Programming GPCR signalling within the endocytic network; mechanisms and therapeutic applications

Cells communicate to each other by sending and responding to chemical messages. Coordinating this communication is essential for correct functioning of every organ in the body. These messages include photons of light, chemicals that we smell, the food that we eat, mineral ions such as calcium, hormones and neurotransmitters (chemical messengers of the brain). The chemical messages are received when they bind to specific proteins on the cell surface called receptors, which relay the message in to the cell. Our research is focussed on a family of receptors called G protein-coupled receptors (GPCRs). Our genes encode for more than 800 different kinds of GPCRs that are capable of responding to numerous different messages. Each organ in the body has many different GPCRs. Importantly, the function of these receptors is disrupted in various diseases and disorders, including cancer, obesity, diabetes, blindness, heart disease, depression, Parkinson's Disease, recurrent miscarriage and pre-term birth, to name but a few. Although many prescribed drugs target GPCRs, there is a high demand for new compounds that are more specific, have fewer side effects, and that are active for longer. Developing these new drugs requires an in-depth understanding of the molecular machinery that controls the activity these receptors.

Once any cell receives external messages that bind to GPCRs, the activated receptors will relay specific signals to elicit an appropriate cellular response. This process is tightly controlled by the cell. One important mechanism is the rapid removal, or trafficking, of receptors from the cell surface in to interior compartments called endosomes, as a means of switching off these signals. However, we discovered that certain GPCRs recruited to specialized endosomes, which we termed very early endosomes (VEE), can switch on new signalling pathways. So, both duration and location of signals generated by GPCRs creates patterns that are critical for the cell, telling it whether to release more chemicals, activate particular genes, divide, or even die. If such signalling patterns are disrupted, or misdirected, they can lead to disease. However, our discovery also raises the possibility that drugs can be developed to redirect the receptor and change its function in a cell.

The aim of this project is to understand how a cell controls receptor activity by examining the molecules involved in trafficking and signal decoding of GPCRs from the VEE. Our recent work also revealed that trafficking to the VEE of a GPCR important in reproduction and in early pregnancy (the LH receptor) may be important in how the uterus responds to hormones produced by the embryo in early pregnancy, and that these pathways may be altered in women suffering recurrent miscarriage. We will also assess the role of this compartment to other GPCRs by studying a receptor important in sensing carbohydrates in our food (the FFA2 receptor) and another that is targeted in IVF, cancer and menopause (the FSH receptor). For FSH and LH receptor we will determine whether drugs can alter the trafficking of specific GPCRs to the VEE or other endosomal compartments, and in turn impact on receptor activity. The outcome of this work will help us understand fundamental mechanisms of how cells communicate. As GPCRs are common drug targets, detailed knowledge of how GPCRs are regulated by the VEE may even provide new avenues for more effective treatments of a number of conditions that involve this superfamily of receptors.

Membrane trafficking of receptors represents a fundamental mechanism to translate complex signalling networks into specific downstream responses. This is the case for numerous cellular signalling systems, including the pathways activated by the superfamily of G protein-coupled receptors (GPCRs). Thus, a detailed molecular description of how membrane trafficking pathways are indispensable in controlling GPCR signalling are key in the fundamental understanding of cellular regulation and how aberrant signalling can result in disease. This an area of precedence in biology and medicine due to the ongoing search to identify GPCR modulators with high specificity. We have identified a novel endosomal compartment critical for the sorting and signalling of distinct GPCRs, termed the very early endosome (VEE). Using a model GPCR for the VEE, the luteinizing hormone receptor (LHR), this project will identify the fundamental mechanisms spatially encoding GPCR activity at the VEE and its downstream roles. The adaptor protein APPL1 is so far, the only VEE protein known to regulate sorting from the VEE and endosomal GPCR signalling. Spatially encoded signalling from the VEE has a critical impact on downstream cellular responses. Altering LHR trafficking to the VEE in the human endometrium is linked to impaired decidualization (cell differentiation) with implications in recurrent miscarriage. Other GPCRs, including the FSH receptor and short chain fatty acid receptor FFA2, also require internalization to VEE/APPL1 endosomes for their function. Employing super-resolution imaging, coupled with quantitative proteomic profiling, and biochemical signal analysis in heterologous and primary human endometrial 2D and 3D cultures, we will identify the core molecular machinery directing regulated sorting and signalling from this novel compartment, profile the GPCR signal pathways that are spatially encoded by the VEE, and identify impact both physiologically and pharmacologically.

Who might benefit from this research?
This project aims to address fundamental questions of how G protein-coupled receptor (GPCR) signal diversity is translated to specific cellular and physiological responses. This an area of precedence in biology and medicine due to the intense interest and demand in identifying GPCR modulators with high specificity. Thus, identified beneficiaries outside the academic arena are primarily those in industry and the commercial sector. GPCRs are the most successfully exploited class of drug targets, and their role in pathogenesis of human disease maintains the strong interest in new basic science discoveries by these sectors. In addition to companies investing in targeting these receptors (Merck, Pfizer, Heptares, Trevena Inc, Ferring, GSK, TocopheRx, EMD Serono), new basic science findings in GPCR research are of high interest across this sector for other GPCRs and commercial sectors creating GPCR screening tools for research (e.g. DiscoveRx, CisBio). This work could have an impact long-term for clinical translation and public health, but also on the UK economy by reducing financial burdens of the NHS (in addition to commercialization). This work has also identified those involved in Education as a beneficiary, from the training of the postdoctoral posts on this project to secondary schools, information resource of public and patient education directly or via Charities.

How might they benefit from this research?
GPCRs are an important target for therapeutic intervention and many successful drugs on the market today modulate their activity. The research proposed has the potential to provide information for design of novel screening platforms, which could be developed commercially as an assay, used in industry or academic labs. Also in identifying strategic targets that feature defined modulation of receptor activity, rather than global agonists/antagonists. A rapidly growing area of high interest in the drug discovery industry is to design GPCR compounds that show bias towards specific receptor activities. Although we will focus on specific model GPCRs for these pathways, the fundamental nature of the findings from our proposed project will uncover mechanisms that will be pertinent to other GPCRs and associated conditions involving these receptors. We intend to establish advisory panel meetings with members from Industry, Academia and Third sector, throughout the project and mediated by Imperial's Corporate and Enterprise Partnerships Team, that will facilitate in steering the path to commercialization and knowledge exchange platforms for stakeholders. In addition, current industry collaborators of the PI and Co-PIs will be contacted during the award in addition to engagement with the Imperial College Cross-Faculty Centre for Drug Discovery Science, led by Co-PI (E.T). If Industrial collaboration and/or commercial exploitation are successful this could escalate in to long-term benefits in UK economy from revenue, employment and ultimately in health benefits. The Co-PI (J.B) will enable translational opportunities by forming links with clinicians and awareness opportunities for patients and their families e.g. via Tommy's highly successful social media campaign. Imperial College London and University of Warwick also have strong commitments to both engaging general public and in education in Secondary schools. Training that the posts will receive would be beneficial for any employment sector by the development of transferable skills. These include communication skills, project and time management, problem solving, public engagement, information technology and mentoring.