Neuroendocrine (NE) cancers are tumours whose cells express features of neuroendocrine cells, and they include aggressively invasive and lethal cancers such as small-cell lung cancer (SCLC). In addition to the many types of primary NE cancer, breast and prostate cancers can also evolve towards a small-cell NE phenotype during progression and metastasis. Improved understanding of NE cancers and new therapeutic strategies are urgently needed. Intriguingly, unlike other cancer types, NE cancer cells are electrically excitable, firing action potentials like neurons. They also express synaptic receptor ion channels, secrete neurotransmitters and peptides, and often metastasize to the brain. Here, I discuss our recent work on the electrophysiology of two NE cancer types: pancreatic neuroendocrine tumours (PNETs) and small-cell lung cancer (SCLC), and the significance of electrical activity in their progression. PNET cells express functional NMDA-type glutamate receptor ion channels - well-known as mediators of excitatory synaptic transmission and its long-term potentiation in the central nervous system. These channels are opened by a continuous autocrine release of glutamate, and also powerfully by necrosis of nearby cells, and the resulting calcium influx drives downstream signalling which promotes invasiveness. SCLC tumours actually contain two distinct classes of cancer cells: excitable NE and inexcitable non-NE. These two cell types appear to collaborate in a metabolic symbiosis, whereby the high energetic demand of electrical activity in the NE cells is met by secretion of lactate from non-NE cells, in a process analogous to the “lactate shuttle” between neurons and astrocytes.
Automated electrophysiology platforms like the Sophion Qube 384 and SURFE2R are transforming drug discovery by enabling high-throughput analysis of ion channel activities. This presentation outlines their role in early safety profiling and target-based screening of ligand-gated ion channels, emphasizing efficient detection of cardiotoxic and neurotoxic risks to minimize late-stage attrition. The discussion will also cover strategic approaches in screening, enhancing the identification and characterization of potent, selective, and safe modulators. In addition, we will explore their utility in mechanistic studies, providing deeper insights into ion channel function and pharmacology. By streamlining these processes, automated electrophysiology improves the predictability of drug efficacy and safety, expediting the development of new therapeutics.
To follow
Christian Grimm1,2
1Walther Straub Institute of Pharmacology and Toxicology, Faculty of Medicine, Ludwig-Maximilians-University, Munich, Germany.
2Immunology, Infection and Pandemic Research IIP, Fraunhofer Institute for Translational Medicine and Pharmacology ITMP, Munich/Frankfurt, Germany
Research on endolysosomal ion channels and transporters has gained considerable momentum in recent years, as new technologies are now available to investigate the function and physiology of these important intracellular membrane proteins in more detail. For example, endolysosomal patch-clamp electrophysiological techniques, intracellular sensors for endolysosomal Ca2+ and pH measurements or new animal models have been developed. Using these new techniques, it has been discovered that endolysosomal cation channels such as TRPML channels or two-pore channels (TPCs) play important physiological roles in a variety of organs and that defects or alterations in their function are associated with lysosomal storage diseases and neurodegenerative diseases, infectious diseases, immune cell dysfunction, lung, liver and heart diseases as well as cancer (e.g. melanoma). Thanks to significant advances also in the availability of pharmacological tools for these endolysosomal membrane proteins, accompanied by numerous cryo-EM structures research in this area is progressing rapidly. The current lecture will present recent and current (unpublished) findings on the physiological and pathophysiological role of TRPML channels and TPCs with a particular focus on its suitability as drug targets.
This work was supported, in part, by funding of the German Research Foundation (SFB/TRR152 P04, SFB1328 A21, GRK2338 P08, DFG GR4315/2-2, DFG GR4315/6-1 and DFG GR4315/7-1).
Selected publications: Xu M et al. & Grimm C#, Dong, X#: TRPML3/BK complex promotes autophagy and bacterial clearance by providing a positive feedback regulation of mTOR. PNAS 20(34):e2215777120, 2023. • Scotto Rosato A et al. & Grimm C#: TPC2 rescues lysosomal storage in mucolipidosis type IV, Niemann-Pick type C1 and Batten disease. EMBO Mol Med 5:e15377, 2022. • Spix B et al. & Grimm C#: Lung emphysema and impaired macrophage elastase clearance in mucolipin 3 deficient mice. Nature Commun 13(1):318, 2022. • Chen C-C et al. & Grimm C#: TRPML2 is an osmo-/mechano-sensitive cation channel in endolysosomal organelles. Science Adv 6(46):eabb5064, 2020. • Chao Y-K et al. & C, Grimm C#: TPC2 polymorphisms associated with a human hair pigmentation phenotype result in gain of channel function by independent mechanisms. PNAS 114:E8595-E8602, 2017. • Chen C-C et al. & Grimm C#: Patch clamp technique to characterize ion channels in individual intact endolysosomes. Nature Protoc 12:1639-1658, 2017. • Sakurai Y, Kolokoltsov AA, Chen C-C, Tidwell MW, Bauta WE, Klugbauer N, Grimm C, Wahl-Schott C, Biel M, Davey RA: Two pore channels control Ebolavirus host cell entry and are drug targets for disease treatment, Science 347:995-998, 2015. • Chen C-C et al. & Grimm C#: A small molecule restores function to TRPML1 mutant isoforms responsible for mucolipidosis type IV. Nature Commun 5:4681, 2014. • Grimm C et al. & Wahl-Schott C: High susceptibility to fatty liver disease in two-pore channel 2-deficient mice. Nature Commun 5:4699, 2014.
To follow
Much evidence points to lysosomes as non-canonical stores of Ca2+. This extends the role of lysosomes to signalling processes. A number of Ca2+-permeable lysosomal ion channels have been identified and shown to regulate numerous functions including many aspects of membrane trafficking. Malfunction of these channels is linked to a growing number of diseases. In this talk, I will cover recent advances in understanding how two-pore channels (TPCs) control flux of Ca2+ and other ions across the lysosomal membrane. The picture that emerges is an unusual one in which TPCs are able to switch their ion selectivity in an agonist dependent manner to differentially control lysosomal function.
The TMEM16 proteins constitute a family of membrane proteins, which comprises lipid scramblases and Cl- channels that are involved in diverse physiological processes. Whereas TMEM16 channels contribute to Cl- secretion in airway epithelia and electrical signaling in smooth muscles and certain neurons, TMEM16 scramblases participate in blood clotting and the fusion of trophoblasts, osteoclasts and myoblasts. Members of both functional branches share a common architecture and are activated by intracellular Ca2+ by a similar mechanism. TMEM16 proteins form homodimers of subunits that are composed of ten membrane-spanning helices. The subunits are functionally independent and contain a regulatory Ca2+-binding site embedded within the transmembrane domain and a close-by site of catalysis located at the periphery of the protein, which either facilitates ion or lipid permeation.
In the fungal lipid scramblase nhTMEM16, Ca2+-binding opens a hydrophilic membrane-spanning furrow, which is hidden in the Ca2+-free protein. This furrow provides a pathway for lipid headgroups to move between both leaflets of the bilayer. In contrast, in the anion channel TMEM16A, Ca2+-binding triggers the opening of a protein-enclosed ion conduction pore located in the same region, which remains shielded from the membrane in its activated state. In this case, Ca2+ serves a dual role in promoting a conformational change to release a gate that impedes conduction in the closed state and by shaping the electrostatics to enhance anion permeation. Finally, in the lipid scramblase TMEM16F, we find both functions as lipid scramblase and ion channel contained within the same protein. Despite its close relationship to the anion channel TMEM16A, in its activated state TMEM16F assumes a structure that is distinct from either nhTMEM16 or TMEM16A, which presumably catalyzes both processes in a single conformation.