Facebook
Twitter
LinkedIn
Google+The Lysosome Group
Our main goal is to identify lysosomal cancer targets and develop novel lysosome-based cancer therapies.
Who are we?
Current researchers:
Atul Anand, MSc, Line Christoffersen, MSc, Anne Marie Ellegaard, PhD, Elena Favaro, PhD, Line Groth-Pedersen, PhD, Saara Hämälistö, PhD, Bin Liu, PhD, Mikkel Rohde, PhD, and Marja Jäättelä, MD PhD
Technicians:
Dianna Skousborg Larsen, Tiina Naumanen, Louise Bro, and Louise Vanderfox
Former postdocs (since 2000):
Sonja Aits, Carla Cardoso, Jennifer Kricker, Nikolaj Havnsøe Torp Pedersen
Former PhD students (since 2000):
Annika Baude, Mads Daugaard, Nicole Fehrenbacher, Lasse Foghsgaard, Mads Gyrd-Hansen, Thomas Kirkegaard, Ulrik Lademann, Jesper Nylandsted, Ida Stenfeldt Mathiasen, Ole Dines Olsen, Marie Stampe Ostenfeld, Dorte Wissing
Why lysosomes?
The central genes controlling evolutionary conserved apoptotic cell death program (i.e. caspases and Bcl-2) were discovered in the late 1980s and early 1990s. During this apoptosis boom, Marja Jäättelä studied tumor necrosis factor-induced cell death in fibrosarcoma cells but failed to detect markers of apoptosis in the dying cells. This non-apoptotic cell death caught her attention and she decided to figure out how those cancer cells were dying. That gave rise to a long and rocky road of trials and errors, which some ten years later resulted in the identification of lysosomal membrane permeabilization as an alternative cell death mechanism in cancer cells.
At that time, apoptosis resistance was emerging as a major hurdle in cancer treatment, and thus the next obvious step was to find cancer-specific ways to permeabilize lysosomes and thereby circumvent apoptosis resistance. In order to do that, Marja Jäättelä established the Lysosome Group, whose research aims and achievements are described below.
Background
Lysosomes are membrane-enclosed acidic organelles found in all mammalian cells except for mature erythrocytes. They are responsible for the digestion and recycling of cellular macromolecules and organelles as well as extracellular material delivered to them by autophagy and endocytosis (Figure 1).
Figure 1. Multiple lysosomal functions in cancer cells.
The main function of the lysosomes is the degradation of cargo derived by autophagy (see Autophagy Group) and endocytosis. They can also fuse with the plasma membrane during cell injury, and have more specialized secretory functions (lysosomal exocytosis) in some cell types including e.g. osteoclasts and invasive cancer cells. In the extracellular space lysosomal hydrolases can promote cancer progression by multiple means (see Signaling Group). Oncogene activation can lead to the destabilization of lysosomal membranes and increased sensitivity to the leakage of lysosomal hydrolases into the cytosol, where they can contribute to the demise of the cancer cell (see text for details). Upon receptor tyrosine kinase activation endolysosomal compartment regulates the recycling of receptors and their ligands. ECM, extracellular matrix; MVB, multivesicular body; RTK, receptor tyrosine kinase. (Kallunki et al, Oncogene 2012)
The digestion of the cargo is brought about by over 50 hydrolases (proteases, glycosidases, phosphatases, sulfatases, nucleases and lipases) that normally reside in the lysosomal lumen and function optimally in the acidic pH. Notably, lysosomal hydrolases can also perform important extralysosomal functions e.g. in cell death as discussed below (Figure 2), and lysosomes have recently been identified as a central hub in the control of cellular metabolism.
Figure 2. Lysosomal cell death pathways.
Numerous treatments trigger lysosomal membrane permeabilization (LMP) either directly or indirectly resulting in the release of lysosomal content into the cytosol. Here, lysosomal hydrolases, especially cathepsins, can mediate cell death either in a mitochondrion-independent manner or through cleavage-mediated activation of proapoptotic Bcl-2 family members and subsequent release of apoptogenic factors (e.g. cytochrome c and AIF) from the mitochondria. (Groth-Pedersen et al., Cancer Lett 2010)
The role of lysosomes and lysosomal hydrolases in cell death was introduced already almost 60 years ago by Christian de Duve, a Belgian scientist who was awarded the Nobel prize for his discovery and characterization of lysosomes. Due to the potent hydrolytic capacity of lysosomal hydrolases, de Duve defined lysosomes as "suicide bags" that can cause cell death and tissue damage upon rupture and subsequent release of lysosomal hydrolyses to the cytosol and the extracellular space.
This definition triggered an intensive search for pharmaceutical agents that either stabilize or destabilize lysosomal membranes for the treatment of degenerative disorders and cancer, respectively. Interest in lysosomal cell death pathways waned, however, rapidly. This was largely due to the lack of appropriate assay systems that could differentiate lysosomal rupture that causes cell death from postmortal alterations in autolytic cells as well as the fear that lysosomotropic detergents would be equally toxic to normal and transformed cells. Accordingly, novel more sensitive assays to study lysosomal membrane permabilization and our data indicating that cancer cell lysosomes are less stable than normal lysosomes (Figure 3) were needed to initiate a new wave of interest in lysosomal cell death pathways in the beginning of the 21st century.
Figure 3. Transformation-associated changes in lysosomal composition.
The transformation process imposes changes in the expression and/or activity of lysosomal proteins. Some of these changes are "pro-tumor"-like processes contributing to tumor growth, invasion, and angiogenesis, and some modulate "anti-tumor" mechanisms by increasing lysosomal membrane permeabilization that can lead to lysosomal destabilization and cell death. These changes open several windows for anti-cancer therapy as discussed in the text. (Kallunki et al., 2012)
Research aims
Our main goal is to identify lysosomal cancer targets and develop novel lysosome-based cancer therapies. For this purpose, we study the cancer-associated changes in the lysosomal protein and lipid composition, lipid metabolism and lysosomal trafficking, and various genetic and small molecule screens to identify genes and compounds that alter lysosomal membrane stability. Most of the work is performed in established cancer cell lines and tumor xenografts in mice, but we have recently also initiated programs for the study of the lysosomal compartment in acute lymphatic leukemia and acute myeloid leukemia patient samples as well as in colon cancer initiating cells. In addition to the various research projects, a large part of our work is devoted to the development of novel methods to study the lysosomal stability and function.
Our Main Contributions
- Tumor necrosis factor induces lysosomal cell death in various cancer cells
Lasse Foghsgaard and coworkers were the first to discover lysosomal destabilization and lysosomal cathepsins as essential mediators of tumor necrosis factor-induced killing of several cancer cells and to demonstrate that cathepsins can trigger caspase-independent cell death (Foghsgaard et al., J Cell Biol 2001). This study together with the data from Gregory Gores' group demonstrating tumor necrosis factor-induced lysosomal membrane permeabilization in murine hepatocytes initiated a new wave of interest in lysosomal cell death - Cancer cell lysosomes are less stable than normal lysosomes
Nicole Fehrenbacher and coworkers realized that the malignant transformation decreases lysosomal stability and identified enhanced lysosomal cathepsin activity and cathepsin-mediated proteolysis of lysosomal membrane proteins as underlying mechanisms (Fehrenbacher et al, Cancer Res 2004 and Cancer Res 2008) (Figure 2). The realization that cancer cell lysosomes are less stable than normal lysosomes formed the basis for our present studies aiming at the development of lysosome-targeting anti-cancer drugs and triggered a broad interest in lysosomes as targets for cancer therapy world-wide - Hsp70 is a guardian of lysosomal stability
Jesper Nylandsted, Mads Gyrd-Hansen and coworkers identified heat shock protein 70 (Hsp70) as a cancer-specific survival factor and guardian of lysosomal stability (Nylandsted et al., PNAS 2000, Cancer Res 2002 and J Exp Med 2004; Gyrd-Hansen et al., Mol Cell Biol 2006). Continuing in this line of research, Thomas Kirkegaard and coworkers showed that Hsp70 stabilizes lysosomes by binding to an endo-lysosomal anionic phospholipid bis(monoacylglycero)phosphate, an essential co-factor for lysosomal sphingomyelin metabolism, which in turn is essential for the lysosomal stability (Kirkegaard et al., Nature 2010). These data reveled lysosomal lipid metabolism as a putative target for cancer therapy and initiated our present research program on cancer-associated alterations in lipid metabolism - Lysosomotropic detergents are potential anti-cancer agents
Marie Ostenfeld and coworkers demonstrated that a lysosomotropic detergent siramesine shows cancer-specific cytotoxicity both in cell culture and in cancer xenograft models in mice (Ostenfeld et al., Cancer Res 2005 and Autophagy 2008). These data opened new possibilities for lysosomotropic detergents and related compounds in future cancer therapy and our further research identified a large group of clinically relevant compounds (antihistamines, antidepressants, antimalarials, etc.) as putative anti-cancer agents that de-stabilize lysosomes and sensitize cancer cells to chemotherapy by inhibiting lysosomal acid sphingomyelinase (Petersen et al., Cancer Cell 2013; Ellegaard et al., Mol Cancer Ther 2013). A subsequent collaboration with our in-house pharmacoepidemiology group then identified a positive association between the use of common antihistamines and reduced mortality among cancer patients (Ellegaard et al., EBiomedicine 2016). Encouraged by these data, we are presently planning clinical trials to test the effect of antihistamines in cancer treatment - Disturbance of the function of microtubules trigger lysosomal cell death
Line Groth-Pedersen, Carla Cardoso, Sonja Aits, Jesper Nylandsted and coworkers identified functional microtubules and specific microtubule- and actin-associated motor proteins as essential stabilizers of lysosomal membranes (Groth-Pedersen et al., Cancer Res 2007 and PLoS One 2012; Cardoso et al., PLoS One 2009). These data opened interesting possibilities for combination therapies consisting of microtubule-targeting drugs and lysosomotropic agents - Non-lethal functions of lysosomal membrane permeabilization
Using our recently developed and highly sensitive method to detect lysosomal membrane permeabilization (Aits et al., Autophagy 2015), Saara Hämälistö and Elena Favaro came up with an exciting hypothesis that tightly regulated lysosomal leakage could also occur in viable cells and regulate normal cellular functions including cell division and cell adhesion (unpublished; Figure 4) - Hsp70-2 and LEDGF stabilize lysosomes and enhance homologous recombination DNA repair
Mikkel Rohde, Mads Daugaard and co-workers identified Hsp70-2 as a cancer-associated survival protein whose depletion in cancer cells causes lysosomal leakage, senescence, and cell death via pathways involving the up-regulation of macrophage inhibitory cytokine-1 and down-regulation of LEDGF, respectively (Rohde et al., Genes Dev 2005; Daugaard et al., Cancer Res 2007). Together with Annika Baude and other co-workers they then showed that LEDGF and its close relative HDGFRP2 promote the repair of DNA double strand breaks by the homologous recombination repair pathway by tethering CtIP to DNA double strand breaks (Daugaard et al., Nature Struct Mol Biol 2012; Baude et al., Nucl Acid Res 2016). In order to keep the focus of the laboratory in lysosomes and autophagy, we terminated this project after successfully defining the DNA repair-associated roles of these proteins - Orphazyme A/S
Prompted by Thomas Kirkegaard's data showing that recombinant heat shock protein 70 (Hsp70) activates lysosomal lipid metabolism and reverts the lysosomal pathology in cells from patients suffering from Niemann Pick disease, a lysosomal storage disorder caused by mutations in the SMPD1 gene encoding for acid sphingomyelinase (Kirkegaard et al., Nature 2010), Thomas Kirkegaard and Marja Jäättelä founded Orphazyme A/S, a biotechnology company devoted to develop innovative Hsp70-based therapies for the treatment of lysosomal storage disorders. The related research showed that recombinant Hsp70 as well as Hsp70-inducing small molecule arimoclomol effectively inhibited glycosphingolipid accumulation and attenuated a wide spectrum of disease-associated neurological symptoms in murine models of lysosomal storage disorders (Kirkegaard et al., Science Transl Med 2016). Encouraged by these data, Orphazyme initiated clinical trials for arimoclomol in Niemann Pick Disease C in 2016
Figure 4. Non-lethal functions of lysosomal membrane permeabilization
In physiological conditions, such as chromosomal segregation during mitosis or deadhesion of cellular protrusions, a subpopulation of lysosomes undergo lysosomal permeabilisation (LMP). This may involve a regulated release of specific lysosomal hydrolases from inside of lysosomes to the cytoplasm to promote processing of their extra-lysosomal targets. The induction and signaling of this subtle LMP is still a mystery. However, these observations have broadened the view in studying lysosomal functions and is currently an area of intense investigation in our laboratory.