Yu-Tsueng Liu

Network-based classification of breast cancer metastasis

Mapping the pathways that give rise to metastasis is one of the key challenges of breast cancer research. Recently, several large‐scale studies have shed light on this problem through analysis of gene expression profiles to identify markers correlated with metastasis. Here, we apply a protein‐network‐based approach that identifies markers not as individual genes but as subnetworks extracted from protein interaction databases. The resulting subnetworks provide novel hypotheses for pathways involved in tumor progression. Although genes with known breast cancer mutations are typically not detected through analysis of differential expression, they play a central role in the protein network by interconnecting many differentially expressed genes. We find that the subnetwork markers are more reproducible than individual marker genes selected without network information, and that they achieve higher accuracy in the classification of metastatic versus non‐metastatic tumors.

Maintenance of colonic homeostasis by distinctive apical TLR9 signalling in intestinal epithelial cells

The mechanisms by which commensal bacteria suppress inflammatory signalling in the gut are still unclear. Here, we present a cellular mechanism whereby the polarity of intestinal epithelial cells (IECs) has a major role in colonic homeostasis. TLR9 activation through apical and basolateral surface domains have distinct transcriptional responses, evident by NF-κB activation and cDNA microarray analysis. Whereas basolateral TLR9 signals IκBα degradation and activation of the NF-κB pathway, apical TLR9 stimulation invokes a unique response in which ubiquitinated IκB accumulates in the cytoplasm preventing NF-κB activation. Furthermore, apical TLR9 stimulation confers intracellular tolerance to subsequent TLR challenges. IECs in TLR9-deficient mice, when compared with wild-type and TLR2-deficient mice, display a lower NF-κB activation threshold and these mice are highly susceptible to experimental colitis. Our data provide a case for organ-specific innate immunity in which TLR expression in polarized IECs has uniquely evolved to maintain colonic homeostasis and regulate tolerance and inflammation.

Antibodies Elicited by Inactivated Propionibacterium acnes-Based Vaccines Exert Protective Immunity and Attenuate the IL-8 Production in Human Sebocytes: Relevance to Therapy for Acne Vulgaris

Propionibacterium acnes is a key pathogen involved in the progression of inflammation in acne vulgaris. We examined whether vaccination against P. acnes suppressed P. acnes-induced skin inflammation. Inactivation of P. acnes with heat was employed to create a P. acnes-based vaccine. Intranasal immunization in mice with this inactivated vaccine provoked specific antibodies against P. acnes. Most notably, immunization with inactivated vaccines generated in vivo protective immunity against P. acnes challenge and facilitated the resolution of ear inflammation in mice. In addition, antibodies elicited by inactivated vaccines effectively neutralized the cytotoxicity of P. acnes and attenuated the production of proinflammatory cytokine IL-8 in human sebocyte SZ95 cells. Intranasal immunization using heat-inactivated P. acnes-based vaccines provided a simple modality to develop acne vaccines. These observations highlight the concept that development of vaccines targeting microbial products may represent an alternative strategy to conventional antibiotic therapy.

Vaccination Targeting a Surface Sialidase of P. acnes: Implication for New Treatment of Acne Vulgaris

Background Acne vulgaris afflicts more than fifty million people in the United State and the severity of this disorder is associated with the immune response to Propionibacterium acnes (P. acnes). Systemic therapies for acne target P. acnes using antibiotics, or target the follicle with retinoids such as isotretinoin. The latter systemic treatment is highly effective but also carries a risk of side effects including immune imbalance, hyperlipidemia, and teratogenicity. Despite substantial research into potential new therapies for this common disease, vaccines against acne vulgaris are not yet available. Methods and Findings Here we create an acne vaccine targeting a cell wall-anchored sialidase of P. acnes. The importance of sialidase to disease pathogenesis is shown by treatment of a human sebocyte cell line with recombinant sialidase that increased susceptibility to P. acnes cytotoxicity and adhesion. Mice immunized with sialidase elicit a detectable antibody; the anti-sialidase serum effectively neutralized the cytotoxicity of P. acnes in vitro and P. acnes-induced interleukin-8 (IL-8) production in human sebocytes. Furthermore, the sialidase-immunized mice provided protective immunity against P. acnes in vivo as this treatment blocked an increase in ear thickness and release of pro-inflammatory macrophage inflammatory protein (MIP-2) cytokine. Conclusions Results indicated that acne vaccines open novel therapeutic avenues for acne vulgaris and other P. acnes-associated diseases.

Staphylococcus aureus Hijacks a Skin Commensal to Intensify Its Virulence: Immunization Targeting β-Hemolysin and CAMP Factor

The need for a new anti-Staphylococcus aureus therapy that can effectively cripple bacterial infection, neutralize secretory virulence factors, and lower the risk of creating bacterial resistance is undisputed. Here, we propose what is, to our knowledge, a previously unreported infectious mechanism by which S. aureus may commandeer Propionibacterium acnes, a key member of the human skin microbiome, to spread its invasion and highlight two secretory virulence factors (S. aureus β-hemolysin and P. acnes CAMP (Christie, Atkins, Munch-Peterson) factor) as potential molecular targets for immunotherapy against S. aureus infection. Our data demonstrate that the hemolysis and cytolysis by S. aureus were noticeably augmented when S. aureus was grown with P. acnes. The augmentation was significantly abrogated when the P. acnes CAMP factor was neutralized or β-hemolysin of S. aureus was mutated. In addition, the hemolysis and cytolysis of recombinant β-hemolysin were markedly enhanced by recombinant CAMP factor. Furthermore, P. acnes exacerbated S. aureus-induced skin lesions in vivo. The combination of CAMP factor neutralization and β-hemolysin immunization cooperatively suppressed the skin lesions caused by coinfection of P. acnes and S. aureus. These observations suggest a previously unreported immunotherapy targeting the interaction of S. aureus with a skin commensal.

A technological update of molecular diagnostics for infectious diseases

Identification of a causative pathogen is essential for the choice of treatment for most infectious diseases. Many FDA approved molecular assays; usually more sensitive and specific compared to traditional tests, have been developed in the last decade. A new trend of high throughput and multiplexing assays are emerging thanks to technological developments for the human genome sequencing project. The applications of microarray and ultra high throughput sequencing technologies for diagnostic microbiology are reviewed. The race for the

The essentiality of α‐2‐macroglobulin in human salivary innate immunity against new H1N1 swine origin influenza A virus

1000 genome technology by 2014 will have a profound impact in diagnosis and treatment of infectious diseases in the near future.

Optimization of primer design for the detection of variable genomic lesions in cancer

A novel strain of influenza A H1N1 emerged in the spring of 2009 and has spread rapidly throughout the world. Although vaccines have recently been developed that are expected to be protective, their availability was delayed until well into the influenza season. Although anti‐influenza drugs such as neuraminidase inhibitors can be effective, resistance to these drugs has already been reported. Although human saliva was known to inhibit viral infection and may thus prevent viral transmission, the components responsible for this activity on influenza virus, in particular, influenza A swine origin influenza A virus (S‐OIV), have not yet been defined. By using a proteomic approach in conjunction with beads that bind α‐2,6‐sialylated glycoprotein, we determined that an α‐2‐macroglobulin (A2M) and an A2M‐like protein are essential components in salivary innate immunity against hemagglutination mediated by a clinical isolate of S‐OIV (San Diego/01/09 S‐OIV). A model of an A2M‐based “double‐edged sword” on competition of α‐2,6‐sialylated glycoprotein receptors and inactivation of host proteases is proposed. We emphasize that endogenous A2M in human innate immunity functions as a natural inhibitor against S‐OIV.

A Novel Approach for Determining Cancer Genomic Breakpoints in the Presence of Normal DNA

Primer approximation multiplex PCR (PAMP) is a new experimental protocol for efficiently assaying structural variation in genomes. PAMP is particularly suited to cancer genomes where the precise breakpoints of alterations such as deletions or translocations vary between patients. The design of PCR primer sets for PAMP is challenging because a large number of primer pairs are required to detect alterations in the hundreds of kilobases range that can occur in cancer. These sets of primers must achieve high coverage of the region of interest, while avoiding primer dimers and satisfying the physico-chemical constraints of good PCR primers. We describe a natural formulation of these constraints as a combinatorial optimization problem. We show that the PAMP primer design problem is NP-hard, and design algorithms based on simulated annealing and integer programming, that provide good solutions to this problem in practice. The algorithms are applied to a test region around the known CDKN2A deletion, which show excellent results even in a 1:49 mixture of mutated:wild-type cells. We use these test results to help set design parameters for larger problems. We can achieve near-optimal designs for regions close to 1 Mb.

Porphyrin Metabolisms in Human Skin Commensal Propionibacterium acnes Bacteria: Potential Application to Monitor Human Radiation Risk

CDKN2A (encodes p16INK4A and p14ARF) deletion, which results in both Rb and p53 inactivation, is the most common chromosomal anomaly in human cancers. To precisely map the deletion breakpoints is important to understanding the molecular mechanism of genomic rearrangement and may also be useful for clinical applications. However, current methods for determining the breakpoint are either of low resolution or require the isolation of relatively pure cancer cells, which can be difficult for clinical samples that are typically contaminated with various amounts of normal host cells. To overcome this hurdle, we have developed a novel approach, designated Primer Approximation Multiplex PCR (PAMP), for enriching breakpoint sequences followed by genomic tiling array hybridization to locate the breakpoints. In a series of proof-of-concept experiments, we were able to identify cancer-derived CDKN2A genomic breakpoints when more than 99.9% of wild type genome was present in a model system. This design can be scaled up with bioinformatics support and can be applied to validate other candidate cancer-associated loci that are revealed by other more systemic but lower throughput assays.