Continuous exposure to UV irradiation (both UV-A and UV-B) leads to the skin cancer and other photoaging complications, which are typically mediated by the reactive oxygen species (ROS), generated in the oxidative pathways. Many of the anti-oxidative and anti-photoaging compounds have been identified previously, which work efficiently against photodamage of the skin. Recently, marine originated photoprotective or anti-photoaging behavior was observed in the methanol extracts of Corallina pilulifera (CPM). These extracts were found to exert potent antioxidant activity and protective effect on UV-A-induced oxidative stress in human dermal fibroblast (HDF) cells by protecting DNA and also by inhibiting matrix metalloproteinases (MMPs), a key component in photoaging of the skin due to exposure to UV-A. The present review depicts various other photoprotective compounds from algae and other marine sources for further elaborative research and their probable use in cosmeceutical and pharmaceutical industries.Marine organisms form a prominent component of the oceanic population, which significantly contribute in the production of cosmeceutical and pharmaceutical molecules with biologically efficient moieties. In addition to the molecules of various biological activities like anti-bacterial, anti-cancerous, anti-inflammatory and anti-oxidative  Continuous exposure to UV light leads to numerous complications that are correlated with various pathological consequences of the skin damage and sunburn occurs when exposure to UV light exceeds the protective capacity of an individual’s melanin ,2. UV irisorders ,9.Several investigations have revealed that collagen gets degraded in photoaged skin due to the inhibition of collagen synthesis mediated by matrix metalloproteinases (MMPs), which are a family of secreted or transmembrane zinc endopeptidases that are capable of digesting ECM. MMPs are divided into subclasses of collagenases, gelatinases, stromelysins, matrilysins and membrane-type MMPs (MT-MMPs) according to their substrate specificity and domain structure . It is ret al. developed a database that contains information on the photoprotective compounds such as MAAs, scytonemin and other unidentified compounds that absorb in the UV range, which are reported in aquatic organisms like cyanobacteria, phytoplankton and macroalgae of diverse habitats at different collection times [Photoprotection is a group of mechanisms that nature has developed to minimize the damages that an organism suffers, when exposed to UV radiation. These mechanisms can be controlled or organized by certain organic and inorganic compounds or substances  produced by different terrestrial and aquatic sources. A number of photo protective compounds such as scytonemins (exclusively in cyanobacteria), mycosporines (in fungi), mycosporine-like amino acids , phenyl propanoids and flavonoids (higher plants), melanins , and several other UV-absorbing substances of unknown chemical structures from different organisms have been developed to counteract the photodamage ,21. Nearon times .The induction of cellular phototoxicity in response to UV radiation has previously been demonstrated in different cell lines –25. Recea from Eisenia bicyclis, fucoxanthin from Hijikia fusiformis and phlorotannin from Ecklonia stolonifera [More than 12,000 novel chemicals from marine plants, animals, and microbes have been explored, and a large number of pharmaceutical products, enzymes and biomaterials have been developed until date. Few of the important marine derived photoprotective compounds are presented in lonifera –31. PresMarine algae have been an important source to produce a variety of secondary metabolites including phenols and polyphenols with unique linkages . From paet al. demonstrated that methanol extracts of alga Corallina pilulifera (CPM) possess high phenolic content, which reduces the expression of UV-induced MMP-2 and -9 in human dermal fibroblast (HDF) cells in dose dependent manner, there by attaining the capability of inhibiting free radicals [Eckloina cava against the photo-oxidative stress induced by UV-B radiation via DCFH-DA, MTT, comet assay, and microscopic analysis [E. Cava can be used as an effective natural source to make cosmeceutical or pharmaceutical products.Recently, Ryu analysis . AccordiSargassum sagamianum suggested to be effective therapeutic agents in protection against UV-B [in vitro and in vivo by the activation of caspases. These same compounds from S. macrocarpum were found to have resulted in neurite outgrowth and supporting the survival of neuronal PC12D cell [et al. documented the cutaneous photoprotective ability of the high UV-absorbing MAAs, Porphyra-334 and shinorine (P-334 + SH), isolated from the red alga Porphyra rosengurttii, by in vivo procedures on mouse skin [Sargassum siliquastrum can also produce xanthin compounds and more recently, Heo and Jeon isolated fucoxanthin, a natural carotenoid that exerts a potential photoprotective effect against UV-B induced cell damage [Since a combination of phototherapy using UV-B with topical reagents has been clinically applied to treat hyperproliferative skin disease; sargaquinoic acid and sargachromenol from nst UV-B . As per 12D cell . Later, use skin . They hause skin . Their sl damage .in vivo and among them; MMP1 and MMP3 are thought to be the two major contributors to photoaging [As discussed earlier, skin damage by reactive oxygen species (ROS) leads to the activation of Protein kinase C (PKC), thus increasing the expression of matrix metalloproteinases (MMPs) and degradation of collagen. UVB is known to induce the expressions of MMP1, MMP3, and MMP9 in the normal human epidermis otoaging ,43. Hencet al. have reported the occurrence of many UV-A and UV-B-absorbing compounds in 206 strains of 152 marine microalgal species [Algal phytoplanktons have tremendous impact on the sustainability of the marine ecosystem by being a food source to other marine fauna and by possessing the photoprotecting principle to protect the other inhabitants and algal consumers of marine environment. Jeffrey  species .Thalassiosira weissflogii is a common species of coastal marine phytoplankton, which might be forming extensive, non-toxic occasional blooms during early summer in the upper layers of the seawater [Marine plants are majorily (around 90%) occupied by the phytoplanktonic species and nowadays especially, diatoms are attracting the drug science , because of their vast species diversity and distribution all over the world . The diaseawater . It suggseawater .Diatoms often dominate the algal blooms during spring and fall at temperate latitudes and in summer in Polar Regions, which show a wide range of response to UV exposure. Centric diatoms seem more UV-resistant than pennate diatoms, after a suitable acclimation period . HoweverCollema cristatum [et al. later identified two different mycosporines and three unidentified UV-absorbing compounds in fungal isolates from hyper-saline waters and polar glacial ice [UV-B absorbing mycosporines with photoprotective activity are present day targets from fugal species e.g., lichenized ascomycete, ristatum . The purcial ice .et al. have evaluated the effect of two lichen compounds, sphaerophorin (depside) and pannarin (depsidone) on pBR322 DNA cleavage induced by hydroxyl radicals (•OH), and by nitric oxide (NO), and their superoxide anion (O2−) scavenging capacity [Extensive studies on the secondary metabolites of lichens have led to the isolation of many novel substances, which by number are over 800 ,55. Thescapacity . These cet al. have analysed the light-filtering power of natural substances like boldine, a major alkaloid found in the leaves and bark of boldo tree Peumus boldus (Molina), and other aromatic compounds  extracted from Chilean lichens [tert.-butyl-4′-methoxy dibenzoylmethane (BM-DBM).Rancan  lichens . These cet al. recently described the bacterial (Klebsiella aerogenes) photoprotection through extracellular cadmium sulfide crystallites (CdS), where these semiconductor particles absorb radiation in the UV spectral region [K. aerogenes produces extracellular CdS material in response to the stressed environments containing cadmium ions, a photoprotective layer is formed.Many sunscreen/cosmetic compositions have been discovered from bacteria, which have been adopted for the photoprotection of human skin and/or hair, because of their underlying anti-photoaging principle. More interestingly, bacterial melanin, an active photoprotecting pigment protects against the DNA damage under full UV-B irradiation . The bacl region . Hence, Bacteria, especially the ones like archaea and other extremophiles species have tremendous implication for the survival in extraterrestrial habitats and are very advantageous in present day astrobiological research for the detection of the protectant biomolecules .Cyanobacteria are phylogenetically a primitive group of Gram-negative prokaryotes that possess higher-plant-type oxygenic photosynthesis . As theyCalothrix sp.; a naturally occurring cyanobacterium [There are a number of adaptation strategies, by which cyanobacteria try to avoid high white light and ultraviolet radiation stress. These adaptations range from migration into habitats of reduced light exposure along with phototactic, photokinetic and photophobic responses ; and veracterium . It was Chlamys farreri and its possible protective role against UV-B induced apoptosis in murine thymocytes has been investigated [et al. has depicted the role of this polypeptide from C. farreri in the inhibition of UVA induced apoptotic pathway in human HaCaT keratinocytes [Marine animals are largely unexplored for the production of biologically active secondary metabolites. A new marine polypeptide (molecular mass 879) has been recently isolated from the Chinese scallop stigated . This stinocytes . The molinocytes .Holothuria atra (Jaeger) and their probable role in photoprotection has been hypothesized. It is believed that MAAs could probably function as broad-spectrum UV absorbers [Patinopecten yessoensis in 2008 by Oyamada et al. [Presence of MAAs has also been reported in the black sea cucumber bsorbers . UV absoa et al. . It was Aplysia dactylomela by Carefoot et al., where the role of MAAs as protective sunscreens was investigated, but the significant mechanisms and commercial prospectives are still open for discussion [In certain cases, the debate is still on to determine photoprotective compounds and their sure role in photoprotection in marine organisms. One such example is the study on spawn of the sea hare scussion .in-situ and ex-situ approaches of marine floral and faunal culture for the development of novel photoprotective molecules, by considering the global warming and other catastrophic causes of ozone destruction.UV irradiation is a present day concern, as the impact of global warming and other climatic catastrophic factors are severely effecting the biological population. Hence, photoprotection is a major biological concern with respect to the source of natural bioactive molecules that have the anti-photoaging effect and especially, the safer marine sources have been identified in past few decades. The advances in molecular biology and culturing technologies are bridging the gap between the challenges pertaining to the exploitation of marine environment as a potential source of natural protective compounds. As presented in the current review, various compounds which support potentially the photoprotective mechanisms of strong cosmeceutical and pharmaceutical value, have already been isolated from different marine sources like algae, fungi, lichens, bacteria, phytoplanktons, animals and plants. Several marine based pharmaceuticals are under active commercial development, as the ecosystem health is high on the public’s list of concerns, and hence, aquaculture is providing an ever-greater proportion of the seafood in developed countries. An extensive research still has a major role to play to uncover the enormousity of the vast marine sources in terms of photoprotective compounds that promote DNA protection by reducing oxidative stress after UV exposure. This would develop a proper understanding of the In the light of present review, it is worthwhile to state that anti-photoaging and photoprotective natural products research from such deceptively simple marine organisms will pave a way for newer and novel products, which find immense use in the pharmaceutical and cosmeceutical industry.


Enzymes which traditionally have played no role in cell-directed cytotoxicity are finding their way into schemes for prodrug activation and immunotoxins owing to such useful enzymatic activity. Alkaline phosphatase, carboxypeptidases, beta-glucosidases and beta-lactamases among many others are being utilised to regenerate potent anti-cancer drugs or toxic small molecules from precursors in a bid to enhance their activity in tumours. These prodrug activation systems require the pretargeting of the enzyme to the surface of a tumour cell, usually by an antibody or its immunoreactive fragment. A recent novel approach proposes the intracellular delivery of appropriate enzymes, such as phosphodiesterases, to particular cellular compartments. There, enzyme activity can cause substantive damage resulting in cell death. Cell targeting of mammalian phosphodiesterase promises to improve upon conventional immunotoxins because of their increased cytotoxicity when targeted to the appropriate compartment and their expected lack of, or lower, immunogenicity in clinical use.


Biomarkers are pivotal for cancer detection, diagnosis, prognosis and therapeutic monitoring. However, currently available cancer biomarkers have the disadvantage of lacking specificity and/or sensitivity. Developing effective cancer biomarkers becomes a pressing and permanent need. The cancer secretome, the totality of proteins released by cancer cells or tissues, provides useful tools for the discovery of novel biomarkers. The focus of this article is to review the recent advances in cancer secretome analysis. We aim to elaborate the approaches currently employed for cancer secretome studies, as well as its applications in the identification of biomarkers and the clarification of carcinogenesis mechanisms. Challenges encountered in this newly emerging field, including sample preparation, in vivo secretome analysis and biomarker validation, are also discussed. Further improvements on strategies and technologies will continue to drive forward cancer secretome research and enable development of a wealth of clinically valuable cancer biomarkers. Cancer remains the major devastating disease throughout the world. It is estimated that cancers are responsible for over 6 million lives per year worldwide with an annual 10 million or more new cases. In developing countries, cancers are the second most common cause of death, which comprise 23–25% of total mortality. Despite advances in diagnostic imaging technologies, surgical management, and therapeutic modalities, the long-term survival is poor in most cancers. For example, the five-year survival rate is only 14% in lung cancer and 4% in pancreatic cancer .Over the past several decades, enormous efforts have been made to screen and characterize useful cancer biomarkers. Some important molecules including carcinoembryonic antigen (CEA), prostate specific antigen (PSA), alpha-fetoprotein (AFP), CA 125, CA 15-3 and CA 19-9, have been identified. They are commonly employed in clinical diagnosis. Unfortunately, most biomarkers are not satisfactory because of their limited specificity and/or sensitivity . TherefoCurrently, we are in an era of molecular biology and bioinformatics. Many novel approaches have been introduced to identify markers associated with cancer. Proteomic profiling is one of the most commonly applied strategies for cancer biomarker discovery. There are two general differential proteomic strategies: comparing protein patterns in cancer tissue with their normal counterparts, and comparing plasma/serum from cancer patients with those from normal controls. As suggested by Liotta : "the blGiven the above-mentioned major limitations in blood proteomics, scientists are seeking other methods for cancer biomarker discovery. The term "secretome" was first proposed by Tjalsma et al.  in a genIn recent years, the emerging technologies in life science, especially that of proteomic research, have greatly accelerated studies on the cancer secretome. Generally, these methods can be categorized into two groups, namely genome-based computational prediction and proteomic approaches.These approaches are characterized by a combined method of transcript profiling and computational analysis. Computational analysis depends on the prediction of signal peptides, which is viewed as a hallmark of classically secreted proteins. According to the famous signal hypothesis , the majThese genome-based methods can provide a comprehensive list of potentially secreted proteins quickly. However, there are two major inherent problems that restrain the broad use of these approaches. First, this approach relies on prediction of signal peptides or cell retention signals, thus making some genuine secreted proteins lacking signal peptide or presenting cell retention signals unpredictable. About 50% of secreted proteins can be predicted by signal peptides or other specific cell retention signals . Second,Nowadays, proteomic technologies are the mainstay of cancer secretome studies. With the massive progress in mass spectrometry (MS), bioinformatics and analytical techniques, proteomic approaches greatly promote the cancer secretome analysis and biomarker discovery. Currently, there are roughly three major proteomic technologies in secretome researches: gel-based methods, gel-free MS-based methods and surface-enhanced laser desorption/ionization time-of-flight mass spectrometric (SELDI-TOF-MS).Two-dimensional gel electrophoresis (2-DE) coupling MS is the most classic and well-established proteomic approach. This method allows the separation of complex mixtures of intact proteins at high resolution. These protein mixtures are first separated according to their charge in the first dimension by isoelectric focusing (IEF) and size in the second dimension by SDS-PAGE, and then analyzed by peptide mass fingerprinting using MS or MS/MS after in-gel trypsin digestion. It has been widely used in secretome studies of cancers, such as malignant glioma , lung caAlthough 2-DE currently remains the most efficient method for separation of complex protein mixtures, it is clear that this technique has several disadvantages, including poor reproducibility between gels, low sensitivity in the detection of proteins in low concentrations and hydrophobic membrane proteins, limited sample capacity and low linear range of visualization procedures . In addiTo circumvent some of these inherent problems of the standard 2-DE procedure, a modified method, differential in-gel electrophoresis (DIGE) has been developed by GE Healthcare . This teTo overcome the inherent drawbacks of gel-based approaches, great efforts have been made recently on gel-free MS-based or shotgun proteomics. In these newly emerging approaches, instead of depending on gels to separate and analyze proteins, complex mixtures of proteins are first digested into peptides or peptide fragments, then separated by one or several steps of capillary chromatography, and finally analyzed by MS/MS. Multidimensional protein identification technology (MudPIT), which was introduced and termed by Yates and colleague , is one 13C or 12C isotopes [The major progress in proteome/secretome study is the technology of quantitative proteomics which introduced isotopes or other molecular labeling methods in proteomic analysis -49. In tisotopes ,54. Thisisotopes . Howeverisotopes .Stable isotope labeling by amino acids in cell culture (SILAC) is another common stable isotope labeling technique. In SILAC, stable isotope-labeled essential amino acids are added to amino acid deficient cell culture media, and then are absorbed and secreted by cells in the synthesis of proteins in vitro. Thus the proteome from different cell cultures can be compared as being grown in media with carbon-isotopically modified amino acids. A differential SILAC secretome study between pancreatic cancer cells and non-neoplastic pancreatic ductal cells identified 145 differentially secreted proteins (> 1.5-fold change), including several common biomarkers of pancreatic cancer and novel proteins that have not been reported previously . Nearly Isobaric tag for relative and absolute quantization (iTRAQ) is a recently developed isotope labeling approach that is increasingly accepted in secretome analysis . This neSELDI-TOF-MS is an exciting approach in cancer proteomics, particularly plasma proteomics -61. The The major application of cancer secretome analysis is to search for cancer biomarkers. As mentioned above, the cancer secretome contains a treasure trove of novel biomarkers, which make cancer diagnosis using secretome markers attractive. Recently, investigation of secretomes from a variety of cancers has led to the identification of a number of potential cancer biomarkers Table . It is kAs shown in table There is a growing consensus that no single cancer biomarker is sensitive and specific enough to meet stringent diagnostic criteria given the substantial heterogeneity among cancers. A feasible strategy to circumvent the drawbacks of individual markers is to measure a combination of proteomic biomarkers. To get panels of serum biomarkers for lung cancer detection, Xiao et al  comparedTaken together, these studies demonstrate that secretome analysis is a feasible and efficient method to find, identify, and characterize clinical relevant biomarkers.In addition to the identification of candidate biomarkers, cancer secretome analysis can provide new insights into the molecular mechanisms of carcinogenesis. Extracellular events such as cell-to-cell interactions and cell-to-extracellular matrix interactions are crucial during carcinogenesis. To characterize extracellular events associated with breast cancer progression, secreted protein-encoded gene expression profiles were investigated in a cell line model of human proliferative breast disease (PBD). Differentially expressed genes from microarray data were searched for genes encoding secreted proteins in three public databases. The analysis displayed two clusters of secretome genes with expression changes correlating with proliferative potential, implicating a role in breast cancer progression . In a reDifferential cancer secretome analysis can also advance our understanding on the functions of interesting genes. It is known that tumor-suppressive p21 is a negative regulator of cell cycle progression; however, several studies have shown that p21 expression in tumor cells mediates an anti-apoptotic and mitogenic paracrine effect ,76. In oTo gain reliable insights into the cancer secretome, it is first necessary to prepare samples for analysis which are as pure as possible. Secreted proteins in vivo occur in body fluids, thus the direct analysis for them is hindered by the high complexity. It is generally accepted that proteins secreted by tumor cells in vitro may, to some extent, reflect the proteins released by tumors in vivo. Therefore, the routine method used to date is to obtain secreted proteins from the media of in vitro cancer cell cultureFigure .Although cells are commonly cultivated in serum-supplemented media, serum-free media (SFM) are needed to guarantee the successful analysis of the cancer secretome in vitro. The reason lies in the fact that the highly abundant serum proteins such as albumin may mask and dilute the secretome, whereas cell growth is much slower in SFM, and these cells tend to autolyse and liberate cytosolic proteins. Mbeunkui et al  performeIn consideration of the significant masking effects of bovine serum albumin (BSA) and other serum constituents, washing the cells thoroughly to reduce serum contaminations before incubation in SFM is a necessary step, whereas stringent washes can damage or kill the cells and lead to the nonspecific liberation of cytoplasmic proteins. Thus, how to keep a balance between serum contaminations removal by washing and cell survival is the key. Pellitteri-Hahn et al  used ratThere is no doubt that optimizing the cell culture conditions and employing an appropriate washing technology can significantly reduce serum or cytosolic protein contamination. Nevertheless, some serum constituents are still present in culture media even after thorough rinsing treatment, and even under optimum culture conditions, cell cultivation in vitro is unavoidably accompanied by cell death and subsequent release of cytosolic proteins. Because the concentration of secreted proteins is always very low, the contamination by non-secreted proteins may easily mask the proteins of interest. Consequently, how to discriminate genuine secreted proteins from non-secreted proteins is a major question that remains to be answered. Zwickl et al  have estSecreted proteins present in the culture media are usually in low concentrations, which can go down to the ng/mL range, as in the case of some cytokines. Thus, proteins secreted in the culture media should be concentrated before subsequent proteomics analysis. Various methods have been used to concentrate the proteins; nonetheless, these methods are not all well suited for the secretome analysis. For example, precipitation with acetone can not concentrate large volumes of culture medium because a minimum five-fold volume excess of acetone should be used, and dye precipitation selects against an important class of secreted proteins – the proglycoproteins . Among tCurrently, most studies on the cancer secretome involve collecting secreted proteins from supernatants of cancer cell lines cultivated in vitro and then analyzing their properties in vivo. Nevertheless, the in vitro cell culture systems are far from physiological situations. Then, the question is whether the in vitro cell culture systems are able to completely replicate the in vivo conditions, or whether the data from in vivo secretome can match well with that achieved in vitro. Considering the great challenges for obtaining pure secretome, to date, only a minority of studies have investigated cancer secretome under in vivo situations. Varnum et al  characteFor achieving reliable and clinically worthwhile biomarkers, the interesting protein markers discovered from the cancer secretome need to be further validated. To some extent, validation is more arduous than discovery , and theAnalysis and characterization of a cancer secretome is a critical step towards the biomarker discovery process, which represents a challenge for current technologies. Though genome-based approaches are convenient and comprehensive, the accuracy for predicting secreted proteins is always far from satisfactory owing to the inherent drawbacks. Furthermore, there is always a discrepancy between the expression levels of mRNA and the corresponding secreted proteins. For allowing direct analysis for secreted proteins, proteomic methods are considered as a more powerful means to investigate the cancer secretome. While classic gel-based proteomic technologies have produced significant contributions to biomarker discovery, the emergence of gel-free MS-based proteomic approaches, such as MudPIT and SELDI-TOF-MS, greatly facilitates the secretome analysis with increased sensitivity and automation. Proteomic approaches currently used are not as rapid and high-throughput as genomic profiling with microarrays – hence improving proteomic methods towards higher comprehensiveness, throughput, reproducibility and accuracy is of vital importance. Considering genomic-based and proteomic approaches provide closely related but distinct information about the cancer secretome, they can be combined as complementary methods. Searching for biomarkers from cancer secretome analysis also challenges bioinformatics, which needs to cope with the vast amounts of data from MS. To gain more reliable insights into the cancer secretome and develop valuable cancer biomarkers, the optimization of sample preparation procedure should be fully established, and more efforts should be focused on in vivo secretome research and biomarker validation. Overall, investigating the cancer secretome opens up new avenues in the search for clinically worthwhile biomarkers. With the rapid development of new strategies and technologies, this newly emerging field will reveal more valuable information on cancer diagnosis, monitoring and therapy.The authors declare that they have no competing interests.HX wrote the manuscript. BJL edited the manuscript. MDL organized and revised the manuscript. All authors read and approved the final manuscript.


During meiosis, the template for homologous recombination is not chosen at random as described in this Primer by Jessica Lao and Neil Hunter. Connections between chromosomes are essential for their accurate segregation during cell division. In somatic cells dividing by mitosis, newly replicated sister chromatids are connected by cohesin proteins. When the sister chromatids become attached to microtubules emanating from opposite poles of the spindle, cohesins resist the pulling forces and the ensuing tension stabilizes the chromatid–microtubule attachments. In this way, each pair of sister chromatids achieves a stable bipolar attachment to the spindle. Consequently, when cohesion is destroyed at the onset of anaphase, sister chromatids are pulled to opposite poles and each new cell receives a full complement of maternal and paternal chromosomes.In the germline, meiosis employs two successive rounds of nuclear division to produce gametes containing half the number of chromosomes as the original precursor cell. During the first division, sister chromatids remain connected while the paternal and maternal homologs are segregated (one homolog comprises a pair of sister chromatids). Homolog segregation during meiosis is governed by the same mechanical principles as sister segregation during mitosis and, as such, homologs must be connected. These connections are called chiasmata, and they are established via a process called homologous recombination, a DNA repair process that involves interaction between a broken chromosome and a homologous template chromosome.To ensure that each pair of homologs is connected by at least one chiasma, homologous recombination during meiosis is regulated at several levels. A key aspect of this regulation is the choice of recombination template. The sister chromatid is the preferred template for recombinational repair in cells dividing by mitosis. However, during meiosis this bias must be overcome so that homologs recombine and become connected by chiasmata. How template choice is regulated remains unclear, but studies of meiotic recombination in budding yeast have suggested a number of possible mechanisms.In most organisms, meiosis produces haploid gametes from diploid precursor cells Interactions between maternal and paternal homologs are the central theme of meiosis. While cohesin maintains the connections between newly replicated sister chromatids Meiotic cells induce recombination by forming numerous programmed DNA double-strand breaks (DSBs). In most organisms the homology-dependent DNA-pairing aspect of recombination then mediates the two-by-two association of the parental homologs that culminates in the intimate synapsis of the homolog pairs along their entire lengths . SubsequMeiotic recombination occurs during a protracted G2 period that follows DNA replication  gel electrophoresis of JM intermediates Mutations in a number of genes diminish meiotic inter-homolog bias in budding yeast. Most, though not all, of these genes appear to be broadly conserved suggesting that the basic mechanisms underlying inter-homolog bias are also conserved. These genes fall into two distinct functional categories:Components of a phosphokinase signal transduction pathway that responds to meiotic DSBs and modulates recombination and progression through meiotic prophase Factors involved directly in DNA strand exchange including Rad51, Dmc1, and several associated factors dmc1 mutants arrest in meiotic prophase with unrepaired DSBs, but additional mutation of Hop1, Red1, or Mek1 alleviates this arrest. In these cases DSBs are repaired, but repair occurs primarily via inter-sister recombination Several studies have demonstrated genetic interactions between mutations in these two categories of genes. For example, If meiotic DSB repair were allowed to proceed unchecked, the expectation is that most DSBs will be rapidly and unproductively repaired using the sister template. This expectation is borne out by analysis of mutations in the Hop1–Red1–Mek1 pathway dmc1 mutants (described above) has led to the idea that a barrier to inter-sister recombination is established during meiosis, essentially forcing the use of homolog rather than sister templates The behavior of PLoS Biology, Goldfarb and Lichten provide direct evidence that the sister template is used efficiently for meiotic DSB repair when allelic homolog templates are absent. They also infer that inter-sister repair may occur much more frequently than previously estimated from analyses of JM intermediates. This conclusion echoes previous inferences that the sister template is frequently engaged during meiotic recombination Numerous studies make it clear, however, that the sister template is available (or becomes available) for recombination during meiosis. In this issue of mek1 mutants. Under this model, the block to inter-sister repair observed in dmc1 mutants and in haploid cells (described above) is proposed to reflect a general block to recombination caused by pathological pan-nuclear hyperactivation of Mek1.Thus, in wild-type cells, any barrier to inter-sister recombination appears to be, at most, transient. Counter to the idea of a barrier, Goldfarb and Lichten suggest a “kinetic impediment” model in which Mek1 promotes inter-homolog bias by specifically slowing down the normally faster rate of inter-sister recombination, such that the rates of inter-sister and inter-homolog recombination are now effectively equalized. This idea is consonant with the established observation that inter-sister and inter-homolog JMs form with identical timing The “barrier” and “kinetic impediment” models are broadly similar in their basic premise that by negatively regulating inter-sister recombination, inter-homolog recombination is promoted as the only possible alternative. Contrasting, albeit non-exclusive, models propose that inter-homolog recombination is positively regulated. Such models do not dictate that access to the sister chromatid be blocked per se, but that inter-homolog bias is implemented by preferentially promoting inter-homolog interactions Unrepaired DSBs are fatal. Therefore, in addition to the primary goals of homolog pairing and chiasmata formation, the meiotic cell must ensure that all DSBs are efficiently repaired. The logical way to accomplish this is to use all available templates, the homologs and the sister. Goldfarb and Lichten More generally, the sister chromatid may regularly be engaged to more efficiently complete recombination To ensure that each pair of homologs becomes connected by chiasmata, meiotic recombination must be regulated at multiple levels: (i) DSB formation, to ensure that recombination is initiated on all homologs; (ii) template choice, to favor inter-homolog interactions; (iii) the crossover/non-crossover outcome, to produce at least one crossover; and (iv) spatial–temporal integration with the other events of meiotic prophase, i.e., homolog pairing, synapsis and segregation. Despite stunning progress in recent years, our understanding of these regulatory processes remains vague.initial recombinational interactions must be developed.Currently, the analysis of template choice during meiotic recombination in yeast is limited by the fact that only relatively late-arising, metastable JMs can be monitored. The levels and ratios of these JMs do not necessarily provide an accurate readout of the initial template choice made during the critical period when homologs are being paired. Moreover, several lines of evidence indicate that a single DSB end can engage different templates, perhaps multiple times, before forming a stable JM or recombinant product, e.g., Understanding the regulation of template choice in organisms other than yeast remains a major challenge. Cytological approaches that allow the visualization of inter-homolog and inter-sister crossovers


Rikkunshito is a popular Japanese traditional medicine that is prescribed in Japan to treat various gastrointestinal tract disorders. In a double-blind controlled study, rikkunshito significantly ameliorated dysmotility-like dyspepsia and brought about a generalized improvement in upper gastric symptoms such as nausea and anorexia when compared with a control group. Several studies in rats have shown enhanced gastric emptying and a protective effect on gastric mucosa injury with rikkunshito administration. In addition, rikkunshito in combination with an anti-emetic drug is effective against anorexia and vomiting that occur as adverse reactions to chemotherapy in patients with advanced breast cancer. However, the mechanism by which rikkunshito alleviates gastrointestinal disorders induced by anticancer agents remains unclear. It has recently been shown that rikkunshito ameliorates cisplatin-induced anorexia by mediating an increase in the circulating ghrelin concentration. Moreover, Fujitsuka et al. found that decreased contractions of the antrum and duodenum in rats treated with a selective serotonin reuptake inhibitor were reversed by rikkunshito via enhancement of the circulating ghrelin concentration. These findings show that rikkunshito may be useful for treatment of anorexia and may provide a new strategy for improvement of upper gastrointestinal dysfunction. Atractylodis Lanceae Rhizoma, Ginseng Radix, Pinelliae Tuber, Hoelen, Zizyphi Fructus, Aurantii Nobilis Pericarpium, Glycyrrhizae Radix and Zingiberis Rhizoma. It has recently been shown that oral administration of rikkunshito stimulates secretion of the orexigenic peptide, ghrelin, from the stomach -GHRP-6, with rikkunshito abolished this effect. These findings suggest that the mechanism of improvement of anorexia by rikkunshito may involve ghrelin receptor activation via stimulation of ghrelin secretion from the stomach into the plasma. Heptamethoxyflavone, an active ingredient flavonoid in rikkunshito, has been shown to have a pivotal effect on stimulation of ghrelin secretion. In addition, P388-bearing mice showed a tendency for improved survival with rikkunshito treatment, and survival was further improved by treatment with cisplatin in combination with rikkunshito, although the difference was not significant [Cisplatin has been shown to cause a significant decrease in plasma ghrelin and food intake in rodents , and intFujitsuka et al. showed that oral administration of rikkunshito restores disturbed motor activity in the gastrointestinal tract and improves anorexia in rats administered SSRIs . IntrapeCisplatin and SSRIs are widely used in clinical practice. 5-HT is a key factor in adverse reactions to these drugs, since both stimulate production of excess 5-HT and suppress 5-HT metabolism in vivo. Thus, it is of interest that involvement of the 5-HT2 receptor in appetite control has recently been shown ; specifiMore recent report demonstrated that administration of rikkunshito improve anorexia of aging via inhibiting a reduced hypothalamic ghrelin receptor reactivity . The datIn summary, administration of rikkunshito stimulates secretion of ghrelin from stomach via peripheral 5-HT2B and central 5-HT2C receptor antagonism as seen in


It is worthwhile to concentrate on the molecular implications underlying the loss of these cellular components which posed to be significant and crucial in the successful propagation of the apoptotic signals. Microarray and real-time PCR analyses demonstrated several ion and water channel genes are regulated upon the onset of lactacystin -mediated apoptosis. A time course study revealed that gene expressions of water and ion channels are being modulated just prior to apoptosis, some of which are aquaporin 4 and 9, potassium channels and chloride channels. In this review, we shall looked into the molecular protein machineries involved in the execution of AVD in the central nervous system (CNS), and focus on the significance of movements of each cellular component in affecting PCD commitment, thus provide some pharmacological advantages in the global apoptotic cell death.Programmed cell death (PCD), is a highly regulated and sophisticated cellular mechanism that commits cell to isolated death fate. PCD has been implicated in the pathogenesis of numerous neurodegenerative disorders. Countless molecular events underlie this phenomenon, with each playing a crucial role in death commitment. A precedent event, apoptotic volume decrease (AVD), is ubiquitously observed in various forms of PCD induced by different cellular insults. Under physiological conditions, cells when subjected to osmotic fluctuations will undergo regulatory volume increase/decrease (RVI/RVD) to achieve homeostatic balance with neurons in the brain being additionally protected by the blood-brain-barrier. However, during AVD following apoptotic trigger, cell undergoes anistonic shrinkage that involves the loss of water and ions, particularly monovalent ions e.g. K The mechanism of cell death is classified into necrosis and apoptosis. Necrosis is commonly caused due to injury or cell damage or exposure to toxic substances. Cells undergoing necrosis swell, the cellular contents leak out causing damage to the surrounding cells and resulting in inflammation. In contrast cells undergoing apoptosis shrink, the DNA gets broken down, mitochondria releases cytochrome C, cells exhibit “eat me” signals such as phosphatidylserine and they break up into small membrane wrapped fragments . This orHowever an imbalance of this orderly process can often result in various disease states. In fact, apoptosis is a common mode of cell death frequently observed in various neurodegenerative diseases. Increased apoptosis within the central nervous system is associated with neurodegenerative diseases such as Alzheimer’s disease (AD) and Parkinson’s disease (PD) in which a massive progressive loss of different populations of neurons is observed , whereasvia three known modes. It can be triggered via an intrinsic or extrinsic stimulus or via reactive oxidative species (ROS). Intrinsic stimulus includes proteins such as Bcl-2, Bax and cytochrome C. This usually results in apoptosis via the mitochondrial pathway whereas extrinsic stimuli are molecules such as the Fas Ligand (FasL) and Tumor Necrosis Factor (TNF) which bind to their respective receptors and initiate apoptosis via the death receptor pathway. ROS such as hydroxyl radicals or superoxide can cause direct damage to the lipid membranes and nucleic acids. Triggers of PCD are usually the absence of survival factors or the presence of negative signals. PCD occurs Though occurrence of PCD could be ascertained together by various apoptotic characteristics, cell shrinkage as a pre-cedent event of apoptosis is a single exclusive feature consistently observed in all physiologic apoptotic models. In the recent years, a new term ‘apoptotic volume decrease’ (AVD) has been coined to denote the loss of cellular volume in the absence of osmotic imbalance in PCD resulting in visible morphological cell shrinkage observation. Cell shrinkage is a ubiquitous and fundamental feature of programmed cell death that is independent of the apoptotic insults and conserved among species. Various drug-induced apoptotic models to mimic the pathological conditions of neurodegenerative disorders such as AD and PD ubiquitously demonstrated the occurrence of cell shrinkage. In a healthy brain, the presence of glial cells and blood-brain-barrier allows buffering of any osmotic fluctuations in the brain during normal physiological neuronal activity. +, Na+, Cl–, H+ and HCO3-, transport of osmolytes amino acids, methylamines, polyols, sugars and urea and also changes in gene expression.  f f+ efflupoptosis . Neverthpoptosis . An incrpoptosis . Up-regupoptosis . In a Papoptosis .+ concentrations in staurosporine- , ceramide- and serum deprivation-mediated apoptosis through suppression of enhanced K+ current in cultured mouse cortical neurons enhance the importance of K+ efflux in apoptotic activation  de de78] denditions , 58, 118nditions , 99, 139nditions . Microar3). mClic demonstrated a significant mRNA downregulation in staurosporine-mediated apoptosis in cultured cortical neurons. On the contrary, an insignificant mClic gene upregulation was observed in lactacystin- and colchicine-mediated neuronal apoptosis. Upregulation of mClic mRNA expression is further substantiated by the microarray analysis demonstrating temporal increase of mClic1 over a 48 h time course in lactacystin-mediated neuronal death (Table 1).Real-time PCR data showed a variable gene expression of mClic, a member of the CLIC intracellular chloride channel which showed localization to the cytoplasmic and inner mitochondrial membrane on different models of neuronal death (Fig. However, recently surfacing evidence denies CLC channels to be the volume-sensitive ORCC . CLC-3 k- channel blockers against apoptosis are inconsistent and controversial in numerous studies. Study conducted by [- channel blocker was more effective than that of K+ and Na+ channel blockers against staurosporine-induced apoptosis, and this inhibition of apoptosis followed a dose-dependent manner, suggesting that Cl- rather than K+ or Na+ plays a pivotal role in apoptosis. Recent findings conducted by [- channels could prevent the occurrence of AVD and downstream apoptotic events in ischemia-reperfusion-induced apoptosis in cardiomyocytes and brain neurons. In contrast, data from [- channel blockers were able to inhibit AVD in cortical neurons treated with staurosporine, however typical apoptotic morphological changes such as chromatin condensation and DNA fragmentation were still exhibited resulting in programmed cell death. Only application of K+ channel blockers, TEA and clofilium was able to successfully suppress AVD occurrence together with other features of apoptosis [- channels are mainly involved in cell volume regulation in the event of apoptosis by triggering cell shrinkage, in contrast to the K+ channels which plays a decisive role in central apoptotic events such as caspase activation and DNA fragmentation. Moreover, the cell shrinkage is not a crucial step in the activation of the apoptotic cascade such that apoptosis can still proceed in the absence of cell shrinkage. Effectiveness of Clucted by  using pructed by  supporteata from  proved opoptosis . This im+ ion is given very little attention by researchers in understanding it functional role in apoptosis since it is present intracellularly at low concentrations. However, consistently observed in various apoptotic stimuli-induced cell death models, an early transient augmentation of intracellular Na+ associated with plasma membrane depolarization preceded cell shrinkage and other features of apoptosis [+ during apoptosis was due to a shrinkage-activated sodium conductance in accordance to the RVI response [+ is short-lived and subsequently followed by a drop in both intracellular K+ and Na+ levels. The initial rise in intracellular Na+ is suggested to be contributed by two major sources: the inhibition of the Na+/K+-ATPase and the opening of the plasma membrane Na+ channels. Neuronal apoptosis has been shown to result from voltage-gated Na+ channel activation leading to Na+ entry into the cells [+ channel opener, due to the constitutive activation of voltage-gated Na+ channels leading to a prolonged state of depolarization [+ channel inhibiting free radical scavenging drug, was able to attenuate apoptosis by inhibiting rise in intracellular Na+ and reactive oxygen species production [+ channel blocker, tetradotoxin, was able to block hypoxia-induced neuronal apoptosis [Napoptosis , 61, 114response . This elhe cells . This isrization . Co-treaoduction . On the poptosis . Apoptosis is the predominant form of cell death pathway that most cells adopted in numerous neurodegenerative disorders. Apoptosis is a highly sophisticated and elaborate mode of cell death, which requires precise and coordinated cooperative among different cellular signaling pathways, to ensure the continuation of the transmission of death signal. It is an elegant and isolated mechanism of induction of cell death without causing massive killing of neighboring cells. Apoptosis is a highly regulated process with numerous feedback loops be it positive or negative at various stages, ensuring that alternative signaling cascades are present to compensate for the deficiency or inefficiency of any one signaling pathways. As such, numerous classical morphological features of cells undergoing apoptosis, e.g. cell shrinkage, membrane blebbing, chromatin condensation and DNA fragmentation, though are frequently observed, their occurrences are not universal in all apoptotic cells. Though it has been shown that apoptosis can still occur in cells in the absence of AVD, the contribution of the underlying movements of ions and water during AVD to ensure the smooth proceeding of apoptosis cannot be denied. In this review article, we have looked into the rationale and importance of the individual ion and water movements in the event of AVD, an event precedent to apoptosis, particularly with respect to cells of CNS. Although controversy exists with respect to the mitochondrial AQPs expression and their involvement in apoptosis, the importance of plasma membrane AQPs in apoptosis causes no doubt. The need for functional AQPs during AVD and for the eventual progress of apoptosis is clearly evident. By understanding the significance of the ion and water fluxes during pathological states, in correlation to that during physiological state, presents an opportunity for identification of therapeutic targets directed at the initial stage of apoptosis, i.e. AVD to allow manipulation of apoptosis or apoptosis-related diseases, thus allowing a more effective inhibition of neuronal death.


Hypertrophic cardiomyopathy (HCM) is characterized by gross cardiac and myocyte hypertrophy, myocyte disarray, and interstitial fibrosis. This condition is relatively common, with a prevalence of about 1:500 in the general population. Most patients with HCM are either asymptomatic or have only minimal symptoms. In general, HCM is a relatively benign disease with an annual mortality rate of slightly less than 1% in unselected HCM populations . HoweverThe high arrhythmic propensity in HCM is contributed by a combination of many primary substrate abnormalities like hypertrophy, myocardial fibre disarray, interstitial fibrosis etc, and possible secondary triggers like ischemia, physical exercise, and excessive sympathetic stimulation.  Marked heterogeneity in substrate and triggers would substantially reduce the predictive accuracy of any risk stratification model as known to occur in patients with many other cardiac arrhythmic conditions. To compound this, heterogeneity in the studied population, low event rate, effect of drug therapy etc would also limit the application of any predictive model.However, a few revelations in the last decade have contributed to the better understanding of the fatal arrhythmia risk. Nonsustained ventricular tachycardia (NSVT) detected on Holter monitoring in younger (less than 30 years) patients have been identified to be associated with higher mortality rate . Unlike Is there any role for 12-lead electrocardiogram to predict arrhythmia risk in them? In a cohort of patients with HCM selected because of their high risk for SCD, none of the studied electrocardiographic features  did not predict subsequent appropriate implantable defibrillator intervention for ventricular tachyarrhythmias and was not useful in risk stratification for SCD . SimilarAnother possible predictor could be exercise-induced arrhythmias. Though occurrence of ventricular arrhythmias during exercise is rare in HCM, its presence has long been identified to be associated with an increased SCD risk. In a recent study, the presence exercise-induced nonsustained VT or ventricular fibrillation was associated with a 3.73-fold increase in the risk of SCD or hemodynamically compromising sustained VT during follow-up [Of late, a few studies with cardiac magnetic resonance (CMR) imaging in HCM also have shown some promising insights into risk stratification -12. DelaIs there a role of genotype in deciding risk of SCD in these patients? Since the discovery of first causal gene and mutation for HCM in 1990, more than a dozen sarcomeric genes have been implicated in this disease . Of thesIn conclusion, despite being half a century down the initial detailed clinical description of the entity, the predictors of fatal arrhythmias in this disease still remain largely elusive. However, in future, more advanced research into the causal genes and genotype-phenotype correlation, and larger natural history studies may likely enable us to predict the arrhythmia risk in HCM in a better way.


Vulpes lagopus) rabies virus variant in most of southern Ontario, Canada, with ancillary benefits of elimination extending into Quebec and the northeastern US. Progress continues with ORV toward preventing the spread and working toward elimination of a unique variant of gray fox (Urocyon cinereoargenteus) rabies in west central Texas. Elimination of rabies in coyotes (Canis latrans) through ORV contributed to the US being declared free of canine rabies in 2007. Raccoon (Procyon lotor) rabies control continues to present the greatest challenges among meso-carnivore rabies reservoirs, yet to date intervention has prevented this variant from gaining a broad geographic foothold beyond ORV zones designed to prevent its spread from the eastern US. Progress continues toward the development and testing of new bait-vaccine combinations that increase the chance for improved delivery and performance in the diverse meso-carnivore rabies reservoir complex in the US.Steps to facilitate inter-jurisdictional collaboration nationally and continentally have been critical for implementing and conducting coordinated wildlife rabies management programs that rely heavily on oral rabies vaccination (ORV). Formation of a national rabies management team has been pivotal for coordinated ORV programs in the United States of America. The signing of the North American Rabies Management Plan extended a collaborative framework for coordination of surveillance, control, and research in border areas among Canada, Mexico, and the US. Advances in enhanced surveillance have facilitated sampling of greater scope and intensity near ORV zones for improved rabies management decision-making in real time. The value of enhanced surveillance as a complement to public health surveillance was best illustrated in Ohio during 2007, where 19 rabies cases were detected that were critical for the formulation of focused contingency actions for controlling rabies in this strategically key area. Diverse complexities and challenges are commonplace when applying ORV to control rabies in wild meso-carnivores. Nevertheless, intervention has resulted in notable successes, including the elimination of an arctic fox ( Canis familiaris) populations, where parenteral vaccination is impractical. The integration of ORV into conventional prevention and control strategies signifies a paradigm shift toward the achievement of effective rabies control in terrestrial wild Carnivora and feral dog reservoirs. Inherent to interventions targeting specific reservoirs are the benefits associated with reduced risk of human and animal exposure to rabies. Moreover, success in geographically confining or reducing terrestrial rabies virus diversity should facilitate an enhanced focus on the public and animal health dilemma associated with rabies in bats in the US and elsewhere in North America.Oral rabies vaccination (ORV) represents a socially acceptable methodology that may be applied on a broad geographic scale to manage the disease in specific terrestrial wildlife reservoirs, as well as in free-ranging or feral dog (Vulpes vulpes) in several European countries V. lagopus) variant of rabies virus, once widespread in red foxes throughout the southern part of the province Mephitis mephitis) C. latrans) in south Texas To date, ORV has been successfully applied to eliminate rabies among red foxes (Procyon lotor) rabies from expanding its geographic foothold beyond the eastern US and Canada, and as a component of potential strategies for raccoon rabies elimination Urocyon cinereoargenteus) rabies in west Texas, and preventing the reemergence of canine rabies from Mexico Currently, ORV use in the US remains focused on preventing raccoon  in October 2008 by representatives from Canada, Mexico, the US, and the Navajo Nation extends collaboration across national boundaries and multiple disciplines in four focus areas: communications, surveillance, control, and research. A fundamental tenet of the NARMP is that rabies prevention and control programs can be enhanced through an international collaborative framework. The formalization of this plan has spawned several proposed collaborative initiatives for 2009. Examples include a comparison of ORV performance between New Brunswick, Canada, and Maine, US, and closer coordination between border states and provinces involved in raccoon rabies control. Other examples include replication of the first ORV campaigns targeting dogs in Mexico, improved enhanced rabies surveillance along the Mexico-US border, and captive studies with the GnRH immunocontraceptive GonaCon in Mexico Rabies surveillance is traditionally based on human or domestic animal exposure events brought to the attention of public health officials Enhanced rabies surveillance to support ORV in the US focuses on the following types of samples: strange acting (extremely aggressive or docile) animals where no human or domestic animal exposure has been reported, road kills, animals found dead in addition to road kills, animals with injuries or lesions indicative of highly aggressive behavior, and euthanized animals from focal trapping at sites where rabid animals were recently confirmed. Not uncommonly, raccoons captured by nuisance wildlife control operators are included as enhanced rabies samples The enhanced surveillance zone for raccoons typically extends from the areas where raccoon rabies is enzootic, through the ORV zone, to approximately 80 km into areas suspected to be free of raccoon rabies . The spaSubstantial increases in suspect rabid animal samples under the enhanced rabies surveillance protocol often create an undue burden on state laboratories for timely diagnosis. Application of a direct rapid immunohistochemistry test (dRIT) developed at the Centers for Disease Control and Prevention (CDC) in the US The value of enhancing rabies surveillance beyond levels typically conducted for the protection of public health was illustrated in northeast Ohio during 2007. This area is characterized by heavy commercial business and residential developments typical of suburbia (Cleveland metropolitan area). Since 2004, when rabies was first detected in this area , surveillance has been enhanced resulting in WS submitting 5,554 animals (through 2008) for rabies testing. In each year except 2007, rabies positive cases were detected through the public health surveillance system as well. From 2004–2006, there had been declining rabies cases, suggesting a trend toward rabies-free status in this contingency action area. In 2007, all rabies positive cases in this area were detected as a function of enhanced surveillance . This waRaccoon rabies was first described in Florida in the 1940s During the 1990s, coordinated operational raccoon rabies control programs in the US expanded from small scale projects in five states  to include portions of 16 states by 2005 Obviously, lower indices to population immunity connote a greater risk of ORV zone compromise and a reduced ability to sustain effective programs, highlighting the need for improved or new baits and vaccines and strategy refinements. Frequent spillover of raccoon rabies virus variant into the striped skunk 2), intensive enhanced surveillance, and TVR. In 2008, 4,196 raccoons were parenterally vaccinated in treatment cells  rabies virus to skunks near Flagstaff, Arizona, are other major examples Notable examples of this dilemma have been documented in Alaska and Canada, where arctic fox rabies virus variant has spilled over into other animals, such as dogs and red foxes 2) recombinant rabies vaccine may hold promise based upon preliminary results obtained in captivity upon a variety of species In Ontario, field trials were conducted recently with a human adenovirus recombinant vaccine (ONRAB) developed with the intent that its strategic use will facilitate the elimination of the fox rabies foci maintained by skunks in southern Ontario Tetracycline has had a long history as a biomarker in wildlife management, including extensive use in evaluating bait uptake in ORV campaigns In 2001, apparent spillover of big brown bat rabies virus variant resulted in 19 documented cases in skunks near Flagstaff, indicative of transmission among skunks 2 during fiscal year 2008. Consequently, accelerated emphasis toward the development and production of low cost, highly effective baits and oral vaccines that can clear the regulatory process for broad scale use is a critical need. Concomitantly, improvements to the currently licensed bait-vaccine appear warranted if effectiveness can be enhanced while cost/units are conserved. In turn, costs may be conserved by refining baiting strategies based on validated models that point to areas where reduced baiting density can be applied without sacrificing effectiveness The cost effectiveness of the shifting paradigm toward controlling rabies in meso-carnivore reservoirs should and will receive continual scrutiny. To date, studies of the benefits and costs have concluded that under specific assumptions and scenarios, ORV is a viable rabies management alternative Concerns linger among some wildlife managers over potentially negative consequences from rabies control to species of conservation concern Establishment of a rabies management team composed of a coalition of diverse expertise from the public health-agriculture-wildlife management interface has been critical to facilitate coordination among rabies control programs targeting meso-carnivores in the US. The NARMP has established a continental framework that extends collaboration and coordination, capacity for rabies communications, surveillance, control, and research among Canada, Mexico, and the US. Enhanced surveillance as a complement to public health surveillance has improved decision-making capability regarding allocation of rabies control resources, including contingency actions to address emergencies, as illustrated in Ohio. Raccoon rabies has not spread appreciably since ORV intervention has expanded in the eastern US, yet rabies virus neutralizing antibody levels in raccoon populations as an index to immune buffers in existing ORV zones point to the need for improved or new baits, oral vaccines, and strategy refinements. Achieving advances that lead to improved field performance should allow for a more aggressive movement of ORV zones into raccoon enzootic areas. Measureable successes beyond containment would be expected to enhance program sustainability toward the goal of broader scale elimination of raccoon rabies and ultimately other meso-carnivore rabies virus variants. Economic analyses will remain integral to ORV planning and as a means to characterize successes in costs and benefits. Conservation concerns related to control programs cannot be ignored and require additional study to better understand the role of rabies and other diseases on the population dynamics of meso-carnivores, such as the raccoon.ORV represents a socially acceptable methodology that has helped eliminate canine rabies from the US, and restricted the distribution of raccoon, arctic fox, and gray fox variants of rabies in North America.An international rabies management team composed of experts from the public health-agriculture-wildlife management interface has been vital to the establishment of viable rabies control programs in North America.ORV in the US remains focused on the raccoon variant of rabies, while work continues to contain and eliminate the gray fox rabies variant in west Texas, and prevent canine rabies from re-emerging into the US from Mexico.Advances in enhanced rabies surveillance that relies largely on a direct rapid immunohistochemistry test have led to improved real-time management decisions for meso-carnivore rabies reservoir species in the US.ORV-related dog research in the southwestern US has potentially broad application in developing countries, where most of the 55,000 human rabies cases per year occur as the result of dog bites.Lembo T, Niezgoda M, Velasco-Villa A, Cleaveland S, Ernest E, et al. (2006) Evaluation of a direct, rapid immuno-histochemical test for rabies diagnosis. Emerg Infect Dis 12: 310-313.MacInnes CD, Smith SM, Tinline RR, Ayers NR, Bachmann P, et al. (2001) Elimination of rabies from red foxes in eastern Ontario. J Wildlife Dis 37: 119-132.Rupprecht CE, Hanlon CA, Blanton J, Manangan J, Morrill P, et al. (2005) Oral vaccination of dogs with recombinant rabies virus vaccines. Virus Res 111: 101-105.Sidwa TJ, Wilson PJ, Moore GM, Oertli EO, Hicks BN, et al. (2005) Evaluation of oral rabies vaccination programs for control of rabies epizootics in coyotes and gray foxes. J Am Vet Med Assoc 227: 785-792.Velasco-Villa A, Reeder SA, Orciari LA, Yager PA, Franka R, et al. (2008) Enzootic rabies elimination from dogs and reemergence in wild terrestrial carnivores, United States. Emerg Infect Dis 14: 1849-1854.


Current standard adjuvant therapies for early breast cancer include tamoxifen and chemotherapy, depending on the disease prognosis and menopausal status. Luteinizing hormone-releasing hormone (LHRH) analogues offer a different approach to the management of early breast cancer in pre- and perimenopausal women. The most widely studied LHRH analogue is goserelin. It acts on the hypothalamic-pituitary axis to suppress ovarian function, decreasing luteinizing hormone and oestradiol levels to post-menopausal values. Pooled data from 228 premenopausal and perimenopausal patients with advanced breast cancer enrolled in 29 studies worldwide demonstrated an objective response rate for goserelin, 3.6 mg, of 36.4%, with a median duration of response of 44 weeks. These results fall well within the ranges of reported response rates for ovarian ablation and for tamoxifen in similar patient populations. By virtue of its mode of action, goserelin does not stimulate the ovaries and is unlikely to have detrimental effects on the endometrium. In addition, given that goserelin has no oestrogen agonist-like effects, unlike tamoxifen, there is no potential for tumour stimulation in those patients becoming resistant to treatment. Goserelin is generally well tolerated, and the main side-effects are related to ovarian suppression, which is potentially reversible. Preliminary results in premenopausal women with early breast cancer indicate that endocrine treatment with goserelin plus tamoxifen may be as effective as standard combination chemotherapy (cyclophosphamide-methotrexate-5-fluorouracil), but has significantly less acute toxicity. A number of large, randomized trials are now in progress to assess the potential role of goserelin as adjuvant therapy for early breast cancer.


A software workflow to translate known differentially methylated regions into clinical biomarkers http://methmarker.mpi-inf.mpg.de/ software to help design robust and cost-efficient DNA methylation assays for six widely used methods. Furthermore, MethMarker implements a bioinformatic workflow for transforming disease-specific differentially methylated genomic regions into robust clinical biomarkers.DNA methylation is a key mechanism of epigenetic regulation that is frequently altered in diseases such as cancer. To confirm the biological or clinical relevance of such changes, gene-specific DNA methylation changes need to be validated in multiple samples. We have developed the MethMarker  Aberrant DNA methylation is a common event in many cancers ,2. FunctFor many years, research in cancer epigenetics has focused on the use of CpG island hypermethylation events of certain genes as cancer biomarkers, with the aim of improving cancer treatment through more accurate diagnosis, prognosis and therapy selection ,6. EarlyIn spite of significant investment in genome-wide screening and subsequent validation studies, few DNA methylation biomarkers have been confirmed by clinical trials. This bottleneck in the process of translating basic research findings into the clinic is partially due to a discontinuity of methods between the discovery phase and the validation phase. The methods used most commonly in the discovery phase  are too time-consuming and expensive to be used in the clinical setting. Hence, candidate biomarkers have to be adapted to high sample-throughput methods such as MethyLight , bisulfiO6-methylguanine DNA methyltransferase (MGMT) gene [Based on our experience from a pilot study on the MT) gene , we haveMT) gene ). The siApart from two key experimental analyses - the generation of high-resolution DNA methylation data in step one and assay validation in step six - this workflow is essentially bioinformatic in nature. We developed the MethMarker software as a user-friendly implementation of the bioinformatic steps, including automatic assay design for six widely used experimental methods  and computational biomarker optimization. MethMarker integrates well with existing bioinformatic tools for analyzing DNA methylation (reviewed in ): epigenMGMT gene promoter, highlighting important decisions, necessary validation experiments and potential stumbling blocks. The raw data for this case study are taken from a recent experimental study [To illustrate the biomarker development workflow outlined in Figure al study  and are MGMT gene encodes a DNA repair protein, which removes alkyl groups from the O6-position of guanine, therefore protecting the DNA from accumulating excessive damage [MGMT promoter is a frequent event in various cancers , that it is associated with decreased survival if the cancer is untreated , and that it renders tumors susceptible to alkylating drugs such as temozolomide . However, until recently no assay for measuring MGMT promoter methylation had been available that was robust enough for routine clinical use and fully compatible with DNA extracted from formalin-fixed, paraffin-embedded samples [The e damage . It has e damage  and refe samples .MGMT gene is an excellent target region for demonstrating the systematic development of a DNA methylation biomarker, such that the resulting assay is accurate, robust and cost-efficient enough for clinical use. To start with, we obtain the genomic DNA sequence of the MGMT promoter region from the UCSC Genome Browser [MGMT promoter methylation, as well as three normal brain samples for use as healthy tissue controls. Next, bisulfite-specific PCR primers are designed For these reasons, the promoter of the  Browser . We also Express ), and cl Express . The seq Express , resultiMGMT promoter on human chromosome 10, visualizes the position of the first exon and aligns the DNA methylation profiles of all 25 training samples  comprises tumor samples only, presumably those that are susceptible for chemotherapy using alkylating drugs such as temozolomide [Next, the genome sequence of the target region, the corresponding primer sequences and the BiQ-Analyzer processed DNA methylation profiles are imported into MethMarker. The software tool automatically identifies the correct location of the zolomide .MGMT promoter hypermethylation are already available [Based on this classification, our goal is to find a DNA methylation assay  that provides accurate, robust and cost-efficient separation between cases and controls. First, we let MethMarker design all feasible DNA methylation assays for the target region, using COBRA, bisulfite SNuPE, bisulfite pyrosequencing, MethyLight and MeDIP-qPCR. We chose to exclude MSP because several MSP-based assays for vailable  and becavailable . Next, wet al. [et al. [From the resulting list, we select several assay combinations that appear to provide a suitable balance between accuracy, robustness and cost . For each of these assay combinations, we let MethMarker optimize logistic regression models that predict whether a sample belongs to the cases or to the controls Figure . During et al.  and Bock [et al.  for a moMGMT gene in routine clinical use: the COBRA biomarker comprising CpG sites 5/6 and 18, utilizing the Hpy99I and HpyCH4III restriction endonucleases (r = 0.985), and the bisulfite pyrosequencing biomarker comprising CpG sites 13, 18 and 20 (r = 0.990). Both biomarkers achieve 100% test set accuracy during leave-one-out cross-validation. Compared to the biomarkers that we previously established for the same dataset [MGMT promoter methylation continue using the previously published biomarkers [For each candidate biomarker, MethMarker also calculates an extensive performance evaluation summary Figure . We use  dataset , the bioomarkers  because MGMT promoter, two key steps remain: experimental assay validation and experimental biomarker validation. First, it is essential to make sure that the DNA methylation assays included in the selected biomarker work well in the lab and result in roughly the same DNA methylation measurements as predicted based on the high-resolution DNA methylation profiles. To that end, the assays are applied to DNA from the training samples, and each assay's empirical measurement value is compared with the simulated measurement value that MethMarker calculated from the high-resolution profiles. Assays showing low correlation or high deviation should be rejected from practical use as biomarkers. Second, the most important step for any new DNA methylation biomarker is to validate its sensitivity, specificity and practical utility in a large number of patients, both by retrospective studies based on archival material with known clinical history and in prospective clinical trials. While several clinical trials have already confirmed the effect of MGMT hypermethylation on chemotherapy resistance in gliomas [MGMT's predictive role in other cancers.Having completed the design, optimization and computational validation of candidate biomarker DNA methylation assay for the  gliomas  and glio gliomas ,32, the Recent advances in genome-wide DNA methylation mapping have provided researchers with rapid and cost-efficient ways to contribute to the ever-growing list of genomic regions reported as differentially methylated in specific cancers and/or patient subgroups. However, a comparable advance for the efficient conversion of DMRs into clinical biomarkers is lacking. Thus, the rate with which new DNA methylation biomarkers are tested and confirmed in clinical trials has remained disappointingly low. While it is inevitable that a large percentage of candidate biomarkers will fail in clinical trials , a more systematic approach to epigenetic biomarker development could help discard many of these unsuccessful candidates early and at low cost. Conversely, careful selection and optimization of candidate biomarkers can reduce the risk of losing effective biomarkers due to contingencies of the validation process, such as accidental selection of DNA methylation assays that measure highly noisy CpG positions in a promoter region that would otherwise provide reliable classification. The workflow described in this paper provides a starting point toward a more systematic way of transforming disease-specific DMRs into robust and cost-efficient clinical biomarkers. The MethMarker software was developed to facilitate the implementation of this workflow. To enable further refinement and adaptation to local requirements, we are happy to share MethMarker's source code with interested researchers.MGMT gene. MethMarker's user interface reflects the workflow for biomarker design, optimization and validation outlined in Figure MethMarker is implemented in Java (version 1.5 or later required). It is platform-independent and can be launched directly from within a web browser. The software comes with a case-study tutorial demonstrating the design, optimization and validation of a DNA methylation biomarker based on the As the first step, the DMR of interest is imported. MethMarker supports several sequence formats, including FASTA, GenBank and EMBL. Typical regions of interest include the promoters of tumor suppressor genes and CpG islands that exhibit cancer-specific hypermethylation. However, MethMarker imposes no restrictions on the type of region to be analyzed. MethMarker can thus be applied not only to human cancers, but more generally to epigenotyping in all kinds of organisms that exhibit CpG dinucleotide methylation.High-resolution DNA methylation profiles for a subset of cases and controls are crucial for MethMarker's optimization process, as they provide the training set on which all candidate biomarkers are optimized and computationally validated. These profiles are usually derived by clonal bisulfite sequencing  or mass On completion of data import, MethMarker displays a high-resolution DNA methylation profile of the region of interest, visualized as lollipop diagrams or as methylation propensity diagrams. Internally, MethMarker uses Needleman-Wunsch sequence alignment  in orderOptionally, MethMarker can annotate the region with transcription start site and exon positions retrieved from the UCSC Genome Browser . To thatMethMarker implements automatic assay design for six experimental methods commonly used for DNA methylation analysis: COBRA, bisulfite SNuPE, bisulfite pyrosequencing, MethyLight, MSP and MeDIP-qPCR. The first five methods utilize bisulfite treatment of genomic DNA to detect DNA methylation indirectly. However, they differ in the way they interrogate the amount of DNA methylation, leading to specific experimental constraints that limit the application of each method to assaying a subset of CpG positions. The sixth method, MeDIP-qPCR, uses an antibody-based approach to enrich for methylated genomic DNA, which leads to quite different experimental constraints . For allAll automatically designed DNA methylation assays can be visualized, revised or excluded by the user, for example, based on results of previous experiments. Furthermore, MethMarker allows users to define and incorporate custom assays, which enables the software to include experimental methods that are not directly supported.in silico), they provide a suitable indication for identifying the most predictive DNA methylation assays to be included in the optimization step.Based on the samples for which high-resolution DNA methylation profiles are available (see step 1), MethMarker scores all DNA methylation assays in terms of their correlation with the overall level of DNA methylation in each sample. The measurement values of the DNA methylation assays are calculated directly from the high-resolution DNA methylation profiles, using a set of method-specific rules. For COBRA, bisulfite SNuPE and bisulfite pyrosequencing, the measurement value is calculated simply as the average DNA methylation level of the assayed CpG site(s), based on the high-resolution DNA methylation profiles of the respective sample. For MSP, MethyLight and MeDIP-qPCR, the measurement value is calculated as the percentage of individual clones in which all participating CpG sites are simultaneously methylated. To better resemble real PCR conditions, for MSP and MethyLight a single CpG position is allowed to have an incorrect methylation value. While simulated measurements cannot replace experimental validation of the resulting DNA methylation assays  or based on the DNA methylation profiles themselves, using one of the following methods: a fixed threshold on the average DNA methylation level, hierarchical clustering, or Second, logistic regression is used to optimize the weight with which the individual measurements contribute to the overall biomarker score, accounting for the fact that different CpGs vary in their predictiveness of the overall level of DNA methylation. Internally, MethMarker uses the WEKA package  to trainThird, the predictiveness of the logistic regression models is validated by leave-one-out cross-validation - that is, the logistic regression models are repeatedly trained on all but one training samples and their prediction performance is assessed on the remaining sample. The results of the optimization step, including a cross-validation-based estimate of the prediction performance on new data, are displayed in the biomarker summary window Figure .While the results of the leave-one-out cross-validation step 4) already provide an important selection criterion for identifying the most suitable DNA methylation biomarkers, they do not account for potential errors and experimental problems that can occur during practical use. MethMarker therefore provides an additional validation step, which assesses the robustness of each candidate biomarker toward noisy data, sequencing errors and unknown single nucleotide polymorphisms. In this step, the optimal logistic regression model is re-applied to all samples for which high-resolution DNA methylation profiles are available , and the biomarker's prediction confidence for a given sample is plotted against its mean DNA methylation level, as calculated from the DNA methylation profiles. It is thus possible to visually assess how well each candidate biomarker separates between the (methylated) cases and (unmethylated) controls. Furthermore, MethMarker assesses the robustness toward erroneous data - such as sequencing errors or unknown single nucleotide polymorphisms - by randomly changing the DNA methylation measurement of a subset of CpGs. The error rate is varied over a wide range, and the impact on the prediction accuracy is visualized in the biomarker summary window . This experimental validation step is important because it corrects for any deviations from the theoretically optimal measurement conditions that underlie the computational simulation of measurement values.When the optimization and validation steps are completed and the user is satisfied with the overall performance, one or more candidate biomarkers are typically selected for further development. MethMarker provides two ways of facilitating the steps toward comprehensive clinical testing and widespread practical use. First, MethMarker can generate a comprehensive PDF report describing the key properties of a selected biomarker. This report includes the final sample classification formula as well as a summary of the accuracy and robustness assessment. Based on this file, it is straightforward to apply the biomarker assay to new data, requiring no statistical or bioinformatic tools beyond a pocket calculator. Second, a selected biomarker can be exported in a standardized data format, PMML, which is supported by several statistics packages and can be imported into diagnostics software. PMML has been developed by the Data Mining Group  to facilO6-methylguanine DNA methyltransferase; MSP: methylation-specific polymerase chain reaction; PMML: predictive model markup language; SNuPE: single-nucleotide primer extension.COBRA: combined bisulfite restriction analysis; DMR: differentially methylated region; MeDIP-qPCR: methylated DNA immunoprecipitation quantitative PCR; MGMT: The authors declare that they have no competing interests.CB initiated the project and conceptualized workflow and software. PS designed and implemented MethMarker, developed the case study tutorial, set up the website and drafted the paper. TM devised the assay design rules and contributed his experience with COBRA, bisulfite SNuPE, bisulfite Pyrosequencing, MSP, MethyLight and MeDIP-qPCR. He also provided experimental data and performed extensive beta testing. AW provided experimental data. TL contributed advice and ideas throughout the project. All authors were involved in the writing of the paper.The following additional data are available with the online version of this paper: a screenshot of MethMarker's performance ranking of DNA methylation assays and candidate biomarkers (Additional data file Screenshot of MethMarker's performance ranking of DNA methylation assays and candidate biomarkers.Click here for fileThe XML-based PMML model that MethMarker uses for exporting, importing and storing candidate biomarkers.Click here for file


Drug resistance is a major obstacle to successful chemotherapy of primary liver cancer, which is associated with high expression of the multidrug resistance (MDR) gene product P-glycoprotein (Pgp), a multidrug efflux transporter. The most effective single agents in treatment of primary liver carcinoma belong to the anthracycline family, yet several anthracyclines are known to be substrates for Pgp. In the present study, we compared four anthracyclines with respect to cell growth inhibition, intracellular accumulation and cellular efflux using the HB8065/R human hepatoma cell line which is rich in Pgp, and the Pgp-poor parental line HB8065/S. The anthracyclines were also administered in conjunction with the Pgp-modifying agents verapamil and SDZ PSC 833 to assess modulation of resistance. The HB8065/R cells were sensitive to aclarubicin (ACL) and highly resistant to epirubicin (EPI), doxorubicin (DOX) and daunorubicin (DNR). SDZ PSC 833 enhanced accumulation, decreased efflux and increased cytotoxicity of EPI, DOX and DNR in the HB8065/R cells, but none of these effects was seen with ACL. In conclusion, ACL is apparently not transported by Pgp and retains its activity in a multidrug-resistant human hepatoma cell line; such properties can be exploited for clinical purposes.


The use of intensity-modulated radiation therapy (IMRT) is rapidly advancing in the field of radiation oncology. Intensity-modulated radiation therapy allows for improved dose conformality, thereby affording the potential to decrease the spectrum of normal tissue toxicities associated with IMRT. Preliminary results with IMRT are quite promising; however, the clinical data is relatively immature and overall patient numbers remain small. High-quality IMRT requires intensive physics support and detailed knowledge of three-dimensional anatomy and patterns of tumour spread. This review focuses on basic principles, and highlights the clinical implementation of IMRT in head and neck and prostate cancer. Intensity-modulated radiation therapy (IMRT) represents a promising new advance in the field of radiation oncology. Intensity-modulated radiation therapy allows for improved ‘shaping’ of radiation dose profiles around tumour and at-risk nodal structures while sparing adjacent normal tissue structures. This capacity for improved dose distribution affords considerable opportunity to reduce the overall toxicity profile associated with radiation therapy. The use of IMRT is advancing rapidly worldwide as this technology has become commercially available. Despite considerable promise, IMRT use remains in relatively early stages, and must be delivered with strict attention to quality assurance as the clinical data and patient follow-up mature. Furthermore, IMRT is quite labour-intensive, with strong dependence on physics and quality assurance support, thus leaving open the possibility for significant heterogeneity in the precision of IMRT practice.Intensity-modulated radiation therapy refers to a specific technique of linear accelerator-based radiation therapy whereby beams are modulated in such a manner to produce highly conformal dose distributions. A primary objective of IMRT is to reduce dose to selected normal tissue structures in an effort to preserve function, while maintaining full dose delivery to tumour targets. For example, in conventional head and neck (H&N) radiotherapy, static fields are shaped by blocks and modulated by simple beam wedges or tissue compensators  and IMRT planning, the radiation oncologist designates specific targets  and avoidance structures . In 3D-CRT, simple beam arrangements are used with generous field margins to account for daily set-up variation and physical characteristics of the beam itself. The radiation dose and profile is then calculated in a process known as Intensity-modulated radiation therapy was first conceptualised in the 1960s. However, it was not until the 1980s–1990s that the computing capability required for complex inverse planning algorithms became commercially available . In 1994Clinical research efforts in IMRT have generally considered two basic paradigms. The first research strategy seeks to maintain current tumour control rates while decreasing toxicity profiles. For example, IMRT studies in H&N cancer commonly strive to maintain conventional dose to primary tumour and at-risk nodal regions, while diminishing dose to adjacent normal tissue structures such as salivary glands and spinal cord. The second strategy attempts to escalate tumour target dose while maintaining acceptable levels of toxicity. This approach has been taken in dose-escalation and hypofractionation trials for prostate cancer and more recently in lung cancer. Regardless of the primary objective, precise, reproducible accurate patient/target set-up and rigorous physics quality assurance are critical to successful IMRT delivery. In this review, we highlight the role of IMRT in two of the most common anatomic sites of current use; H&N and prostate cancer.Radiation plays a central role in the treatment of H&N cancer. New radiation delivery techniques offer powerful potential to diminish the spectrum and severity of radiation toxicities for H&N cancer patients. For many decades, conventional H&N radiation techniques have involved treatment with generous opposed lateral beams to encompass the known primary tumour and upper cervical lymphatics. This classical technique produces a relatively homogeneous dose distribution that allows excellent target dosing while minimizing hot and cold spots. However, due to the tight proximity of tumour targets and normal tissue in the H&N region, many uninvolved structures including salivary glands, spinal cord, auditory apparatus, optic apparatus, mandible, and vocal cords can unnecessarily receive high doses of radiation.In H&N cancer, one of the most common rationales for IMRT is to preserve salivary gland function and thereby diminish the severity of chronic xerostomia with associated adverse impact on taste, swallowing, dentition, speech, and overall quality of life. In addition, the capacity of IMRT to limit dose to normal tissue structures may also allow dose escalation and differential dose painting, thereby accomplishing ‘in-field tumour boosting’ . Prospective, multiinstitutional protocols that incorporate IMRT for H&N cancer patients remain in very early stages.Early reports have described clinical promise with H&N IMRT, both for tumour control and reduction of xerostomia. However, these data generally represent limited single-institution experience, often with heterogeneous cohorts of patients . It may, therefore, prove advantageous to use larger daily fractions of >2.0 Gy (hypofractionation), rather than conventional 1.8–2.0 Gy fractions commonly employed, when treating most other tumour types. Using linear quadratic formulation, one can predict an improved ratio of tumour control to normal tissue toxicity (the therapeutic ratio) for prostate cancer by using hypofractionation .As with any new technology, there is enthusiasm mixed with caution regarding the use of IMRT. Some have voiced concern regarding the embracement of IMRT as a standard approach until the completion and comparative evaluation of systematic clinical trials . NeverthThere exists a lack of global standardisation in IMRT planning and delivery. The literature describes several distinct IMRT techniques, and several aspects of the IMRT planning processes remain highly practitioner dependent. A variety of fractionation regimens and target delineation techniques are in common use. In a recent study, substantial variation in both target delineation and fractionation recommendations were observed among worldwide H&N experts when provided the identical tonsil cancer case   required for IMRT delivery and the potential for increased radiation exposure. With increased modulation of the radiation beam, more monitor units are generated to deliver the prescribed dose. Consequently, there can be increased leakage from the linear accelerator, increasing the total body exposure by 2–3 fold . In a reAs H&N IMRT steadily advances into more common use, the design of clinical trials that explore the use of chemoradiation, altered fractionation, and molecular targeted therapy becomes more complex. Indeed, the cooperative oncology groups are struggling currently with systematic methodology to credential and quality-assure the process of H&N IMRT for participating institutions. The successful accomplishment of future H&N cancer treatment trials will need to acknowledge the increasing use of IMRT despite inherent difficulties in trial design and quality assurance. This represents a significant challenge in that a broad series of promising molecular agents, that may enhance radiation response are becoming available. However, the profound variation in IMRT expertise and delivery technique across institutions currently renders this an added variable that serves to complicate the evaluation of new molecular agents with radiation.In prostate cancer, novel hypofractionated regimens have been proposed to exploit radiobiological properties with significant shortening of treatment time without loss of efficacy or increase in toxicity  that render patients most likely to truly benefit from IMRT will be valuable. The time, expense, and expertise required to realise optimised and reproducible IMRT delivery across institutions warrants acknowledgement regarding the most appropriate usage and necessary quality assurance processes to ensure patient benefit and safety. The systematic accumulation of clinical data is of great importance for advancement in this field. Ideally, this data will come in the form of controlled clinical trials that rigorously examine not just radiation physics and dosimetry, but acute and late normal tissue effects with long term clinical follow up.


Lymphoid cells stimulated by soluble tumour antigens in the MCA-induced murine fibrosarcoma system have been identified by subclass and protective capacity in adoptive syngeneic hosts. Lymph-node or spleen cells taken at weekly intervals after inoculation of syngeneic chemically induced fibrosarcomas were enriched by 3 methods in T, B, and "null" cell subclasses, and assayed for proliferative kinetics in response to soluble membrane antigens. The stimulated subpopulations were found to be heterogeneous, their composition varying with time and tumour burden. Initial proliferative responses after tumour inoculation were limited to the T-enriched subpopulation. Later during tumour growth, T, B and null cell fractions were vigorously and equally stimulated by tumour antigen. The ability of the same T, B or null-cell subpopulations to inhibit tumour growth was measured in adoptive hosts by a modified Winn assay. Only the T-cell subpopulation responding to tumour antigen in vitro effectively and consistently retarded tumour growth in vivo. In contrast to the shared specificities on syngeneic tumours identified by the proliferative assay, tumour-growth inhibition was limited to the specific tumour borne by the cell donor.