We can restore the mitochondrial function in your brain that is stolen by aging and degenerative diseases!
Mitochondria produce the majority of our energy, can initiate cell death, allow us to live in oxygen and likely bring about many of the challenges of aging. We forget how dependent we are on these remarkable organelles that appear to have developed in cells from primitive bacterial invaders that established residence over 2 billion years ago. A notable biologist named Lynn Margulis popularized this concept of “endosymbiosis”, whereby early cells engulfed primitive bacteria that ultimately became modern-day mitochondria.
Consider the following:
- Mitochondria contain their own small, circular DNA that is exclusively inherited from mothers.
- Mitochondria have two membranes, one (the outer membrane) that is high in protein and resembles in composition their host cells, and the other (the inner membrane) that is high in fat (called cardiolipin) and is more like the bacteria they originated from.
- The molecular machinery mitochondria use to make energy is genetically derived from ~85 separate proteins that are assembled into 5 large “complexes”. Of these ~85 proteins, 13 are made inside mitochondria and come from the circular mitochondrial DNA, with the other ~72 being imported from outside the mitochondria and genetically derived from both parents equally.
- Mitochondria have a life of their own; they fuse, divide and replicate independently of their host cells. However, the machinery for mitochondrial replication genetically comes from their host cells, thus from both parents.
- This means that for cells like those in brain, heart and muscle that divide very little if at all, their mitochondria reproduce themselves over the lifetime of their hosts.
- As you go through life, the mitochondria in these tissues are not the same ones you began your life with.
- Each of the things that mitochondria do can go wrong, leading to “mitochondrial diseases”, which tend to be deficits of energy production and are typically found in high-energy tissues like brain, eye, heart and muscle.
- A substantial body of evidence suggests that mitochondria become damaged as we age, and that in diseases of aging that can affect brain, heart and muscle, mitochondrial impairments may be responsible for many if not all of the symptoms.
We now have the tools to “fix” damaged mitochondria. This advance results from increased understanding of how mitochondria replicate their DNA, how they make energy (called oxidative phosphorylation) and how host cells control production of new mitochondria (called mitochondrial biogenesis, or “mitobiogenesis”).
At the center of these processes is a naturally occurring protein called TFAM, for “mitochondria transcription factor A”. TFAM is essential for normal development, and if it is genetically deleted from embryos, they fail to develop and die.
Figure 1. rhTFAM protein, shown in cartoon fashion in the lower left, has two “high mobility groups” (HMG) that can bind tightly to DNA or RNA; a “zip code” (MLS=mitochondrial localization signal) that sends it to the inside of mitochondria; and a “protein transduction domain” (PTD) that lets the entire protein quickly pass through membranes and enter the inside of cells. Once inside mitochondria, it can bind to DNA (upper left) by virtue of its HMG’s (upper right) and can coat DNA (lower right). Images courtesy of Donna Bennett, SlateBranch Images.
TFAM appears to do many things, some of which we understand. Its name derived from control of transcription, the molecular process by which DNA is converted to RNA that in turn is made into proteins. In this case, TFAM inside mitochondria controls the transcription of mitochondrial DNA into mitochondrial RNA, which in turn is used to guide the synthesis of the 13 mitochondrial proteins involved in energy production.
TFAM is also essential for replication of mitochondrial DNA, that uses a RNA guide just like in viruses. In fact, the enzyme responsible for replication of mitochondrial DNA, known as polymerase gamma (POL-g), can be inhibited by drugs that inhibit HIV replication, so-called “reverse transcriptase inhibitors”.
TFAM itself is not an effective mitochondrial drug. Instead, it has been engineered to have two additional components, called domains (need Figure). The first is a small stretch of amino acids that allow TFAM to pass easily through cell membranes. This process is known as “protein transduction” and is used by natural substances like viruses or man-made proteins to allow these large molecules to enter cells rapidly.
The second component of engineered TFAM is a larger stretch of amino acids that function as a “zip code” to send TFAM to the inside space of mitochondria (called the matrix). This is known as a “mitochondrial localization signal”, and the discovery of these zip codes inside cells resulted in the awarding of a Nobel Prize. Once rhTFAM passes into cells and enters mitochondria, the added domains are removed, and natural TFAM remains.
TFAM engineered with the protein transduction domain and the mitochondrial localization signal attached to one end is known as “recombinant human TFAM”, rhTFAM. rhTFAM applied to human cells or injected into animals stimulates energy production and mitochondrial protein levels in brain, heart and muscle mitochondria, stimulates mitobiogenesis (we’re still working to figure out how this happens), and increases exercise capacity of aging animals.
rhTFAM has the potential to treat a myriad of human conditions that derive from impaired mitochondria. We are particularly interested in using rhTFAM to treat degenerative brain diseases and weaknesses of aging. Because damaged mitochondrial DNA may contribute to these conditions, we are also interested in using rhTFAM to bind healthy mitochondrial DNA and transport it into mitochondria as a “genetic rescue” form of treatment.
At this point, over 3 million dollars have already been spent to develop and test rhTFAM in cells and animals. We need funding to enable synthesis of very pure rhTFAM that can be given to humans, for testing of rhTFAM toxicity in animals, and for the initial human clinical studies of rhTFAM. In total, this funding requirement is 1.5-2 million dollars.
rhTFAM could be used to treat early Alzheimer’s disease or ALS
One of the earliest signs of Alzheimer’s disease (AD) is loss of recent memories. In mouse AD models, memory loss is studied by loss of where a platform is located in a water tank. Mice and rats can swim in water but want to get out as soon as they can. If they are offered a platform in a water tank, they will remember where the platform is located and quickly swim to it. The time course of their learning where the platform is located in the “Morris water maze test” is a measure of their recent memory.
Mice with experimental AD induced by expression of human familial AD genes develop impaired recent memory function in the Morris water maze test. We treated such mice with weekly shots of i.v. rhTFAM. Their recent memory function improved compared to mice given buffer shots without any rhTFAM. Because these mice were at an “early” stage of brain AD pathologically, our findings suggest that rhTFAM might be effective in improving memory function in humans with early AD.
Figure2: These are the Morris water maze results of a study done in mice that express three familial AD genes in their brains (3xTg). One group of mice received treatment with buffer alone (“VEH”) and the other group received treatment with rhTFAM (“TFAM”). Both groups of mice received weekly i.v. injections and were trained to remember where the platform was located in the circular water tank. The mice treated with rhTFAM learned faster than the mice treated with buffer (not shown). After the 10-day period of learning where the platform was located, the mice were rested for 2 days and then retested to see how long it took (seconds, s) to remember where the platform was located. These results indicated that the mice treated with rhTFAM took ~1/3 as long to reach the platform compared to those treated with buffer, indicating that their recent memory function was improved. The two groups of mice had the same swimming speeds, so this improvement is not due to the rhTFAM-treated mice swimming faster.
ALS (amyotrophic lateral sclerosis, “Lou Gherig’s disease”) is a degenerative and fatal brain disease that arises from loss of motor neurons in the brainstem and spinal cord. Rats treated with rhTFAM had marked increases in spinal cord mitochondrial DNA amounts, and also increases in spinal cord levels of mitochondrial energy producing proteins. Because we have reported that human ALS spinal cords show markedly reduced mitochondrial function, rhTFAM might be useful to restore function in those with ALS.
Figure 3: This adult rat study measured the effect of rhTFAM treatment on spinal cord levels of mtDNA genes. A single rhTFAM treatment increased spinal cord mtDNA genes >10-fold.
Figure 4: This study, done in adult rats, showed that weekly treatment with rhTFAM increased spinal cord levels of energy-producing oxphos proteins. This finding is important because human TFAM is only 61% identical to rat TFAM. The likely effect in humans would be much greater.