David Good

Linkage of Paget disease of bone to a novel region on human chromosome 18q23.

Paget disease of bone (PDB) is characterized by increased osteoclast activity and localized abnormal bone remodeling. PDB has a significant genetic component, with evidence of linkage to chromosomes 6p21.3 (PDB1) and 18q21-22 (PDB2) in some pedigrees. There is evidence of genetic heterogeneity, with other pedigrees showing negative linkage to these regions. TNFRSF11A, a gene that is essential for osteoclast formation and that encodes receptor activator of nuclear factor-kappa B (RANK), has been mapped to the PDB2 region. TNFRSF11A mutations that segregate in pedigrees with either familial expansile osteolysis or familial PDB have been identified; however, linkage studies and mutation screening have excluded the involvement of RANK in the majority of patients with PDB. We have excluded linkage, both to PDB1 and to PDB2, in a large multigenerational pedigree with multiple family members affected by PDB. We have conducted a genomewide scan of this pedigree, followed by fine mapping and multipoint analysis in regions of interest. The peak two-point LOD scores from the genomewide scan were 2.75, at D7S507, and 1.76, at D18S70. Multipoint and haplotype analysis of markers flanking D7S507 did not support linkage to this region. Haplotype analysis of markers flanking D18S70 demonstrated a haplotype segregating with PDB in a large subpedigree. This subpedigree had a significantly lower age at diagnosis than the rest of the pedigree (51.2+/-8.5 vs. 64.2+/-9.7 years; P=.0012). Linkage analysis of this subpedigree demonstrated a peak two-point LOD score of 4.23, at marker D18S1390 (straight theta=0), and a peak multipoint LOD score of 4.71, at marker D18S70. Our data are consistent with genetic heterogeneity within the pedigree and indicate that 18q23 harbors a novel susceptibility gene for PDB.

Potential for Layered Double Hydroxides-Based, Innovative Drug Delivery Systems

Layered Double Hydroxides (LDHs)-based drug delivery systems have, for many years, shown great promises for the delivery of chemical therapeutics and bioactive molecules to mammalian cells in vitro and in vivo. This system offers high efficiency and drug loading density, as well as excellent protection of loaded molecules from undesired degradation. Toxicological studies have also found LDHs to be biocompatible compared with other widely used nanoparticles, such as iron oxide, silica, and single-walled carbon nanotubes. A plethora of bio-molecules have been reported to either attach to the surface of or intercalate into LDH materials through co-precipitation or anion-exchange reaction, including amino acid and peptides, ATPs, vitamins, and even polysaccharides. Recently, LDHs have been used for gene delivery of small molecular nucleic acids, such as antisense, oligonucleotides, PCR fragments, siRNA molecules or sheared genomic DNA. These nano-medicines have been applied to target cells or organs in gene therapeutic approaches. This review summarizes current progress of the development of LDHs nanoparticle drug carriers for nucleotides, anti-inflammatory, anti-cancer drugs and recent LDH application in medical research. Ground breaking studies will be highlighted and an outlook of the possible future progress proposed. It is hoped that the layered inorganic material will open up new frontier of research, leading to new nano-drugs in clinical applications.

Familial Paget's disease of bone: nonlinkage to the PDB1 and PDB2 loci on chromosomes 6p and 18q in a large pedigree.

Pagets disease of bone is a common condition characterized by bone pain, deformity, pathological fracture, and an increased incidence of osteosarcoma. Genetic factors play a role in the pathogenesis of Pagets disease but the molecular basis remains largely unknown. Susceptibility loci for Pagets disease of bone have been mapped to chromosome 6p21.3 (PDB1) and 18q21.1‐q22 (PDB2) in different pedigrees. We have identified a large pedigree of over 250 individuals with 49 informative individuals affected with Pagets disease of bone; 31 of whom are available for genotypic analysis. The disease is inherited as an autosomal dominant trait in the pedigree with high penetrance by the sixth decade. Linkage analysis has been performed with markers at PDB1; these data show significant exclusion of linkage with log10 of the odds ratio (LOD) scores < −2 in this region. Linkage analysis of microsatellite markers from the PDB2 region has excluded linkage with this region, with a 30 cM exclusion region (LOD score < −2.0) centered on D18S42. These data confirm the genetic heterogeneity of Pagets disease of bone. Our hypothesis is that a novel susceptibility gene relevant to the pathogenesis of Pagets disease of bone lies elsewhere in the genome in the affected members of this pedigree and will be identified using a microsatellite genomewide scan followed by positional cloning.

Neuroprotective effects of a standardized flavonoid extract from Diospyros kaki leaves

ETHNOPHARMACOLOGICAL RELEVANCE Flavonoids, extracted from the leaves of Diospyros kaki, are the main therapeutic components of NaoXingQing (NXQ), a potent and patented Chinese herbal remedy widely used in China for the treatment of apoplexy syndrome. AIM OF THE STUDY To investigate the neuroprotective effects of FLDK-P70, a standardized flavonoid extract, using in vivo rat models of both focal ischemia/reperfusion (I/R) injury induced by middle cerebral artery occlusion (MCAO) and on transient global brain ischemia induced by four-vessel occlusion (4-VO). We also aim to examine the effects of FLDK-P70 on glutamate-induced cell injury of hippocampal neurons as well as on hypoxia-induced injury of cortical neurons in primary cell culture. MATERIALS AND METHODS AND RESULTS Administration of FLDK-P70 for 12 days (40, 80 mg/kg body weight, p.o., 5 days before and 7 days after 4-VO) increased the survival of hippocampal CA1 pyramidal neurons after transient global brain ischemia. Similarly, administration of FLDK-P70 for 7 days (40, 80 mg/kg body weight, p.o., 3 days before and 4 days after MCAO) significantly reduced the lesion of the insulted brain hemisphere and improved the neurological behavior of rats. In primary rat hippocampal neuronal cultures, pretreatment with FLDK-P70 (5, 10 microg/ml) protected neurons from glutamate-induced excitotoxic neuronal death in a dose-dependent manner. In primary rat cerebral cortical neuronal culture, pretreatment with FLDK-P70 (25, 100 microg/ml) also reduced hypoxia-reoxygen induced neuronal death and apoptosis in a dose-dependent manner. CONCLUSIONS These in vivo and in vitro results suggest that FLDK-P70 significantly protects rats from MCAO and 4-VO ischemic injury in vivo and protects hippocampal neurons from glutamate-induced excitotoxic injury as well as cortical neurons from hypoxia-induced injury in vitro. The mechanisms of these effects may be due to the antioxidative activity of the flavonoids. These results convincingly demonstrate that FLDK-P70 may be useful for the prevention and treatment of ischemia/reperfusion injury and other related neurodegenerative diseases.

Apoptosis and microRNA aberrations in cancer

Carcinogenesis arises from the malfunction of genes that control cell growth and division. Therefore, the most effective method of hindering tumourigenesis is to induce the death of immortalized cancer cells. Apoptosis or programmed cell death has shown the most promises in impairing cancer growth. A variety of proteins is involved in the regulation of apoptosis and the malfunction of any these regulators may cause cell proliferation. The microRNAs have been shown to play a central role in the regulation of the cell cycle, including apoptosis. The microRNAs are involved in post‐transcriptional gene suppression and have been implicated in the regulation of cell differentiation and development. Aberrations in the microRNA regulation of apoptosis lead to tumourigenesis. The present review assesses the current knowledge of apoptotic regulation in cancer and the effect of microRNA aberrations in tumourigenesis.

Toad Glandular Secretions and Skin Extractions as Anti-Inflammatory and Anticancer Agents

Toad glandular secretions and skin extractions contain many natural agents which may provide a unique resource for novel drug development. The dried secretion from the auricular and skin glands of Chinese toad (Bufo bufo gargarizans) is named Chansu, which has been used in Traditional Chinese Medicine (TCM) for treating infection and inflammation for hundreds of years. The sterilized hot water extraction of dried toad skin is named Huachansu (Cinobufacini) which was developed for treating hepatitis B virus (HBV) and several types of cancers. However, the mechanisms of action of Chansu, Huachansu, and their constituents within are not well reported. Existing studies have suggested that their anti-inflammation and anticancer potential were via targeting Nuclear Factor (NF)-κB and its signalling pathways which are crucial hallmarks of inflammation and cancer in various experimental models. Here, we review some current studies of Chansu, Huachansu, and their compounds in terms of their use as both anti-inflammatory and anticancer agents. We also explored the potential use of toad glandular secretions and skin extractions as alternate resources for treating human cancers in combinational therapies.

Clostridial Spores for Cancer Therapy: Targeting Solid Tumour Microenvironment

Solid tumour accounts for 90% of all cancers. The current treatment approach for most solid tumours is surgery, however it is limited to early stage tumours. Other treatment options such as chemotherapy and radiotherapy are non-selective, thus causing damage to both healthy and cancerous tissue. Past research has focused on understanding tumour cells themselves, and conventional wisdom has aimed at targeting these cells directly. Recent research has shifted towards understanding the tumour microenvironment and its differences from that of healthy cells/tissues in the body and then to exploit these differences for treatmeat of the tumour. One such approach is utilizing anaerobic bacteria. Several strains of bacteria have been shown to selectively colonize in solid tumours, making them valuable tools for selective tumour targeting and destruction. Amongst them, the anaerobic Clostridium has shown great potential in penetration and colonization of the hypoxic and necrotic areas of the tumour microenvironment, causing significant oncolysis as well as enabling the delivery of therapeutics directly to the tumour in situ. Various strategies utilizing Clostridium are currently being investigated, and represent a novel area of emerging cancer therapy. This review provides an update review of tumour microenvironment as well as summary of the progresses and current status of Clostridial spore-based cancer therapies.

The Mechanisms of Chansu in Inducing Efficient Apoptosis in Colon Cancer Cells

Chansu is one of the most widely used traditional Chinese medicines in China, Japan, and other Southeast Asian countries primarily for antipain, anti-inflammation, and recently anticancer. Over 10 recipes and remedies contained Chansu, which are easily available in pharmacies and hospitals, but the mechanisms of action were not clearly articulated. In the present study, Cinobufagin (CBF), the major compound of Chansu, was employed as a surrogate marker to determine its ability in inducing cancer cell death. As expected, CBF has significant cancer-killing capacity for a range of cancers, but such ability differs markedly. Colon and prostate cancers are more sensitive than skin and lung cancers. Interestingly, cancer cells die through apoptotic pathway either being biphasic caspase-3-dependent (HCT116) or independent (HT29). Multipathway analysis reveals that CBF-induced apoptosis is likely modulated by the hypoxia-inducing factor-1 alpha subunit (HIF-1α) as its inhibition was evident in vitro and in vivo. Taken together, these results demonstrate that CBF is a potent apoptotic inducer with potential for further development as a novel and effective anticancer agent for a range of cancers, especially colon cancer.

Identification of Chlamydia pneumoniae proteins in the transition from reticulate to elementary body formation.

Chlamydia pneumoniae is an important human respiratory pathogen that is responsible for an estimated 10% of community-acquired pneumonia and 5% of bronchitis and sinusitis cases. We examined changes in global protein expression profiles associated with the redifferentiation of reticulate body (RB) to elementary body (EB) as C. pneumoniae cells progressed from 24 to 48 h postinfection in HEp2 cells. Proteins corresponding to those showing the greatest changes in abundance in the beginning of the RB to EB transition were then identified from purified EBs. Among the 300 spots recognized, 35 proteins that were expressed at sufficiently high levels were identified by mass spectrometry. We identified C. pneumoniae proteins that showed more than 2-fold increases in abundance in the early stages of RB to EB transition, including several associated with amino acid and cofactor biosynthesis (Ndk, TrxA, Adk, PyrH, and BirA), maintenance of cytoplasmic protein function (GroEL/ES, DnaK, DksA, GrpE, HtrA, ClpP, ClpB, and Map), modification of the bacterial cell surface (CrpA, OmpA, and OmcB), energy metabolism (Tal and Pyk), and the putative transcriptional regulator TctD. This study identified C. pneumoniae proteins involved in the process of redifferentiation into mature, infective EBs and indicates bacterial metabolic pathways that may be involved in this transition. The proteins involved in RB to EB transition are key to C. pneumoniae infection and are perhaps suitable targets for therapeutic intervention.

Efficient c-kit Receptor-Targeted Gene Transfer to Primary Human CD34-Selected Hematopoietic Stem Cells

ABSTRACT We have previously reported effective gene transfer with a targeted molecular conjugate adenovirus vector through the c-kit receptor in hematopoietic progenitor cell lines. However, a c-kit-targeted recombinant retroviral vector failed to transduce cells, indicating the existence of significant differences for c-kit target gene transfer between these two viruses. Here we demonstrate that conjugation of an adenovirus to a c-kit-retargeted retrovirus vector enables retroviral transduction. This finding suggests the requirement of endosomalysis for successful c-kit-targeted gene transfer. Furthermore, we show efficient gene transfer to, and high transgene expression (66%) in, CD34-selected, c-kit+ human peripheral blood stem cells using a c-kit-targeted adenovirus vector. These findings may have important implications for future vector development in c-kit-targeted stem cell gene transfer.