Search Results for: pathway

Mambalgin 1 – Blocking ASIC channels in pain pathways

Acid-Sensing Ion Channels (ASICs) are neuronal voltage-insensitive cationic channels which are activated by extracellular protons. They belong to the ENaC/Deg superfamily of ion channels. Up to now 6 members of the ASIC family have been identified: ASIC1 – ASIC 4, and the splice variants ASIC1a, 1b and 2a, 2b. The ASIC family members are trimeric and can be made up of different combinations of subunits. All ASICs are expressed in the peripheral nervous system. ASIC1a, 2a, 2b and 4 are expressed in the central nervous system. ASICs are promising drug targets for treating a wide variety of conditions linked to both the CNS and PNS especially implied in pain pathways.

Structure of Mambalgin-1

Schwarze Mamba

Fig. 1: Dendroaspis polylepis polylepis

Mambalgin-1 (from Smartox, now available through tebu-bio) was initially isolated by Sylvie Diochot and collaborators from the venom of the black mamba (Dendroaspis polylepis polylepis, Fig. 1).

Mambalgin-1 belongs to the family of three-finger toxins (Fig. 2) and has no sequence/structural homology with either PcTx1 or APETx2. Mambalgin-1 differs from mambalgin-2 by one amino acid. Both have demonstrated a similar activity.

Mambalgin 1 – a blocker of ASIC1 channels implied in pain pathways

It has been shown that Mambalgin-1 is a potent and selective blocker of ASICs implied in pain pathways (1, 2). Mambalgin-1 rapidly and reversibly inhibits recombinant homomeric ASIC1a (IC50=55 nM) and heteromeric ASIC1a+ASIC2a (IC50=246 nM) or ASIC1a+ASIC2b channels (IC50=61 nM) but also human channels hASIC1b (IC50=192 nM) and   hASIC1a+hASIC1b (IC50=72nM).

Mambalgin-1 has no effect on ASIC2a, ASIC3, ASIC1a+ASIC3 and ASIC1b+ASIC3 channels, as well as on TRPV1, P2X2, 5-HT3A, Nav1.8, Cav3.2 and Kv1.2 channels. Thus Mambalgin-1 can be used as a selective inhibitor for the above mentioned homomeric and heteromeric ASICs.

Interested in further information? Please contact me through the form below.

If you’d like an overview of the complete venomous toxins available, take a look to our special page: Synthetic peptide toxins ideal for studying ion channels

References:

(1) S, Dichot et al., Black mamba venom peptides target acid-sensing ion channels to abolish pain, Nature. 490 (7421), pages 552-5 (2012)

(2) Wen M., et al., Site-specific fluorescence spectrum detection and characterization of hASIC1a channels upon toxin mambalgin-1 binding in live mammalian cells. Chem Commun. 51 (38), pages 8153-6 (2015)

9 pathway-specific screening assays in Immunotherapy

The immune system is a system of cells and organs whose function is to defend an organism from foreign pathogens. With the ability to mount a response against virtually any foreign material and return to a quiescent state following neutralization of the threat, this fascinating organ system displays remarkable specificity and plasticity. To achieve this, there is a multifaceted balancing act between the many activators and suppressors which maintains homeostasis of the body’s perhaps most complex organ system.

[Read more…]

Immunotherapy Screening – IDO pathway

In previous blogs, I invited you to join me in exploring the relevance of the following pathways:

PD1 - PD-L1 - PD-L2

 

Today I’d like to focus on the IDO pathway. [Read more…]

Focus on the Calcineurin-NFAT Pathway…

A few weeks ago, I received an email from one of our blog readers. Following my post about the Autophagy pathway, she asked me for the same kind of post about the Calcineurin-NFAT Pathway (Cn-NFAT). With my colleague, Ana (thank you!), we did some research about this pathway and this post is the result. We hope it will help you!

[Read more…]

5 most popular pathways in 2014!

Today, I’d like to invite you to take a look at the 5 posts describing a pathway that saw the most visits on our blog in 2014. 

Just follow the links if you haven’t read them yet (or if you want to browse them again, feel free!).

[Read more…]

Immunotherapy Screening – CD137:CD137L pathway

In previous blogs, I invited you to read about the relevance of the B7-1 : CD28, B7-1 : CTLA4, the BLTA:HVEM, CD47:SIRPα , the GITR:GITRL, the CD40:CD40L and the  PD-1/PD-L1/PD-L2 pathway for immunotherapy screenings and discussed the products available to work on these pathways. Today, I will focus on the CD137:CD137L pathway.

CD137 is another co-stimulatory protein that is expressed on activated T cells. Unlike CD40:CD40L signaling, which primarily involves helper T-cells, CD137 has a crucial role in the development of cytotoxic T-cells and anti-tumor immunity. Its ligand, CD137L, is mainly expressed on antigen-presenting cells, such as activated B cells, macrophages, and dendritic cells, as well as on human tumor cells.

Co-stimulation through CD137:CD137L enhances T-cell activation, promotes the rejection of cardiac allografts and skin transplants, and eradicates experimentally induced tumors in animal models. Several clinical studies are on-going that use agonistic anti-CD137 antibodies to induce an anti-cancer response to solid tumors. [Read more…]

Immunotherapy Screening – CD40:CD40L pathway

In my previous blogs, I invited you to read about the relevance of the B7-1 : CD28, B7-1 : CTLA4, the BLTA:HVEM, CD47:SIRPα , the GITR:GITRL and the PD-1/PD-L1/PD-L2 pathway for immunotherapy screenings and discussed the research products available to study these pathways. Today, I’d like to focus on the CD40:CD40L pathway.

[Read more…]

Immunotherapy Screening – GITR:GITRL pathway

In previous blogs, I invited you to read about the relevance of the B7-1 : CD28, B7-1 : CTLA4, the BLTA:HVEM, CD47:SIRPα , and the  PD-1/PD-L1/PD-L2 pathway for immunotherapy screenings and discussed the products available to work on these pathways. Today, let’s focus on the GITR:GITRL pathway (glucocorticoid-induced tumor-necrosis-factor-receptor-related protein).

The GITR:GITRL pathway is a very important target for immunotherapy drug discovery — in fact, the National Cancer Institute (USA) identified GITR modulation as one of the top 25 most promising research areas! GITR is expressed on most immune cells, including T-cells, B-cells, macrophages, and NK cells. Binding of GITR by the GITR ligand (found on antigen-presenting cells and many tumors) activates the immune response by stimulating macrophages and triggering T-cells to expand, proliferate, and differentiate. Using anti-GITR mAb to trigger GITR signaling is effective in treating viral, bacterial, and parasitic infections, as well in boosting immune response against tumors. Not surprisingly, clinical trials testing anti-GITR mAb in melanoma patients are already in progress (1).GITR - GITRL Figure

Of course, it’s never that simple. Stimulating GITR signaling in NK cells has the opposite effect compared to T-cells and macrophages—GITR signaling inhibits NK cell activity (2). Likewise, stimulation of T-suppressor cells by GITR signaling also down-regulates the immune response.   Researchers are using this to their advantage; down-regulation of the immune response can be used to treat autoimmune and inflammatory diseases, including arthritis, allergy, inflammatory bowel diseases, and graft rejection. Therefore, GITR-Fc fusion protein and mAb that block GITR signaling instead of stimulating the pathway are also being studied as potential drug candidates.

BPS Biosciences recently released a set of products linked to immunotherapeutically relevant targets, amongst them the human glucocorticoid-induced TNF-related ligand (GITRL), also known as AITRL, CD357L, or tumor necrosis factor ligand superfamily member 18 (TNFSF18).

If you are interested in this specific ligand or any other of the topics and reagents covered in previous blogs (there are links to them at the beginning of this post), don’t hesitate to get in touch (you can use the form below).

References:

(1) Nocentini, G., et al., Br J Pharmacol. 165(7): 2089-99 (2012)

(2) Barao, I., Front Immunol. 3: 402 (2012)

Immunotherapy Screening – A focus on PD-1/PD-L1/PD-L2 Pathway in drug discovery

The treatment of diseases by inducing, enhancing, or surpressing an immune response is referred to as Immunotherapy. T-cell activation and inactivation requires the coordination of various co-inhibitory and co-stimulatory signals which are modulated by most immunotherapeutic approaches.

Therapeutic manipulation of immunopathways has lead to promising clinical results for the treatment of a number of diseases (ex. cancer, autoimmune diseases and inflammatory diseases…). Research projects in this field are rapidly evolving as scientists seek to identify the next generation of therapies.

In an earlier post on this blog, I introduced the latest advances regarding the the B7-1 : CD28 and B7-1 : CTLA4 pathways for immunotherapy screenings. Today, I’d like to focus on the PD-1/PD-L1/PD-L2 pathway.

After a brief review of the role of this pathway in immune response, let’s take a look at some of the screening tools I’ve selected (by our partner BPS Biosciences) to investigate this pathway for drug discovery applications. Interestingly, some of these tools are the first ready-to-use assay kits made available for drug discoverers to screen for inhibitors of the PD-1/PD-L1/PD-L2pathway.

Why look at PD-1/PD-L1/PD-L2 pathways in immunotherapeutics?

PD-1 is a receptor present on the surface of activated T-cells and other immune cells.  PD-1 has two ligands, PD-L1 and PD-L2, which are overexpressed in most human cancers.  Binding of PD-L1 to PD-1 negatively regulates T cell signaling and inhibits cytotoxic T-cell activity.

PD1 - PD-L1 - PD-L2

Similarly, engagement of PD-1 by PD-L2 dramatically inhibits T-cell proliferation, especially in CD4+ (T helper) cells. Therefore, this upregulation of PD-L1 and PD-L2 may allow cancers to evade the host immune system.

Neutralizing antibodies can prevent PD-L1 from binding to PD-1 or to B7-1. Blockade of PD-L1 enables the activation of T-cells, restoring their ability to detect and attack tumor cells. Therefore a number of pharmaceutical companies are actively developing monoclonal antibodies targeting PD-1 and PD-L to boost the immune system for the treatment of non-small-cell lung cancer, melanoma, and bladder, renal, and triple-negative breast cancers.

The PD-1/PD-L pathway is also a therapeutic target for chronic viral-infections (including HIV and HCV). This therapeutic appraoch is based on the observation that PD-1 expression is upregulated by virus-specific T cells and that the inhibition of this loop is hoped to decrease viral load by increasing  T cell function. Conversely, since the PD-1/PD-L interaction regulates self-tolerance and immunologic homeostasis, agonists of these proteins are promising drug targets for many autoimmune disorders including multiple sclerosis, arthritis, lupus, and type I diabetes.

The most advanced PD-1/PD-L1/PD-L2 screening reagents

These new therapeutical advances highlight the importance of reliable R&D tools in Drug discovery approaches and translation medicine programs. Access to robust research reagents covering the needs of Drug discoverers from target validation to screening (HTS or secondary) or even DMPK studies is crucial. Below, I’ve made a selection of some of the latest releases for scientists interested in the PD-1/PD-L1/PD-L2 pathway.

         #1 – PD-1  – a receptor for PD-L1

PD-1 is the receptor for PD-L1. Formation of the receptor-ligand complex leads to an inhibitory signal which reduces proliferation of CD8x T cells.

PD-1 is available as a Fc Fusion (Id. nr 71106), as a FLAG-tagged variant (Id. nr 71115), and as a Biotin labeled variant (Id. nr 71109)

        #2 – PD-L1 – a ligand for PD-1 receptor

PL-L1 is one of the ligands for PD-1, which plays a role in surpressing the immune response especially during pregnancy, tissue allografts, autoimmune diseases and diseases such as Hepatitis.

PL-L1 is available as a non labeled variant (Id. nr 71104) and a Biotin-labeled variant (Id. nr 71105).

        #3 – PD-L2 –  aligand for PD-2 receptor

PL-L2 is another ligand for PD-1 which, upon complex formation T cell proliferation, especially of CD4+ (T helper) cells, is dramatically decreased.

PL-L2 is available as a non labeled variant (Id. nr 71107) and a Biotin-labeled variant (Id. nr 71108).

        #4 – PD1-neutralizing antibody

This antibody (Id. nr 71120) can be used as a control inhibitor using the PD-1:PD-L1[Biotinylated] Inhibitor Screening Assay Kit (see below).

Ready-to-use PD-1/PD-L1/PD-L2 screening assay kits

Recently, BPS Biosciences have released 3 unique ready-to-use assay kits to screen and profile inhibitors for the PD-1/PD-L1/PD-L2 signaling pathways

PD-1:PD-L1[Biotinylated] Inhibitor Screening Assay Kit (Id. nr 72003)

PD-1:PD-L2[Biotinylated] Inhibitor Screening Assay Kit Id. nr 72004)

Both are 96-well format assay kits and come with biotinylated PD-L1 or PD-L2, purified PD-1,  streptavidin labeled HRP for 100 binding reactions.

PD-L1-PD-1 Interaction measured by Luminescence

Typical data obtained with the BPS’ Ready-to-use Inhibitor Screening assays.

PD-L2 Inhibitor Screening Assay Kit (Id. nr 72006).

This 96-well format assay kits comes with biotinylated PD-1, purified PD-L2,  streptavidin labeled HRP  for 100 binding reactions.

Looking for PD-1/PD-L1/PD-L2 testings?

Choosing the right reagents or screening kits to raise the value of your compounds of interest by looking to PD-1/PD-L1/PD-L2 pathway can be challenging.

Research tools reviewed in this second post (belonging to a series of 3 posts dedicated to  Immunotherapy screening) will definitely help you there. Otherwise, you can simply contact me with the form below.

Recently released Immunotherapy Screening posts on the blog “being bio-reactive”:

Upcoming Immunotherapy Screening posts on the blog “being bio-reactive”:

  • Immunotherapy Screening – Part 3:  BLTA:HVEM and CD47:SIRPalpha Pathways

Immunotherapy Screening – B7-1 / CD28 and B7-1 / CTLA4 Pathways in Drug Discovery

The treatment of diseases by inducing, enhancing, or surpressing an immune response is referred to as Immunotherapy. T-cell activation and inactivation requires the coordination of various co-inhibitory and co-stimulatory signals and most immunotherapies modulate these signals.

Therapeutic manipulation of immunopathways has lead to promising clinical results for the treatment of a number of diseases such as cancer, autoimmune diseases and inflammatory diseases. Research in this field is rapidly evolving as scientists seek to identify the next generation of therapies.

In the first post of a series of three,  I will describe the B7-1 : CD28 and B7-1 : CTLA4 pathways and summarize the portfolio of tools currently available to researchers to investigate these pathways. In fact, these are the first kits on the market to screen for inhibitors of these pathways. [Read more…]