Sunday, February 2, 2014

Mitochondrial Movements in Cancer

Research projects evolve in a fortuitous manner, often guided by a convergence of novel observations, intuition, helpful colleagues and unique personal circumstances. It is precisely this constellation that prompted two cardiologists to study the mitochondrial networks in lung cancer cells.

In 2008, my colleague and friend Stephen Archer, a Professor of Medicine at the University of Chicago, asked me whether I would be interested in studying the role of mitochondrial networks in lung cancer cells. My first response was the question “Do mitochondria really form networks?”, because at that time the expression “mitochondria” evoked images of scattered oval-like organelles, a textbook image of electron microscopy.
I was also intrigued by my colleague’s request, since we were both cardiologists and it therefore appeared to be somewhat unusual for us to study cancer cells. However, as is often the case in science research, personal motivations lay behind Stephen’s newfound research interest – Stephen’s cousin had recently died from lung cancer.
His cousin’s untimely death and Stephen’s frustration at the lack of therapeutic options for lung cancer victims had served as an incentive for him to expand his ongoing work on the role of mitochondria in cardiovascular cells to also include the investigation of mitochondria in cancer cells. Could we contribute to the identification of novel approaches to treat lung cancer?
During the preceding years, Stephen had focused on the role of glucose oxidation in cancer, in part inspired by the work of the German Nobel prize laureate Otto Heinrich Warburg (1883-1970). In the 1920s, Warburg hypothesized that cancer cells primarily rely on non-oxidative glycolysis instead of glucose oxidation to fuel their energy demands.This metabolic signature of cancer cells was critical for the development and growth of tumors.
As he examined the metabolism of malignant lung cancer cells and non-malignant healthy epithelial cells, Stephen had noticed an important difference in the physical appearance of the mitochondria. The mitochondria in the vast majority of cancer cells appeared to be small and fragmented, while healthy epithelial cells predominantly contained elongated, filamentous-like mitochondria that formed large intact networks. The cause and significance of this difference in the mitochondrial structure between lung cancer cells and healthy lung epithelial cells was unknown and thus a fertile ground for new discoveries.
Even though the planned collaborative project would primarily focus on the mitochondrial network structure and not the mitochondrial metabolism of cancer cells, I also decided to read some of the original Warburg papers in the original German language. German used to be a major language of scientific communication and publication in the 19th century as well as the first half of the 20th century. However, during the latter half of the 20th century and especially in the 21st century, English has become the predominant language of the scientific enterprise, even in Germany.
My nostalgic longing for reading scientific articles in German and my curiosity about how scientists wrote articles in the 1920s prompted me to download some of the Warburg papers. I have to admit that I was quite impressed by the comprehensive nature of the work described. The paper entitled “Über den Stoffwechsel der Carcinomzelle” (Biochemische Zeitschrift 152, 309-344 (1924)) by Warburg and his co-authors Karl Posener and Erwin Negelein contains a comprehensive evaluation of the respiration of tissues from multiple organs, such as the epithelium, connective tissue, brain tissue, retina and various benign and malignant tumors. This 36 page paper includes numerous hypotheses, observations and conclusions about the nature of tumor metabolism that would inspire subsequent generations of scientists.
One observation made by Warburg, for example, toward the end of the manuscript is that tumors with high levels of glycolysis are also associated with high levels of ammonia production and Warburg refers to this observation as an oddity that needs further research. It would take at least 80 years for researchers to understand some of the key underlying molecular mechanisms that explain this observation, when multiple research groups demonstrated that cancer cells use the amino acid glutamine as a major mitochondrial substrate and which upon degradation releases ammonia.
After reading the awe-inspiring Warburg papers, I felt even more enthusiastic about embarking on this new collaboration to study mitochondrial networks in cancer cells.
Peter Toth, a pharmacologist and neuroscientist who directed the confocal microscopy imaging core in Stephen’s group, used his extraordinary live-cell imaging expertise to visualize the mitochondrial networks of malignant and non-malignant lung cells over time. The microscopy data showed that the mitochondrial networks of cells were highly dynamic, continuously undergoing mitochondrial fission (fragmentation or division) and mitochondrial fusion (rejoining). However, at any given time, the majority of cancer cells had smaller, fragmented mitochondria when compared to healthy lung epithelial or vascular cells.
Working with a number of colleagues in our laboratories, we determined that lung cancer cells expressed higher levels of the mitochondrial fission protein Drp-1 when compared to multiple healthy cell types found in the lung. Inhibition of Drp-1 reversed the mitochondrial fragmentation and restored the degree of mitochondrial networking in malignant cancer cells to the levels we observed in healthy non-malignant cells. Similarly, over- expression of the mitochondrial fusion mediator Mitofusin-2 (Mfn-2) also increased mitochondrial networking. Importantly, inhibiting mitochondrial fission resulted in a cell cycle arrest of cancer cells and markedly reduced cancer cell proliferation. In vivo experiments using a tumor xenotransplant model showed a marked reduction in tumor progression when tumors were either treated with a pharmacological inhibitor of Drp-1 or when Mfn-2 was over-expressed.
Our findings are consistent with the observation that mitochondria undergo a cycle of fission and fusion which is coordinated with the cycle of cell division (mitosis). Our experiments suggest that targeting the mitotic fission of mitochondria may be a complementary approach to halt cancer cell proliferation. When we examined the tumor tissues of lung cancer patients, we found that tumor regions indeed expressed markedly higher levels of Drp-1 than healthy lung tissues. Whether Drp-1 levels are also higher in other forms of cancer and whether targeting mitochondrial fission in these other tumor tissues would be equally beneficial still needs to be examined in future studies.
Prior studies have detailed the role of Drp-1 in non-malignant cells where the protein appears to play a role in cell death. Drp-1 induced mitochondrial fission is a characteristic of mitochondrial apoptosis and short-term inhibition of Drp-1 can actually prevent cell death. However, other studies have also linked Drp-1 activation and mitochondrial fission to cell proliferation to mitochondrial fission because the cell cycle regulator Cdk1/Cyclin B regulates the activity of the mitochondrial fission mediator Drp-1. This suggests that mitochondrial fission induced by Drp-1 has two very distinct and nearly contradictory roles: cell death and cell growth.
In the highly proliferative cancer cells that we studied, Drp-1 appeared to be primarily acting as a mediator of mitotic fission, but it is quite possible that in other cell types or settings, Drp-1 may be more closely tied to regulation of apoptotic fission. The fact that the same protein regulates seemingly opposite processes of apoptotic fission during cell death and mitotic fission during cell proliferation may seem surprising. However, it is also reminiscent of the fact that mitochondria themselves can have apparently contradictory roles in cells, acting both as metabolic powerhouses as well as initiators of cell death.
From an evolutionary and teleological standpoint, a cell undergoing division (mitosis) would want to coordinate this process with the dividing and distributing of its mitochondrial organelles. Intact mitochondrial networks are probably difficult to distribute to daughter cells, whereas smaller, fragmented (“fissioned”) mitochondria can be easily distributed. Our study suggests that a reverse signal also exists, by which halting the mitochondrial fission seems to also halt the progression of the cell cycle.
As with any research, our study also points towards many unanswered questions, some of them highlighting the importance of how the nucleus communicates with other organelles: what are the specific mechanisms by which preventing mitotic mitochondrial fission signals back to the nucleus and halts the progression of the cell cycle? How does the cell coordinate the dynamics of other organelles during cell cycle? Could the dynamics of other organelles also be therapeutically targeted in cancer cells?
This work is only the beginning of a journey into the still mysterious realm of organelle movements in cancer cells. It may lead to new treatments for cancer, but such treatments would still require an extensive amount of additional work in pre-clinical studies before they could be tested in humans. The research may also provide fundamental insights of how a cell's nucleus, its "command center", interacts with the other organelles in the cell, some of which hold the keys to the survival and well-being of their "command center". The endosymbiont theory of cellular evolution states that organelles such as mitochondria are derived from lone bacteria (prokaryotes) which were engulfed by complex cells (eukaryotes). Too often, this relationship is portrayed in a one-sided manner: The endosymbiotic mitochondria are like slaves of the cell, continuously providing valuable metabolites for the growth of the cell.  However, it is also known that mitochondria can initiate cell death and that mitochondrial welfare is essential for the survival of the whole cell. The newly discovered role of mitochondrial dynamics in cancer cells, that merely preventing mitochondria from breaking up can bring the whole process of cell growth to a halt, underscores the importance of continuously re-evaluating the relationships and communication routes between the different organelles and the nucleus within a cell.
Notes: An earlier version of this article above was first published on the Scientific American blog. The scientific work described above was published in the FASEB Journal in an article entitled “Inhibition of mitochondrial fission prevents cell cycle progression in lung cancer”. Jalees Rehman, Hannah J. Zhang, Peter T. Toth, Yanmin Zhang, Glenn Marsboom, Zhigang Hong, Ravi Salgia, Aliya N. Husain, Christian Wietholt, & Stephen L. Archer (2012). Inhibition of mitochondrial fission prevents cell cycle progression in lung cancer FASEB Journal DOI: 10.1096/fj.11-196543

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