Tak Wah Wong

DNAzyme targeting c-jun suppresses skin cancer growth

Catalytic DNA molecules that target the transcription factor c-jun inhibit skin cancer growth in mice. Getting Under Cancer’s Skin Summer brings to mind barbecues, baseball, and trips to the local pool. Yet, outdoor fun can be hazardous to one’s health—too much sun exposure can increase the risk of developing skin cancer. Indeed, one in three cancers worldwide is skin-related, and currently available treatments may induce scarring or other toxicities. Cai et al. now report that the DNAzyme Dz13—which targets an mRNA that encodes a cancer-associated transcription factor, c-Jun—inhibits the growth of two common types of skin cancers: basal cell and squamous cell carcinomas. DNAzymes are single-stranded, all-DNA, catalytic molecules that specifically bind and cleave their target RNAs. The authors examined the effects of Dz13, which destroys c-jun mRNA, on animal models of skin cancer. Dz13 inhibited tumor growth, blocked neovascularization, and prevented metastasis in mouse models of skin cancer—effects that were mediated, in part, through the induction of antitumor immunity. Minimal toxicity was observed in Dz13-treated cynomolgus monkeys, minipigs, and rodents, and there were no off-target effects in more than 70 in vitro bioassays. Thus, Dz13 may prove to be a safe, effective therapy for skin cancer. Nonetheless, one is advised to pack the sun block in preparation for extra innings—or a fifth set. Worldwide, one in three cancers is skin-related, with increasing incidence in many populations. Here, we demonstrate the capacity of a DNAzyme-targeting c-jun mRNA, Dz13, to inhibit growth of two common skin cancer types—basal cell and squamous cell carcinomas—in a therapeutic setting with established tumors. Dz13 inhibited tumor growth in both immunodeficient and immunocompetent syngeneic mice and reduced lung nodule formation in a model of metastasis. In addition, Dz13 suppressed neovascularization in tumor-bearing mice and zebrafish and increased apoptosis of tumor cells. Dz13 inhibition of tumor growth, which required an intact catalytic domain, was due in part to the induction of tumor immunity. In a series of good laboratory practice–compliant toxicology studies in cynomolgus monkeys, minipigs, and rodents, the DNAzyme was found to be safe and well tolerated. It also did not interfere in more than 70 physiologically relevant in vitro bioassays, suggesting a reduced propensity for off-target effects. If these findings hold true in clinical trials, Dz13 may provide a safe, effective therapy for human skin cancer.

Histopathological differential diagnosis of keloid and hypertrophic scar

Distinguishing hypertrophic scar (HS) from keloid histopathologically is sometimes difficult because thickened hyalinized collagen (keloidal collagen), the hallmark of keloid, is not always detectable and &agr;-smooth muscle actin (&agr;-SMA), a differentiating marker of HS, is variably expressed in both forms of scar. The aim of this study was to investigate additional distinguishing features to facilitate differentiation between keloid and HS. We compared various histologic features and the expression of &agr;-SMA in 40 specimens of keloid and 10 specimens of HS. The features more commonly seen in keloids were: (a) no flattening of the overlying epidermis, (b) no scarring of the papillary dermis, (c) presence of keloidal collagen, (d) absence of prominent vertically oriented blood vessels, (e) presence of prominent disarray of fibrous fascicles/nodules, (f) presence of a tongue-like advancing edge underneath normal-appearing epidermis and papillary dermis, (g) horizontal cellular fibrous band in the upper reticular dermis, and (h) prominent fascia-like fibrous band. The last three features were found in keloid specimens only, including the ones lacking detectable keloidal collagen. Our study confirmed the diagnostic value of keloidal collagen, but it was only found in 55% of keloid specimens. &agr;-SMA expression was found in both HS (70%) and keloid (45%), thus it would not be a differentiating marker. In scars with no detectable keloidal collagen, the presence of the following feature(s) favors the diagnosis of keloid: non-flattened epidermis, non-fibrotic papillary dermis, a tongue-like advancing edge, horizontal cellular fibrous band in the upper reticular dermis, and prominent fascia-like band.

Bactericidal Effects of Toluidine Blue-Mediated Photodynamic Action on Vibrio vulnificus

ABSTRACT Vibrio vulnificus is a gram-negative, highly invasive bacterium responsible for human opportunistic infections. We studied the antibacterial effects of toluidine blue O (TBO)-mediated photodynamic therapy (PDT) for V. vulnificus wound infections in mice. Fifty-three percent (10 of 19) of mice treated with 100 μg of TBO per ml and exposed to broad-spectrum red light (150 J/cm2 at 80 mW/cm2) survived, even though systemic septicemia had been established with a bacterial inoculum 100 times the 50% lethal dose. In vitro, the bacteria were killed after exposure to a lower light dose (100 J/cm2 at 80 mW/cm2) in the presence of low-dose TBO (0.1 μg/ml). PDT severely damaged the cell wall and reduced cell motility and virulence. Cell-killing effects were dependent on the TBO concentration and light doses and were mediated partly through the reactive oxygen species generated during the photodynamic reaction. Our study has demonstrated that PDT can cure mice with otherwise fatal V. vulnificus wound infections. These promising results suggest the potential of this regimen as a possible alternative to antibiotics in future clinical applications.

Human skin surface lipid film: An ultrastructural study and interaction with corneocytes and intercellular lipid lamellae of the stratum corneum

Sebum is a complex mixture of lipids, which is secreted by mammalian sebaceous glands, and forms a fluid film over the skin surface. After sebum is secreted, it becomes mixed with lipid from the keratinizing epithelium and forms the skin surface lipid film (SSLF). Until now, direct fine structural observation of the SSLF has been lacking. In the present work, we viewed the detailed structures of the human SSLF by ruthenium tetroxide staining. The results showed that the SSLF formed an amorphous sheet of variable thickness on the skin surface instead of forming lipid droplets, as had been the usual assumption. In general, its thickness was < 0.5 μm or even negligible in sebum‐poor extremities. However, in the sebum‐rich face, its thickness was > 4 μm in focal areas. Consistent with the thickness of SSLF, the sebum quantity showed great regional variation. It varied from 1 μg/cm2 (leg) to 189 ± 42.7 μg/cm2 (mean ± SD: face). The SSLF was composed of numerous fine granules of about 4–5 nm in a random orientation. Within the SSLF, variable amounts of deranged lipid lamellae derived from corneocytes were mixed with sebum. As well as on the skin surface, a similar amount of sebum was also found between the desquamating corneocytes in the uppermost several layers of the stratum corneum (SC). We also observed the presence of intercellular lipid lamellae in the outer layers of the SC: their lipid envelope remained intact even in desquamated corneocytes. Our results provide some new insights concerning the structure of the SSLF and its relationship with the SC.

Depigmented genital extramammary Paget’s disease: A possible histogenetic link to Toker’s clear cells and clear cell papulosis

Background: The histogenesis of extramammary Paget’s disease (EMPD) is still controversial. Benign pagetoid cells of the nipple first described by Toker and the similar clear cells found in white maculopapules of clear cell papulosis (CCP) have been proposed to be potential precursor cells giving rise to EMPD and primary intraepidermal Paget’s disease in the nipple. The observation of a rare case of depigmented EMPD provided us with a chance to examine further the interesting Toker’s clear cell/CCP hypothesis.

Pilot study of topical delivery of methotrexate by electroporation.

Background  The topical administration of methotrexate (MTX) for the treatment of psoriasis and neoplastic diseases is restricted by the poor diffusion of MTX across the stratum corneum.

In vitro/in vivo correlations between transdermal delivery of 5-aminolaevulinic acid and cutaneous protoporphyrin IX accumulation and effect of formulation

Summary Background Photodynamic therapy (PDT) using topical application of 5‐aminolaevulinic acid (ALA) has been widely reported for the treatment of a variety of neoplastic and non‐neoplastic cutaneous diseases. Although different formulations containing variable amounts of ALA have been applied in PDT, the dose–response relationships between transdermal ALA delivery and cutaneous protoporphyrin IX (PpIX) accumulation have not been studied.

Molecular mimicry between streptococcal pyrogenic exotoxin B and endothelial cells

Molecular mimicry between group A streptococcus and host antigens has important roles in the development of post-streptococcal sequelae, including glomerulonephritis and rheumatic heart disease (RHD). The etiology of RHD involves host cross-reactivity with M proteins and carbohydrate antigens. In this study, we show that anti-streptococcal pyrogenic exotoxin B (SPE B) antibodies exhibited characteristics of autoantibodies, which cross-react with endothelial cells. Immunoglobulin G (IgG) deposition and complement activation were observed in the heart valve of SPE B-immunized mice. In addition, apoptosis in the heart valve was detected in SPE B-immunized mice. An anti-SPE B monoclonal antibody (mAb) 10G showed cross-reactivity with human microvascular endothelial (HMEC-1) cells and mouse valve endothelial cells. Passive immunization with mAb 10G also caused IgG deposition, complement activation, and apoptotic cell death in the mouse heart valve. We conducted peptide array and ELISA using synthetic peptides to identify the SPE B antigenic epitope recognized by mAb 10G. Results showed that the major epitope of mAb 10G is localized to amino-acid residues 296–310 of SPE B (P7–8). The cross-reactivity of mAb 10G with endothelial cells was inhibited using P7–8 peptides for competition. These results suggest that anti-SPE B antibodies cross-react with endothelial cells, and that a dominant epitope is located within the amino-acid residues 296–310 of SPE B. Moreover, we found that mAb 10G can also bind to N-acetyl-β-D-glucosamine (GlcNAc) conjugated with bovine serum albumin (BSA), but not to BSA or M1 protein. Competition assay showed that the binding activity of mAb 10G with GlcNAc-BSA and P7–8 of SPE B was inhibited by pretreatment with GlcNAc-BSA or P7–8 peptides. Therefore, our results suggest that conformational molecular mimicry may exist between SPE B and GlcNAc.

Solanum incanum extract (SR-T100) induces human cutaneous squamous cell carcinoma apoptosis through modulating tumor necrosis factor receptor signaling pathway

BACKGROUND The Solanum species herbs have been used to treat cancer for centuries; however, the underlying mechanisms and effectiveness in vivo remain unclear. OBJECTIVES SR-T100, extracted from the Solanum incanum, contains solamargine alkaloid as the main active ingredient. Here, we investigated the apoptosis-inducing effects of SR-T100 for targeting squamous cell carcinoma (SCC) in vitro and in vivo. METHODS We elucidated the mechanism by which SR-T100 induces apoptosis of human SCCs (A431, SCC4, SCC9, and SCC25) cells. The efficacy and safety issues were addressed regarding topical treatment of SR-T100 on UVB-induced cutaneous SCC of hairless mice and actinic keratoses (AKs) of human. RESULTS SR-T100 induces apoptosis in human SCCs cell lines by up-regulating the expressions of tumor necrosis factor receptors (TNFRs) and Fas, and downstream adaptors FADD/TRADD of the TNF-α and Fas ligand signaling cascades. SR-T100 also triggered the mitochondrial apoptotic pathway, as up-regulated cytochrome c and Bax, down-regulated Bcl-X(L). Animal experiments showed that all papillomas (35/35) and 27 of 30 UVB-induced microinvasive SCCs in hairless mice disappeared within 10 weeks after once-daily application of topical SR-T100. Furthermore, 13 patients, who suffered with 14 AKs, were treated with once-daily topical SR-T100 gel and 10 AKs cured after 16 weeks, showing negligible discomforts. CONCLUSION Our studies indicate that SR-T100 induces apoptosis of SCC cells via death receptors and the mitochondrial death pathway. The high efficacy of SR-T100 in our preclinical trial suggests that SR-T100 is a highly promising herb for AKs and related disorders.

Methylene blue-mediated photodynamic inactivation as a novel disinfectant of enterovirus 71

OBJECTIVES We tested whether methylene blue, an inexpensive and safe photosensitizer, is feasible for photodynamic inactivation of enterovirus 71 (EV71) in the environment. METHODS By escalating light doses and photosensitizer concentrations, photoinactivation of EV71 and other enteroviruses was examined in vitro. Viral transmission in the environment was simulated with a neonatal mouse model in vivo. Possible mechanisms were analysed with alterations of viral DNA and proteins after treatments. RESULTS Photodynamic inactivation of EV71 in suspensions occurred in a dose-dependent manner. The optimal condition for photoinactivating EV71 required a light dose of 200 J/cm(2) in the presence of methylene blue. This photodynamic condition was also able to inactivate other enteroviruses, including poliovirus 1 and coxsackieviruses A2, A3, A16 and B3. In an imitation environment, EV71 spread on a solid surface was inactivated by methylene blue-mediated photodynamic inactivation and prevented EV71 transmission to mice. Western blot and RT-PCR analysis indicated that both the viral proteins and the genome were disrupted after photodynamic inactivation. CONCLUSIONS Methylene blue-mediated photodynamic inactivation may provide a novel way to eliminate environmentally contaminated sources of EV71 to prevent infection.