based community development in Scotland

In this article, we trace the ideological and social policy roots of asset-based community development (ABCD) in the United States and the United Kingdom, and explore how this approach has been legitimized in Scotland. We argue that ABCD is a capitulation to neoliberal values of individualization and privatization. Drawing on findings from our empirical work, we discuss how ABCD generates dilemmas for community development. Although some practitioners are able to adapt ABCD to focus on renewing Scottish democracy, several practitioners are using ABCD to privatize public issues such as inequality and justify dramatic cuts to the Scottish welfare state

Morphoea with Myositis: A Rare Association

In this case, we describe an unusual presentation of a young woman with a rash typical of morphoea (confirmed on biopsy), who went on to develop myositis in an atypical distribution. Although the association of myositis with diffuse systemic sclerosis is well described, the link with localised scleroderma (morphoea) and myositis has not been described

Superantigenic Activity of emm3 Streptococcus pyogenes Is Abrogated by a Conserved, Naturally Occurring smeZ Mutation

Streptococcus pyogenes M/emm3 strains have been epidemiologically linked with enhanced infection severity and risk of streptococcal toxic shock syndrome (STSS), a syndrome triggered by superantigenic stimulation of T cells. Comparison of S. pyogenes strains causing STSS demonstrated that emm3 strains were surprisingly less mitogenic than other emm-types (emm1, emm12, emm18, emm28, emm87, emm89) both in vitro and in vivo, indicating poor superantigenic activity. We identified a 13 bp deletion in the superantigen smeZ gene of all emm3 strains tested. The deletion led to a premature stop codon in smeZ, and was not present in other major emm-types tested. Expression of a functional non-M3-smeZ gene successfully enhanced mitogenic activity in emm3 S. pyogenes and also restored mitogenic activity to emm1 and emm89 S. pyogenes strains where the smeZ gene had been disrupted. In contrast, the M3-smeZ gene with the 13 bp deletion could not enhance or restore mitogenicity in any of these S. pyogenes strains, confirming that M3-smeZ is non-functional regardless of strain background. The mutation in M3-smeZ reduced the potential for M3 S. pyogenes to induce cytokines in human tonsil, but not during invasive infection of superantigen-sensitive mice. Notwithstanding epidemiological associations with STSS and disease severity, emm3 strains have inherently poor superantigenicity that is explained by a conserved mutation in smeZ.Publisher PDFPeer reviewe

Functional SMEZ is required to stimulate production of cytokines from human tonsil cells.

<p>Human tonsil cell suspensions were cultured with bacterial cell-free culture supernatants from GAS-M3_control(white bars), GAS-M3<i>_smeZ</i>_-M89 (black bars) and GAS-M3<i>_smeZ</i>_-M3 (gray bars). After 2 and 5 days incubation cell-free media were obtained from cultures and production of TNFα, TNFβ, IL-10, IL-17 and IFNγ were measured by ELISA. Horizontal dotted lines represent the mean level of cytokines produced after co-culture with bacterial cell-free culture supernatants from GAS-M1_control on day 2 and day 5. Mean (+ standard deviation) of three replicates measured in duplicate. Representative of two experiments performed on different donors. Statistical analysis was performed using ANOVA with Bonferroni multiple comparison.</p

<i>Emm</i>3 STSS isolates show <i>emm</i>-type specific differences in overall mitogenicity.

<p><i>A.</i>Human MNC proliferation response to culture supernatants from 63 <i>Streptococcus pyogenes</i> streptococcal toxic shock syndrome (STSS) isolates grouped by <i>emm</i>-type (numbers per group: <i>emm</i>1, n = 11; <i>emm</i>3, n = 12; <i>emm1</i>2 and <i>emm</i>87, n = 11; <i>emm</i>18, n = 5; <i>emm</i>28, n = 7; <i>emm</i>89, n = 6). Negative; tissue culture media (RMPI) alone. Outliers are represented as individual circles. Representative of 2 experiments performed on different donors. <i>B.</i> Human MNC proliferation response to sera obtained from CD1 mice 24 hours after being infected with one of three <i>S. pyogenes</i> strains representing each <i>emm</i>-type; two mice were infected per strain. Negative; uninfected mouse serum. Proliferative response is measured as counts per minute (cpm) of tritiated-thymidine uptake. Median and 5th, 25th, 50th, 75th, and 95th centiles shown.</p

<i>emm</i>3 strains have low mitogenicity due to the mutation in <i>smeZ</i>.

<p><i>A.</i> MNC proliferative response to culture supernatants from an <i>emm</i>3 isolate, GAS-M3 carrying the typical M3-<i>smeZ</i> with 13 bp deletion, GAS-M3 over-expressing the functional M89 form of SMEZ (GAS-M3<i>_smeZ</i>_-M89) and GAS-M3 over-expressing the M3-SMEZ (GAS-M3<i>_smeZ</i>_-M3). <i>B.</i> Experimental <i>smeZ</i> mutation in <i>emm</i>1 <i>S. pyogenes</i> (GAS-M1) reduced MNC proliferation (GAS-M1Δ<i>smeZ</i>). Proliferative response was restored when GAS-M1Δ<i>smeZ</i> over-expressed the functional M89 form of SMEZ (GAS-M1<i>_smeZ</i>_-M89) but not with M3-type form of SMEZ (GAS-M1<i>_smeZ</i>_-M3). <i>C</i>. A similar result was obtained using an <i>emm</i>89 strain (GAS-M89) with an experimental mutation in <i>smeZ</i> (GAS-M89Δ<i>smeZ</i>). Proliferation was restored when GAS-M89ΔsmeZ over-expressed the functional M89 form of SMEZ (GAS-M89<i>_smeZ</i>_-M89) but not with M3-type form of SMEZ (GAS-M89<i>_smeZ</i>_-M3). Negative; media alone.Proliferation was measured as counts per minute (cpm) of tritiated-thymidine uptake and the percentage proliferation for each strain was calculated relative to the wild type strain (GAS-M3, GAS-M1, GAS-M89 respectively). Data are mean (+standard deviation) of three measurements. Representative of two experiments performed using different donor MNC.</p

<i>emm</i>3 isolates have a 13 base pair (bp) deletion within the <i>smeZ</i> locus.

<p>Representation of the <i>smeZ</i> locus from <i>emm</i>1 (M1) and <i>emm</i>3 (M3) strains. Nucleotides 1 to 3 encode the start codon (shown in bold). From 73 bp of the nucleotide sequence, the amino acid sequence of the mature SMEZ protein is shown. <i>Emm</i>3 strains have a 13 bp deletion at 316 bp (highlighted by a shaded box) that results in a frameshift and a predicted premature stop codon after 86 amino acids (shown as *). The forward primer and the reverse primer amplify the full length <i>smeZ</i> locus. The 13 bp deletion was detected using a forward (truncated) primer that anneals specifically to the region containing the deletion.</p