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The incidence of human fungal infections is increasing due to the expansion of the immunocompromised patient population. The continuous use of different antifungal agents has eventually resulted in the establishment of resistant fungal species. The fungal pathogens unfold multiple resistance strategies to successfully tackle the effect of different antifungal agents. For the successful colonization and establishment of infection inside the host, the pathogenic fungi switch to the process of metabolic flexibility to regulate distinct nutrient uptake systems as well as to modulate their metabolism accordingly. Glucose the most favourable carbon source helps carry out the important survival and niche colonization processes. Adopting glucose as the center, this review has been put forward to provide an outline of the important processes like growth, the progression of infection, and the metabolism regulated by glucose, affecting the pathogenicity and virulence traits in the human pathogenic fungi. This could help in the identification of better treatment options and appropriate target-oriented antifungal drugs based on the glucose-regulated pathways and processes. In the article, we have also presented a summary of the novel studies and findings pointing to glucose-based potential therapeutic avenues to be explored to tackle the problem of globally increasing multidrug-resistant human fungal infections.

Antibodies represent a well-established class of clinical diagnostics for medical applications as well as essential research and biotechnological tools. Although both polyclonal and monoclonal antibodies are indispensable reagents in basic research and diagnostics but both of them have their limitations. Hence, there is urgent need to develop strategies aimed at production of alternative scaffolds and recombinant antibodies of smaller dimensions that could be easily produced, selected and manipulated. Unlike conventional antibodies, members of Camelidae and sharks produce antibodies composed only of heavy chains with small size, high solubility, thermal stability, refolding capacity and good tissue penetration in vivo. The discovery of these naturally occurring antibodies having only heavy-chain in Camelidae family and their further development into small recombinant nanobodies represents an attractive alternative in drug delivery, diagnostics and imaging. Nanobody derivatives are soluble, stable, versatile, have unique refolding capacities, reduced aggregation tendencies and high-target binding capabilities. They can be genetically customized to target enzymes, transmembrane proteins or molecular interactions. Their ability to recognize recessed antigenic sites has been attributed to their smaller size and the ability of the extended CDR3 loop to quickly penetrate into such epitopes. With the advent of molecular engineering and phage display technology, they can be of potential use in molecular imaging, drug delivery and therapeutics for several major diseases. In this review we present the recent advances in nanobodies for modulating immune functions, for targeting cancers, viruses, toxins and microbes as well as their utility as diagnostic and biosensor tools.

Much recent attention has been given to traditional medicines and natural products with potential and promising anti-inflammatory properties. Leaf extract of Ginkgo biloba GbE-761, possesses several clinical beneficial effects and is now used in a broad spectrum of pharmacological actions. However, mechanisms associated with its anti-inflammatory effect are not very much clear. Previously we established that GbE-761 exerts a neuroprotective effect against ischemic brain injury through an anti-apoptotic mechanism. In this study we investigated the anti-inflammatory effect of GbE-761 in LPS-activated murine macrophage. Our results demonstrate that GbE-761 potently inhibited LPS-induced NO and PGE2 production and these observations were consistent with the inhibition in the protein and mRNA expression levels of inducible iNOS and COX-2 enzymes in a dose dependent manner. Not only this, GbE-761 attenuated the production of pro-inflammatory cytokines like IL-1 , IL-6 and TNFin LPS-activated murine macrophages in concentration dependent manner. In addition we reported inhibition in the protein and mRNA expression levels of IL-1 , IL-6 and TNF. Furthermore, GbE-761 inhibited the NFB activation induced by LPS. Together these results suggest that anti-inflammatory properties of GbE-761 extract might be the results of inhibition of iNOS and COX-2 by upregulating anti-oxidative enzymes and inhibition of pro-inflammatory cytokines (IL-1 , IL-6 and TNF) expression through the down-regulation of NF-kB activation in murine macrophages.

Tuberculosis (TB) also known as white plague is a major infectious disease and a global emergency. It is one of the oldest recorded human afflictions and still continues to cause widespread morbidity and mortality in children and adults worldwide, despite the extensive use of a live attenuated vaccine and several antibiotics. One third of the world’s population is already infected and about 3 million people die and 8 million people develop the active disease each year. In the last decade there is a sharp rise in the cases of TB mainly due to emergence of multi-drug resistant (MTD), extensively drug-resistant (XDR) and totally drugresistant (TDR) strains of Mycobacterium Tuberculosis, which are virtually untreatable.

This study has been undertaken to explore the therapeutic effects of deguelin and specific siRNAs in HeLa cells. The data provided clearly show the silencing of ERK 1/2 with siRNAs and inhibition of ERK1/2 with deguelin treatment in HeLa cells. Additionally, we are providing information that deguelin binds directly to anti-apoptotic Bcl-2, Bcl-xl and Mcl-1 in the hydrophobic grooves, thereby releasing BAD and BAX from dimerization with these proteins. This results in increased apoptotic activity through the intrinsic pathway involved in rupture of mitochondrial membrane and release of cytochrome C. Evidence for inhibition of ERK1/2 by deguelin and escape of BAD phosphorylation at serine 112 through ERK/RSK pathway has been further fortified by obtaining similar results by silencing ERK 1/2 each with specific siRNAs. Increase in BAD after treatment with deguelin or siRNAs has been interpreted to mean that deguelin acts through several alternative pathways and therefore can be used as effective therapeutic agent.

Background: Epilepsy is the second most common neurological condition, and antiepileptic drugs (AEDs) are used as a prophylactic measure. Any amount of poor-quality medicine is unacceptable because it increases morbidity and mortality, thus an assessment of quality of AEDs are important in developing countries like India. Phenytoin (PHT) has become a major first-line AED in the treatment of partial and secondarily generalized seizures, but like other pharmaceuticals, PHT too may develop impurities at various stages of development, transportation, and storage which make the pharmaceutical risky. Hence, the objective of the study was to test PHT concentrations in various injectable medicines available in the local markets of Kashmir Valley. Materials and Methods: Quality analysis of PHT by high-performance liquid chromatography (HPLC) is more precise and accurate than using other analytical methods such as enzyme-based assays and immunoassay as the use of HPLC technique in the analytical field helps in structure elucidation and quantitative determination of impurities. In this study, we studied the different parameters of quality of various PHT drugs available in the local markets using HPLC and compared the results obtained with that of the standard. Results : We observed that all the tested PHT sodium injections available in the local market have standard concentrations as all the samples under study showed the peak exactly where the standard PHT peak was supposed to be. Hence, our results suggest that the quality of various PHT drugs used in Kashmir Valley is satisfactory and safe. Conclusion: It is clear from our results that there is a need for a quick and effective drug quality analysis method. As shown from the experiments performed in this study, HPLC is not only a suitable but also efficient system to carry out drug quality evaluation and help curb or at least keep in check the rampant sale of substandard drugs that go unabated in developing countries like India.

The role of CD80 and CD86 costimulatory molecules is well established in the activation of T cells but not an - tigen presenting cells. Recently, many reports in literature have demonstrated categorically the influence of CD80 and CD86 in the activation of B cells and dendritic cells. Stimulation via CD80/CD86 in B cells can modulate their prolifera- tion, IgG secretion and expression of pro-apoptotic and anti-apoptotic molecules, nuclear localization of NF- B p50 subunit, phosphorylation of Rel A (p65) and I B-alpha and increased oct-2 expression. In case of dendritic cells, it has been shown that signals induced via CD80 and CD86 enhance the production of IL-6 and IFN- which in turn, up- regulates the expression of the enzyme indolamine 2, 3-dioxygenase that results in tryptophan catabolism and affects T cell proliferation. Interestingly, it has been shown that co-stimulation through CD80 can restrict the survival of lympho- mas and can also induce apoptosis in neural stem cells. Consequently, it may be concluded that CD28/CD152-CD80/86 interaction delivers a bi-directional co-stimulation, thereby not only having impact on the function of T cells, but also an- tigen presenting cells.

Alzheimer's disease (AD) and vascular dementia (VaD) are the most common cause of dementia. Cerebral ischemia is a major risk factor for development of dementia. (123)I-FP-CIT SPECT (DaTScan) is a complementary tool in the differential diagnoses of patients with incomplete or uncertain Parkinsonism. Additional application of DaTScan enables the categorization of Parkinsonian disease with dementia (PDD), and its differentiation from pure AD, and may further contribute to change the therapeutic decision. The aim of this study was to analyze the vascular contribution towards dementia and mild cognitive impairment (MCI). We evaluated the utility of DaTScan for the early diagnosis of dementia in patients with and without a clinical vascular component, and the association between neuropsychological function, vascular component and dopaminergic function on DaTScan. One-hundred and five patients with MCI or the initial phases of dementia were studied prospectively. We developed an initial assessment using neurologic examination, blood tests, cognitive function tests, structural neuroimaging and DaTScan. The vascular component was later quantified in two ways: clinically, according to the Framingham Risk Score (FRS) and by structural neuroimaging using Wahlund Scale Total Score (WSTS). Early diagnosis of dementia was associated with an abnormal DaTScan. A significant association was found between a high WSTS and an abnormal DaTScan (p < 0.01). Mixed AD was the group with the highest vascular component, followed by the VaD group, while MCI and pure AD showed similar WSTS. No significant associations were found between neuropsychological impairment and DaTScan independently of associated vascular component. DaTScan seems to be a good tool to discriminate, in a first clinical assessment, patients with MCI from those with established dementia. There was bigger general vascular affectation observable in MRI or CT in patients with abnormal dopaminergic uptake seen on DaTScan.

Cancer is a major health problem worldwide and is one of the most prominent causes of morbidity and mortality in children and adults. According to the American Cancer Society each year around 1.5 million cases of cancer are diagnosed. Presently radiation and chemotherapy are the current “gold standards” for cancer treatment, but these therapies have their own limitations and are marginally effective, toxic, and serve to diminish the quality of life for cancer patients. On the other side immunotherapy represents an attractive alternative to these traditional treatment regimens. Despite overwhelming evidence that the immune system is capable of recognizing and eliminating tumors, both spontaneously and in response to immune-based therapy, such protection is abrogated in the face of compensatory immunosuppressive events characteristic of progressive disease. Thus, a major goal of novel immune-based therapies is the coordinate silencing of regulatory circuits and amplification of protective T-cell function. A successful immune response to tumor depends on a series of specific interactions between a T cell and an antigen-presenting cell. But unfortunately even after the interaction with T cells, an effective immune response is not generated because tumor cells have developed a number of mechanisms to resist recognition and elimination by immune system. Tumors evade immune responses by several mechanisms, like downregulating the expression of costimulatory and major histocompatibility complex molecules; producing immune-suppressive substances; and not expressing tumor antigens and inducing tolerance to tumor antigens. So manipulating the immune system in order to induce clinically relevant responses against cancer is a longstanding goal. Interventions to enhance tumor-specific immunity through vaccination, sustaining effector T-cell activation, or increasing the numbers of tumor-specific T cells using ex vivo expansion, have all resulted in clinical successes. The CD28 and B7 protein families are critical regulators of immune responses and, in the past few years, several new family members have been identified. Preclinical studies exploring the role of members of the CD28 and B7 families support the targeting of these pathways for new therapeutic approaches. Indeed, recent phase I clinical studies using agonists and antagonists of the CD28/CTLA-4/B7 pathway have shown promising results in cancer therapies.

Since the original proposal of a two-signal model for T-cell activation, there has been a continuous discovery of new costimulatory molecules on antigen-presenting cells (APCs) and their receptors on T cells, but till date, CD80 and CD86 remain the best-defined costimulatory molecules. The ability of APC to deliver the costimulatory signal to T cells by CD80 and CD86 molecules is very well established. In contrast, whether the engagement of CD80 and CD86 molecules by CD28 and CTLA-4 also induces bidirectional costimulation that can affect the function of APCs was very poorly documented. However, we for the first time established the concept of reverse costimulation or bidirectional costimulation, where we have categorically demonstrated that the costimulatory signals not only influence the activation of T cells, but by reverse/bidirectional costimulation they can also affect the activity of APCs (B cells). We showed that costimulation through CD80 specifically inhibits the proliferation and antibody secretion by B cells and B-cell lymphoma by upregulating the expression of proapoptotic molecules such as caspase-3, caspase-8, Fas, FasL, Bak, and Bax, and downregulating the levels of antiapoptotic molecule Bcl-x(L). In contrast, costimulation through CD86 augmented the level of antiapoptotic molecules Bcl-w and Bcl-x(L) and decreased the levels of caspase-8. Many groups worldwide after our report have endorsed the concept of bidirectional/reverse costimulation. It has been shown that targeting death receptor-6 on B cells can enhance B-cell expansion, survival, and humoral responses. Further, cross-linking of B7-DC costimulatory molecule on dendritic cells (DCs) with the monoclonal antibody directly potentates DC function by enhancing antigen uptake, DC presentation of major histocompatibility complex–peptide complexes, promoting DC survival, and increasing secretion of interleukin-12p70. Further, this B7-DC cross-linking restores antigen uptake and augments APC function by matured DCs. To translate this field into the clinic, it is urgent to develop novel methods to target currently appreciated costimulatory pathway. Despite the complex roles and interactions within the CD28 and B7 costimulatory families, we predict that novel approaches targeting these families will yield new therapies for the treatment of inflammation, autoimmunity, transplantation, cancer, and infectious diseases.

The immune system has the remarkable ability to defend against a diversity of microbial pathogens, yet not respond to self. Activation of this immune system is dependent on both innate and adaptive responses to environmental challenges such as infection. Activation of adaptive immune responses requires signals through antigen-specific T- and B-cell receptors. However, this antigen-specific signal alone is not sufficient to drive the activation of naive T cells and requires a second antigen-independent nonspecific signal known as costimulatory signal. The primary source of this costimulatory signal is interactions between the T-cell integral membrane proteins and their corresponding ligands on the APC. Currently, the B7 family has seven members (B7-1, B7-2, ICOS-L, PD-L1, PD-L2, B7-H3, and B7-H4), and the TNFR family has six members (HVEM, BTLA, CD70, CD30, 4-1BBL is also known as CD137 is a type II glycoprotein belonging to the tumor necrosis factor superfamily (4-IBB-L), and OX40L). The best-defined costimulatory pathway, to date is the B7-1/B7-2/CD28 pathway, and this has been shown to constitute the primary and strongest costimulatory signal delivered by APCs to amplify T-cell activation. The biologic consequences of CD28 costimulation include increased cytokine gene expression, stabilization of cytokine messenger RNA, augmentation of glucose uptake and utilization, promotion of T-cell survival, and maintenance of T-cell responsiveness upon subsequent restimulation. With the rapid increase in knowledge of the function of a growing number of specific costimulatory molecules, costimulation is now starting to be recognized as the master switch for T-cell activation and modulation of T-cell function. Therefore, understanding the relationships between the costimulatory ligands and their receptors in different immune effector pathways will surely help us in improving immunomodulatory therapeutics, development of improved vaccines, and avoidance of unintended tissue injury.

The prevalence of allergy and allergic asthma is increasing worldwide. More than half of the US population has a positive skin prick test and approximately 10% are asthmatics, which is evidence of the growing prevalence of allergy in industrialized countries. The immune response to any infection or environmental antigen is a tightly regulated process, which normally results in protection and tolerance. However, in allergic disease, the activated immune response results in a chronic proinflammatory state characterized by antibody secretion (immunoglobulin E, IgE) and T-cell activation by normally well-tolerated antigens. T cells play an important role in the pathogenesis of allergic responses. T-cell receptor engagement by antigenic peptides alone is not sufficient to drive the activation of naive T cells and requires a second costimulatory signal. Costimulatory molecules are responsible for second signals that induce T-cell activation and proliferation. The best-characterized costimulatory pathways include CD80/CD86 interacting with CD28/cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4). The positive signals induced by these costimulatory molecules are counter-balanced by other members of the costimulatory family, including CTLA-4, programmed death (PD)-1, and B- and T-lymphocyte attenuator (BTLA), which dampen immune responses. Depending on the type of molecules involved, this secondary signal can promote the development of an inflammatory allergic reaction or may favor immune regulation. Several molecules of the tumor necrosis factor receptor family (OX40, CD30, 4-1BB, Fas, CD27, CD40) and B7 family (CD80, CD86, PD-1, ICOS, CTLA-4) play an important role in delivering costimulatory signals in early and late phases of allergic response. Currently, the treatments of allergic disease are focused on the suppression of key inflammatory mediators or inflammatory cell populations and include anti-histamines, anti-leukotrienes, β2-adrenergic receptor agonists, and corticosteroids. However, these approaches only provide a temporary suppression of disease symptoms. Successful long-term treatment can only be provided by allergen-specific immunotherapy, which restores normal immunity against allergens. Therefore, costimulatory molecules involved in promotion or prevention of allergic immune responses are potential targets for the development of novel therapeutic approaches. This chapter aims to recapitulate the influence of costimulatory signals on allergic responses and to summarize our current understanding of the relationship between allergic diseases.

Activation of T lymphocytes is a critical component of the immunological response to foreign protein molecules. T-cell receptor recognition of peptide-loaded major histocompatibility complex molecules provides antigen specificity and initiates the required steps for T-cell activation, although additional signals are needed for complete T-cell activation. CD28 signaling causes the initial activation of naive CD4 T cells by increasing the sensitivity of the T cell to antigen receptor engagement, and as a result proliferation is induced at otherwise submitogenic concentrations of antigen. A major consequence of CD28 signaling is protection from cell death. The two dominant pathways of apoptosis in T cells are receptor mediated (i.e., tumor necrosis factor family) or mitochondrial-associated proteins (i.e., the Bcl family), both of which are modulated via CD28 signaling. So, an understanding of costimulation and the mechanism by which it regulates immune responses provides new avenues for the development of therapeutics. As CD28 decreases the threshold required to activate naive T cells, as well as increasing the survival of activated T cells, one can envision development of drugs that either potentiate or downregulate an immune response. Further CD28 requirement of memory CD8+ T cells for expansion has been shown in multiple viral infections such as influenza type A virus, HSV, vaccinia virus, and murine gamma herpes virus and is required for rapid pathogen clearance. Costimulatory molecules have been shown to not only help in the generation and maintenance of virus-specific memory CD8+ T cells, but are also important for the reactivation of memory CD8+ T cells and secondary responses. This information has important implications in designing efficient vaccination strategies against pathogens and tumors which can down-regulate costimulatory signals. Moreover the transfusion of T cells, also called adoptive T-cell therapy, is an effective treatment for viral infections and has induced regression of cancer in early stage clinical trials. Adoptive T-cell therapy for cancer is a form of transfusion therapy consisting of the infusion of various mature T-cell subsets with the goal of eliminating a tumor and preventing its recurrence. Second-generation chimeric antigen receptors (CARs) have been generated in which intracytoplasmic domains, derived from costimulatory molecules such as CD2837, or CD134/OX40, or CD137/4-1BB, are incorporated within CARs to fully activate T cells. Second-generation CARs were constructed to provide signaling both through the CD3 zeta chain and, primarily, the CD28 costimulatory molecule by placing the signaling domains in series as a single gene multidomain product. Constructs with the CD28 signaling domain proximal and the zeta chain distal to the membrane were found to be better expressed than constructs with the opposite orientation, and were capable of mediating up to 20 times more interleukin (IL)-2 production upon stimulation with solid-phase antigen compared with first-generation CARs. Subsequently, CAR constructs with costimulatory signaling domains from CD28, inducible costimulator (ICOS), OX40 (CD134), or 4-1BB (CD137) in series with the CD3 zeta signaling region were evaluated using resting human primary T cells. It was found that second-generation CARs, when providing any of these B7 or tumor necrosis factor receptor family costimulatory signals in series with CD3 zeta, confer self-sufficient antigen-driven clonal expansion and enhanced effector function in resting human T cells. Furthermore, addition of the CD28 signaling domain to CARs has been shown to enhance CAR T-cell resistance to regulatory T cells.

Vertebrate immune system is highly evolved to combat and eliminate pathogens. However, some pathogens can successfully subvert the host immune system to establish their intracellular survival via strategies like immunosuppression, molecular mimicry, disguise or sequestration of antigens, circumvention of complements and cytokines cascade, blockade of antigen presentation, escape from apoptosis and autophagy, and modulation of costimulatory signals). T cells and antigen-presenting cells (APCs) play crucial roles in eliminating intracellular pathogens. The optimal activation of naive T cells is achieved by occupancy of T-cell receptor by the major histocompatibility complex (MHC)–peptide complex displayed on the surface of APCs, delivery of costimulatory signals, and the presence of proinflammatory cytokines. The expression of costimulatory molecules on APCs is critical in shaping the extent and nature of the immune response. Thus, an encounter of T cells with MHC–peptide can result in two discrete events: (a) T-cell proliferation and differentiation into effector cells; (b) or anergy or apoptosis. The question of which of these outcomes transpires is determined by the delivery of appropriate costimulatory signals. An array of costimulatory molecules is displayed on the surface of APCs (CD80/B7-1, CD86/B7-2, CD83, CD40, PDL-1, DC-SIGN, 4-1BBL, etc.) and T cells (CD28, CTLA-4, CD40L, PD-1, OX40, 4-1BB, etc.). The level of the expression of the costimulatory molecules may play an important role during the course of acute disease and its remission or relapse. Hence, modulation of these molecules by pathogens can help them to establish their existence in the host.

Background: One of the mechanisms by which tumors evade the immune system is by downregulating the expression of costimulatory molecules like CD80 and CD86. The role of CD80 in the activation of T cells is well established, and current studies are exploring its role in cancer. Objective: To examine the possible role of CD80 signaling in generating an effective immune response against cancer cells. Methods: Many reports have described the influence of CD80 on the growth of B cell and follicular lymphomas. Signaling through CD80 in B cell lymphomas can retard their proliferation by upregulating expression of pro-apoptotic molecules and downregulating antiapoptotic molecules and can therefore induce apoptosis. Recently, a Phase I/II study of treatment with CD80-specific antibody has shown it to be quite effective in relapsed and refractory follicular lymphoma patients. Conclusion: This study shows that anti-CD80 immunotherapy may have a potent role in treating CD80-bearing cancer cells.