1. Biochemistry and Chemical Biology
  2. Cell Biology
Download icon

Bone-Targeting Drugs: From vesicle to cytosol

  1. Michael J Rogers  Is a corresponding author
  2. Marcia A Munoz
  1. Garvan Institute of Medical Research, Australia
Insight
  • Cited 4
  • Views 1,059
  • Annotations
Cite this article as: eLife 2018;7:e38847 doi: 10.7554/eLife.38847

Abstract

Drugs called bisphosphonates are used to treat a range of bone diseases, but how do they reach the enzymes that are their target?

Main text

Despite its appearance, bone is a highly metabolic and dynamic tissue that is composed of a vast network of cells called osteocytes that are embedded in a matrix made mostly of collagen and various salts of calcium and phosphate. These osteocytes sense regions of damaged or weakened bone, and 'instruct' bone-destroying cells (called osteoclasts) and bone-forming cells (osteoblasts) to, respectively, remove old bone and deposit new bone (Figure 1). Hence, like a team of road-repairers, the osteocytes, osteoclasts and osteoblasts work together to repair bone and maintain our skeleton in good health (Crockett et al., 2011).

How bisphosphonates act in bone.

(A) Old and damaged bone is constantly being broken down by cells called osteoclasts in a process called resorption (top left), while new bone is deposited by cells called osteoblasts (top right). Cells called osteocytes (bottom) influence both of these processes through a spidery system of tiny canals called canaliculi. After entering the circulation, the drug bisphosphonate (green) binds very effectively to calcium ions on the bone mineral surface. During resorption, the bisphosphonate on the bone surface is released into the acidic extracellular space beneath the osteoclast. (B) The bisphosphonate in the extracellular space is engulfed into osteoclasts via a process called endocytosis (1). The resulting endosomes mature to form structures called lysosomes, and two proteins, SLC37A3 and ATRAID, then interact in the membrane of the lysosome to allow the bisphosphonate to enter the cytosol (2). Once in the cytosol, the nitrogen-containing bisphosphonates inhibit an enzyme called FDPS and prevent the osteoclast from breaking down bone (3). BP: bisphosphonate; FDPS: farnesyl diphosphate synthase; SLC37A3: solute carrier family 37 member A3.

In young adult life there is usually a balance between the amount of old bone broken down and the amount of new bone formed by this repair process, so there is no net gain or loss of bone mass. However, in diseases that affect the skeleton, such as post-menopausal osteoporosis or cancers growing in bone, this delicate balance can be disturbed by osteoclasts being over-active, which leads to excessive bone destruction and fractures. Drugs called bisphosphonates – which inhibit osteoclasts – have been used for more than three decades to treat such diseases and protect the skeleton from potentially catastrophic bone loss, although researchers still do not fully understand how they work. Now, in eLife, Erin O'Shea of Harvard University and the Howard Hughes Medical Institute (HHMI) and colleagues – including Zhou Yu as first author – report the answer to one of the remaining questions about these drugs (Yu et al., 2018).

Bisphosphonates are synthetic molecules that closely resemble the chemical structure of pyrophosphate, which is a natural by-product of numerous metabolic reactions. Importantly, bisphosphonates have two negatively-charged phosphonate groups that enable them to bind calcium ions very effectively, and hence to localize rapidly to any exposed calcium on the bone surface (Rogers et al., 2011). The mechanisms used by bisphosphonates to inhibit osteoclasts remained a mystery for several decades after they were first used in the clinic, but this did not stop the development of improved versions of the drugs (Russell et al., 2008). Eventually it was discovered that nitrogen-containing bisphosphonates (N-BPs), which are now widely used to treat osteoporosis and other bone diseases, work by inhibiting an enzyme called FDPS inside the osteoclasts (Luckman et al., 1998; Bergstrom et al., 2000; Dunford et al., 2001). The N-BP molecules displace the lipid substrates that the FDPS enzyme usually acts on, locking the enzyme in an inactive state (Kavanagh et al., 2006; Rondeau et al., 2006). Without FDPS activity, osteoclasts are no longer able to degrade bone (Rogers et al., 2011).

However, one question remained: how do the N-BPs and other bisphosphonates actually reach the FDPS enzyme, which is in the cytosol of the osteoclasts? There was little or no evidence that a receptor on the plasma membrane was involved (Thompson et al., 2006). Studies with fluorescently-tagged bisphosphonates showed that they first entered the osteoclasts via endocytosis – a process that involves the cell membrane folding inwards and then pinching off to create a vesicle inside the cell (Coxon et al., 2008). But how do the drugs leave these vesicles – which are enclosed by a membrane – to enter the cytosol? Bisphosphonate molecules have a large negative charge, which rules out passive diffusion across the vesicle membrane, which in turn suggests the possibility of a hitherto unidentified transport mechanism (Thompson et al., 2006).

Yu et al. – who are based at Harvard, HHMI, UCSF, MIT, the Broad, Koch and Whitehead Institutes, and Washington University – report that they have identified a protein called SLC37A3 that is required for the release of N-BP molecules from vesicles into the cytosol. Using a CRISPR-based approach to screen for genes that, when missing, confer resistance to bisphosphonates, they identified SLC37A3 as the gene with the strongest effect. Although the exact function of the SLC37A3 protein remains to be clarified, related members of this protein family are involved in the transport of charged molecules across membranes (Cappello et al., 2018).

Yu et al. found that SLC37A3 interacts and co-localizes with a protein called ATRAID at the vesicle membrane (Figure 1). Importantly, vesicles isolated from cells that did not express SLC37A3 or ATRAID appeared unable to release N-BP molecules, and these cells were much less sensitive to the pharmacological effect of N-BPs.

This new transport mechanism identified by Yu et al. raises interesting questions about how the SLC37A3/ATRAID complex specifically recognizes N-BP molecules, and how it transports them across the membrane of the vesicle. It will also be worthwhile to determine whether differences in the expression of SLC37A3 or ATRAID account for the different sensitivity of osteoclasts and other cell types to N-BP molecules, or whether variants in these genes affect the clinical responsiveness of patients to these drugs. Nevertheless, these elegant studies explain how negatively-charged N-BP molecules can gain access to the cell cytosol after endocytosis and, as a result, go on to benefit huge numbers of patients with potentially devastating bone diseases.

References

  1. 1
  2. 2
  3. 3
  4. 4
  5. 5
    Structure-activity relationships for inhibition of farnesyl diphosphate synthase in vitro and inhibition of bone resorption in vivo by nitrogen-containing bisphosphonates
    1. JE Dunford
    2. K Thompson
    3. FP Coxon
    4. SP Luckman
    5. FM Hahn
    6. CD Poulter
    7. FH Ebetino
    8. MJ Rogers
    (2001)
    Journal of Pharmacology and Experimental Therapeutics 296:235–242.
  6. 6
  7. 7
  8. 8
  9. 9
  10. 10
  11. 11
  12. 12

Article and author information

Author details

  1. Michael J Rogers

    Michael J Rogers is in the Bone Biology Division, Garvan Institute of Medical Research, Darlinghurst, Australia

    For correspondence
    m.rogers@garvan.org.au
    Competing interests
    No competing interests declared
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-1818-9249
  2. Marcia A Munoz

    Marcia A Munoz is in the Bone Biology Division, Garvan Institute of Medical Research, Darlinghurst, Australia

    Competing interests
    No competing interests declared
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-7603-0351

Publication history

  1. Version of Record published: June 27, 2018 (version 1)

Copyright

© 2018, Rogers et al.

This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 1,059
    Page views
  • 108
    Downloads
  • 4
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, PubMed Central, Scopus.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Download citations (links to download the citations from this article in formats compatible with various reference manager tools)

Open citations (links to open the citations from this article in various online reference manager services)

Further reading

    1. Biochemistry and Chemical Biology
    2. Cell Biology
    Haibin Yang et al.
    Research Article Updated

    Communications between actin filaments and integrin-mediated focal adhesion (FA) are crucial for cell adhesion and migration. As a core platform to organize FA proteins, the tripartite ILK/PINCH/Parvin (IPP) complex interacts with actin filaments to regulate the cytoskeleton-FA crosstalk. Rsu1, a Ras suppressor, is enriched in FA through PINCH1 and plays important roles in regulating F-actin structures. Here, we solved crystal structures of the Rsu1/PINCH1 complex, in which the leucine-rich-repeats of Rsu1 form a solenoid structure to tightly associate with the C-terminal region of PINCH1. Further structural analysis uncovered that the interaction between Rsu1 and PINCH1 blocks the IPP-mediated F-actin bundling by disrupting the binding of PINCH1 to actin. Consistently, overexpressing Rsu1 in HeLa cells impairs stress fiber formation and cell spreading. Together, our findings demonstrated that Rsu1 is critical for tuning the communication between F-actin and FA by interacting with the IPP complex and negatively modulating the F-actin bundling.

    1. Biochemistry and Chemical Biology
    Rajendra Uprety et al.
    Research Article Updated

    Controlling receptor functional selectivity profiles for opioid receptors is a promising approach for discovering safer analgesics; however, the structural determinants conferring functional selectivity are not well understood. Here, we used crystal structures of opioid receptors, including the recently solved active state kappa opioid complex with MP1104, to rationally design novel mixed mu (MOR) and kappa (KOR) opioid receptor agonists with reduced arrestin signaling. Analysis of structure-activity relationships for new MP1104 analogs points to a region between transmembrane 5 (TM5) and extracellular loop (ECL2) as key for modulation of arrestin recruitment to both MOR and KOR. The lead compounds, MP1207 and MP1208, displayed MOR/KOR Gi-partial agonism with diminished arrestin signaling, showed efficient analgesia with attenuated liabilities, including respiratory depression and conditioned place preference and aversion in mice. The findings validate a novel structure-inspired paradigm for achieving beneficial in vivo profiles for analgesia through different mechanisms that include bias, partial agonism, and dual MOR/KOR agonism.