Clinical Phenotype and Genetics of mitochondrial disorders

Topic;Clinical Phenotype and Genetics of mitochondrial disorders, Bruce Cohen, March 12, 2022, 09:00 AM Eastern time (US and Canada)

Bruce H. Cohen, MD, FAAN, is the Director of the NeuroDevelopmental Science Center, interim Vice-President and Medical Director of the Rebecca D. Considine Research Institute at The Children’s Hospital Medical Center of Akron, and Professor of Pediatrics as well as Professor of Integrative Medical Sciences at Northeast Ohio Medical University. He holds the Rebecca D. and William Considine Chair in Research at Akron Children's Hospital. Cohen attended Washington University, graduating summa cum laude with a BA in chemistry. He attended the Albert Einstein College of Medicine of Yeshiva University and received his MD degree in 1982. [Read More...] 

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[00:00:00] Bruce Cohen: okay. If everyone's able to see it, I'm going to, I'm going to go ahead and start. It's, hard to even have any discussion about any topic without acknowledging the horror going on in the world today. And and just want to extend our prayers for peace in our prayers. For. Victims of of unwelcome violence in Ukraine and with the moment of silence and, just want to humbly thank ICNA for the invitation to speak this morning, this morning, I'm gonna talking about mitochondrial disease, as it exists in 2022.

[00:00:52] Bruce Cohen: And in doing so I'm going to reflect on the last. About 35 years of my involvement in mitochondrial disease. These are my conflicts of interest. They're mainly research support paid to my medical center for my involvement in performing clinical trials. And as a speaker for the American Academy of Neurology. Our goals and objectives have been distributed, but I'm going to really take you on a tour of mitochondrial disease.

[00:01:25] Bruce Cohen: And I think one of the most important points is topic two, subtopic three of the canonical phenotypes that were described by the founders of mitochondrial disease and these phenotypic descriptions, which evolved between 1950 and 1980 actually play true today. And if we stick with those and we'll list those in a few moments ,even without the best laboratory testing possible.

[00:01:55] Bruce Cohen: You can be rather assure you're getting into the right diagnosis. So my career path I, did my medical trainings starting in 1978 and finished my training in 1989. And really during most of that time I communicated in written expression by a typewriter . When I started my work at the Cleveland clinic, back in 1989, we carried information on a three and a quarter inch disc which held 1.4 megabytes.

[00:02:31] Bruce Cohen: And now we transport information on something even much smaller, a 256 gigabyte in, I would say our knowledge base about mitochondrial diseases analogously as expanding. Although the method what was true and what would still work today worked pretty well back in the early part of mitochondrial, the history of mitochondrial disease.

[00:02:58] Bruce Cohen: What are mitochondria? Mitochondria is obviously sub-cellular organelles. As we understand them on looking at dead cells under a microscope they are bunched up in the cigar shape about one micron in length, but actually in the living cell they tend to form and you really can't measure how big they are because it's one gigantic three-dimensional spider web.

[00:03:26] Bruce Cohen: Throughout the cell that the spiderweb keeps moving tends to accumulate in places where a lot of energy is needed. And you would see that these these spiderwebs division apart and fuse back together, again, they're in constant motion. And that's really one of the beauties of mitochondria is that they're really not these.

[00:03:52] Bruce Cohen: Unique discreet sub-cellular organelles, but it's a network that seems to have a mind of its own. The mitochondria itself is composed of 1200 different proteins. About 90 of these proteins make up the electron transport chain. 13 of these proteins are have their genes located within the mitochondrial DNA.

[00:04:16] Bruce Cohen: The rest are encoded by nuclear DNA. The mitochondrial DNA itself resides within the mitochondria. It's discreet from the, 46 XX and 46 XY package. The mitochondrial DNA has 37 genes. 16,569 base pairs. It's a circular molecule and the genetics of mitochondrial DNA are very different than the mendelian genetics of the nuclear DNA.

[00:04:48] Bruce Cohen: So again, about 1200 of the genes responsible for mitochondrial function and presence are found on the autosomes and display features of mendelian genetics. [00:05:00] The mitochondria are found in every cell except the mature red blood cell and produce 90% of the energy through oxidative phosphorylation.

[00:05:09] Bruce Cohen: These are just pictures. The left the mitochondria are stained with an orange stain and on the right with a red stain and the, staining patterns tend to align with the particular cell where the mitochondria exists. Heart stain on the right is going to look very different than a white blood cell stain of mitochondria on the left.

[00:05:38] Bruce Cohen: Mitochondrial diseases are caused by inheritable or de novo mutations and nuclear DNA and or mitochondrial DNA and these cause primary mitochondrial disease. In fact, that's the definition of primary mitochondrial disease. It's an inheritance or a congenital presence of these mutations at birth. Now, interestingly, spontaneous mutations in the mitochondrial DNA occurs with age and probably contribute to many of the diseases of aging, such as sarcopenia.

[00:06:11] Bruce Cohen: And environmental toxins can lead to mitochondrial dysfunction. We do not consider these as primary mitochondrial disease, but we're fascinated by the presence of disease. And so again, this is a drawing of a mitochondria and the the purpose of the mitochondria is obviously to generate ATP.

[00:06:37] Bruce Cohen: It does so through oxidative phosphorylation, I used to take about 15 minutes of the talk describing oxidative phosphorylation. But we're going to skip through that because it tends to be rather difficult exercise. And I'd rather use my time describing more of the phenotypic and genetic features.

[00:06:58] Bruce Cohen: So it generates ATP. It's also critically responsible, critically responsible for apoptosis. The mitochondria is the judge, jury and executioner of cells. It is the central organelle responsible for apoptosis. Mitochondria are responsible for the generation of free radicals. These free radicals are part of normal, healthy living.

[00:07:23] Bruce Cohen: But when the mitochondria is not working, it tends to generate not only less ATP, but more free radicals. And we're now getting an appreciation that this free radical generation and mitochondrial disease may be a major part of the pathophysiology and some of the disorders. Mitochondria play roles in many different neurodegenerative diseases and some cancers.

[00:07:46] Bruce Cohen: And are the basis of toxicity of many antibacterial drugs and cancer chemotherapy drugs. We're just not going to have time to talk about points four and five. Now this is just a graphical depiction of how much energy a particular tissue can generate in both the resting and non resting state.

[00:08:11] Bruce Cohen: So I'd like to start off on the right with muscles. And you can see muscle at rest uses and generates 9.4 micromole per gram, per minute of ATP. The brain, whether it's resting or thinking uses a little bit more than that. 12.1, the retina 13. The kidney uses 27. So twice it's really twice as much as the retina does.

[00:08:39] Bruce Cohen: And although the kidney can be involved in mitochondrial disease, it most often is not. And it begs the question why do we have so much eye involvement in mitochondrial disease? But not kidney involvement. And the answer to that is really not understood. The heart uses 30 in the heart, never rest, but the heart in a non-exercise state uses 30.

[00:09:06] Bruce Cohen: And the point of the muscle inactivity is that the mitochondria can power up very much like a car engine powers up and in a athlete burn 10 times as much fuel and generate 10 times as much energy. At the exercise mode than at rest. And so that's one of the beauties of the mitochondria.

[00:09:36] Bruce Cohen: It's very much like a car engine and in the working state in a healthy car engine you can accelerate fast and maintain that speed at the expense of burning more energy. So some fun facts about ATP. And I get asked this question all the time by my patients is why don't you just give me ATP if that's what the problem is.

[00:09:57] Bruce Cohen: And there's two reasons. The first [00:10:00] is that in our bodies right now, we have about 24 grams of ATP. That's a fistful of almonds. And I, have a pile of 24 almonds there. That's how much ATP we have in our body, but that gets recycled. It gets hydrolyzed to 80. And wa and phosphate within ourselves as it does its work and then gets re phosphorylated by complex five to form ATP again.

[00:10:28] Bruce Cohen: And the first reason why we can't just take ATP is we would have to take about 72 kilograms of ATP a day. And that's a giant fish sitting on that man's lap. So again, it's an analogy the point is, that recycles each ATP molecule about two or 3000 times a day. Now, the second reason why we can't take ATP as are you put ATP in your mouth, it would explode the, energy of release of even a small amount of ATP.

[00:11:01] Bruce Cohen: It hydrolyzes back to ADP would recreate a fire in your mouth. So I'm not going to go through this history of ATP of mitochondrial disease. It will be supplied for you, but I just want to talk about a few key points. Leigh syndrome was described in 1951, not as a mitochondrial disease, but as a fascinating description of a pathophysiology that affected young infants.

[00:11:31] Bruce Cohen: Followed in 58 by Kearns and Sayre describing their syndrome. And we'll, see examples of this in the talk. The first mitochondrial disease that was labeled the first mitochondrial patient that was labeled as having a mitochondrial disease was by Rolf Luft in 1962. In 1963 soon after that mitochondrial DNA was discovered and in the 1970s, a whole bunch of enzyme assays were developed for which that we spent the next two and a half decades testing patients using these assays to try to diagnose mitochondrial disease.

[00:12:07] Bruce Cohen: The nomenclature was first introduced in 1977 and then formalized in 1985 by Salvatore (Billy) DiMauro. Dr. DiMauro was one of my mentors. I met him in 1984 when I joined the residency program at Columbia Presbyterian. And then there was the, famous syndromic paper describing MERFF, MELAS and Kearns-Sayre syndrome appeared in 1984.

[00:12:33] Bruce Cohen: Also out of Columbia. The first mutations were described in the late 1980s and fast forwarding to 2019. I think the mitochondrial physicians all got together and agree that we should create this description of primary mitochondrial disease specifically. Diseases with a distinct phenotype linked to known mutations that would cause dysfunction of mitochondrial proteins.

[00:13:03] Bruce Cohen: So this is the classic mitochondrial phenotype. I actually made this pretty slide. But didn't develop any of the content of the slide. That was developed by mentors and the early physicians working in mitochondria disease. Interestingly enough, many of these physicians are still involved in mitochondrial diseases today.

[00:13:29] Bruce Cohen: And, these come from descriptions and the literature from the 1980s. And just to talk through them very quickly stepwise deterioration in central nervous system. Ptosis and progressive external ophthalmoplegia, progressive optic atrophy, or retinitis pigmentosa, hearing loss, cardiac conduction defects, myopathy, or cardiomyopathy, and probably smooth muscle myopathy as well.

[00:13:58] Bruce Cohen: Hepatopathy, large fiber neuropathy. Some patients who have this fascinating systemic lipomatosis or lipodystrophy. There's some classic MRI findings, some of the phenotypes and biochemically some, but not all of these patients will have true lactic acidosis, classic amino acid patterns, or classic organic acid patterns.

[00:14:22] Bruce Cohen: Now, obviously not every patient has all of these . Infact most patients don't have the majority of these. But when you, just go through the syndromes that I'll get to in a few moments. You'll see patterns emerge. And I think that if you use this description at the bedside, you'll be able to define what patients are likely to have mitochondrial disease.

[00:14:50] Bruce Cohen: So they got it right back in the 1980s. Let's talk a little bit about genetics for those genetics of the genes [00:15:00] in the nuclear they follow autosomal, recessive, autosomal dominant, and X-Linked patterns. Just like all those diseases we learned about years ago. Mitochondrial DNA follows a pattern that is not X-linked, but is maternal.

[00:15:17] Bruce Cohen: And that's because our mitochondria and mitochondrial DNA are completely inherited from our mothers. There, you see a spermatozoa beginning to enter. Once the head of the spermatozoa gets into the egg any mitochondrial DNA, which actually is located on the mid shaft of the tail of this spermatozoa is instantly de proteinized by enzymes on the inner surface of the ova.

[00:15:47] Bruce Cohen: So we are our mothers' mitochondria and mitochondrial DNA. The four key points about mitochondrial DNA in these pictures. The green circular molecules represent a mitochondrial DNA molecule without mutation and the red circular circles represent mitochondrial DNA that has a specific mutation.

[00:16:17] Bruce Cohen: We're not describing what we're not saying. What mutations it has. But there's a, pathogenic mutation. And so the concept is that a particular mitochondria, a particular cell may have homoplasmy wild mitochondrial DNA or heteroplasmy, and that heteroplasmy can be described as a percentage. You can have 5% mutant, heteroplasmy, 10% mutant, heteroplasmy, 50% mutant heteroplasmy.

[00:16:47] Bruce Cohen: 95% mutant heteroplasmies in some diseases, you get a hundred percent mutant, heteroplasmy. So some mutations are more harmful than others and some mutations cause distinct phenotypic disease. But there is some crossover in, the phenotypes. Again, mutations are not All or None. They can be described as heteroplasmy.

[00:17:12] Bruce Cohen: As I alluded to. The higher percent of mutant heteroplasmy the closer you are to a disease state. And the earlier in life, you get to that disease threshold. And again, there's no good explanation regarding tissue expression activity or why each syndrome is so different than another syndrome. What that means is that we really don't understand why mutations that cause Leber's hereditary, optic neuropathy, or LHON don't cause renal tubular acidosis.

[00:17:46] Bruce Cohen: And we don't understand why. Leber's hereditary optic neuropathy is caused by some particular mutations and very close to that, those same mutation points or other mutations that tend to cause the syndrome of MELAS. We just don't. We just don't get that. There's some explanations coming out, but I've yet to be convinced that we really know why.

[00:18:11] Bruce Cohen: So this is the concept of threshold. Showing that you get to that threshold of disease state, if you have more increased percentage of mutant heteroplasmy, I'm often asked what's the threshold? Is it 77%? If it's 50%, it depends on the individual. It depends on the tissue. It depends on the actual mutation itself.

[00:18:31] Bruce Cohen: So this is maternal inheritance genotype. This is what in the patient. And I'm going to start on the left-hand slide with the female. Women who have a mitochondrial mutation pass that mutation onto all their children. They may not pass the disease on, we'll get to that in the next slide, but they pass the mutation onto all their children, men with a mitochondrial DNA mutation, don't pass it on to any of their children.

[00:18:59] Bruce Cohen: And this gets to the concept that we inherit our mitochondria from her mother and therefore the mitochondrial DNA. Now this slide describes the phenotype. And although a child of a woman with a mitochondrial mutation may have the same disease expression. As you can see in the second generation from the left, the first girl does the first boy does.

[00:19:25] Bruce Cohen: The second girl is subclinical and but may have manifestations later on in life. And the second boy is subclinical, but may have manifestations later on in life. And then if you go down to the next and we're just going to describe the children. Of the left second generation left-hand girl. She has three children.

[00:19:49] Bruce Cohen: The girl has although may have the mutation has no, and will never develop the disease. The first boy gets it young in life and the second boy may have [00:20:00] manifestations later on in life. So in 1992 this slide appeared in the literature. I just like you to look at this circular picture at the top of the page.

[00:20:12] Bruce Cohen: This is a drawing of mitochondrial DNA and the complex one .The seven complex one genes are in purple. The complex III gene is in red. These little tiny slivers that you see of, that are drawn in the background and yellow are the transfer RNA genes and the 10 to 11 o'clock are the two ribosomal RNA genes.

[00:20:40] Bruce Cohen: And that, was that, was all mapped out very nicely in the late sixties, early seventies. And then surrounding the picture you'll see in very tiny print, the mutations that had been described by 1992, and there were about a dozen of them. These mutations include a couple MERFF mutations.

[00:21:02] Bruce Cohen: The NARP mutation associated with Leigh Syndrome. And Kearns-Sayre mutations, some of the PEO mutations and the Leber mutations. So there are about 12 of them. And we knew about these 12 mutations in 1992 and could test for them in research labs. The first ability to test for them in the United States and clinical labs, wasn't till the mid 1990s.

[00:21:25] Bruce Cohen: So I'm going to go through a bunch of phenotypic descriptions and I'm going to start with Leigh syndrome. This girl presented having a perfectly normal three years of life. She had a non-specific viral infection and then woke up one morning, unable to walk because of ataxia, was brought to the hospital.

[00:21:50] Bruce Cohen: And this, took place around the year 2000. When she got sick she was MRI'd. She got an MRI within an hour of getting to the hospital and it showed you could see her midbrain severely affected by this increase, this hyper intensity and was labeled by the emergency room staff, none of which have ever heard of mitochondrial disease or seen mitochondrial disease as having Leigh syndrome.

[00:22:28] Bruce Cohen: And they got it. Just because of the MRI that you do a lactic acid level and performed an LP and found lactic acid increased in both blood and CSF. While she was in the hospital, she developed weakness of one side of her body then weakness of both sides of her body. It was noted that she had eye movement problems and you can see the left exotropia ,swallowing difficulties.

[00:22:53] Bruce Cohen: This has all evolved over about a week. dystonia. She was well enough to be discharged home with a feeding tube and then over the next six months developed a neuropathy, made some recovery over the following over the first six months, but then deteriorated. And this had a stepwise deterioration ever since.

[00:23:18] Bruce Cohen: I decided not to do a muscle biopsy on her because I thought her diagnosis was firm from enough that we could safely call this Leigh syndrome. And the other thing about the clinical manifestations, in Leigh syndrome, it's heavily central nervous system and peripheral nervous system. And not until very later on in the disease presentation, can you get other organ syndrome involvement?

[00:23:47] Bruce Cohen: There's a host of mitochondrial DNA and nuclear DNA mutations that result in this presentation. Why so many genes have a common final pathway of the syndrome? We don't know. Right now there's over 75 genes known to be associated with these. And the age of onset of Leigh's syndrome can be as early as one month to even past 40 years of life.

[00:24:09] Bruce Cohen: But in general, it happens in the first few years of life. Now, once she presented there were over the course of the next five years, I could do a bunch of genetic testing and spent a lot of money trying to find out what the genetic cause of her disease was. Was never able to find it. Although I lost follow-up with her in about 2010, although I did see her grandparents about five years ago, who said she's continuing to get worse.

[00:24:39] Bruce Cohen: I haven't been able to see her because she lives far away and don't know if anyone's done any genetic testing. So that's Leigh syndrome in a nutshell, but I want to tell you about another case on the next slide, of a patient that I saw way before, I met that [00:25:00] girl and it was a 26 year old woman who was previously healthy in a year before she presented to us.

[00:25:06] Bruce Cohen: Had an acute psychosis that resolved with the use of haloperidol. Nine months after that, she presented with psychosis again and Chorea. CAT scan showed calcified basal ganglia. And she was transferred to the adult services at my hospital. And this was her MRI scan showing bilateral basal ganglia injury and she slowly improved over the next several years.

[00:25:43] Bruce Cohen: But let me tell you about her evaluation. It was very interesting. When these patients present, you do the right tests, you can go to the next slide. You really get some pretty rapid answers. Blood lactic acid level was three times a year for limit of normal for CSF lactic acid about four times the upper limit of normal.

[00:26:01] Bruce Cohen: She had elevated alanine to lysine ratio. That's probably the most important marker in amino acids, along with high prolines, , isarc Sarcosines often found as well. She had citric acid cycle intermediates and lactate in her urine. And her muscle pathology was normal, but her muscle enzymology was not normal.

[00:26:24] Bruce Cohen: And at the time in 1997, I could order that panel of 12 mitochondrial DNA mutations. And that was normal. I could do a karyotype again, this is way before microarrays or anything like that. And then that was normal. So the next slide shows her enzymology and I'm not going to spend much time. Except if you drop down to complex one, one in three you'll see the number 6.1 that's , three rows down.

[00:26:57] Bruce Cohen: And a typical value of that enzyme activity is 4 81. So we knew there was something very wrong. Based on enzymology you labeled her as mitochondrial disease. I wasn't able to really discover what happened to her until about 2007, 2008 when we were able to sequence the entire mitochondrial DNA molecule and she was found to have this mutation at the 1644A.

[00:27:27] Bruce Cohen: Nucleoside that encodes for mitochondrial transfer RNA of Leucine gene, and interesting enough had been reported to be associated with adult onset Leigh syndrome. She's waxed and waned throughout the years. I last saw her three or four years ago. She still is. You can have functional speech with her.

[00:27:53] Bruce Cohen: Very pleasant. She does all her normal activities of daily living of keeping her house clean and cooking food for herself and her husband. Talk a little bit about childhood onset, Kearn-Sayre syndrome. This is a patient who presented to her ophthalmologist with the chief complaint of failing his school eye screening.

[00:28:16] Bruce Cohen: And the reason he failed his eye screen tests is when he looked into the machine his head was positioned in such a way that his lids fell over his pupils. So he told the school nurse he couldn't see anything. So she said, you need to go see the eye doctor. So he went in to see one of our ophthalmologists.

[00:28:35] Bruce Cohen: This was when I was working at the Cleveland clinic. And the ophthalmologist called me late on a Friday afternoon saying that he wanted me to see this patient. I reminded my ophthalmologist. It was Friday afternoon. He said, no you'll, appreciate the referral. On examination, his eyes did not move up.

[00:28:56] Bruce Cohen: They didn't move to the left. They didn't move to the right. You can see the ptosis, you can get a general sense that he's myopathic. And then when I listened to his heart, I had to listen long and hard before I heard any heartbeat and send them down to get an EKG. And you can see the EKG, he is in third degree, heart block.

[00:29:19] Bruce Cohen: And he got a pacemaker placed that evening. So this is Kearns-Sayre syndrome. Kearns-Sayre syndrome is the triad that occurs before 20 years of age of high frequency hearing loss, retinitis, pigmentosa, and progressive external Ophthalmoplegia. One get asked what happens if this presents in the thirties and forties, and the answer is what we call a PEO plus.

[00:29:42] Bruce Cohen: But when presenting in the teenage years, we call it Kearns-Sayre syndrome, other features that can occur commonly, but not always include. Remember that laundry list of phenotypes myopathy, diabetes, retinopathy [00:30:00] dementia, seizures, cardiomyopathy, dysphagia, and weight loss. And a very distinct hyper nasal speech.

[00:30:07] Bruce Cohen: Now, if you can look in the upper right-hand corner, that's the circular molecule and mitochondrial DNA. And if I can, in most, if not all patients with Kearns-Sayre syndrome, it's not a point mutation, but it's actually a deletion with the same break points in patient to patient 4,977 base pairs are missing.

[00:30:29] Bruce Cohen: And obviously because of heteroplasmy, a hundred percent mutant heteroplasmy, would be a lethal condition. Most of the patients that were, that I've have measured heteroplasmy, they tend to be 90 to 95% mutant heteroplasmy. So they're surviving off of the 5% wild heteroplasmy.

[00:30:52] Bruce Cohen: Of mitochondrial DNA. And this is generally a progressive disorder described in 1958 by Kearns and Sayre. There's a lot to talk about with Kearns-Sayre syndrome, but I'm just going to keep moving on to the next slide, which is the index patient of my career. This is the picture of a little girl that got me started in mitochondrial disease.

[00:31:20] Bruce Cohen: She was born in 1982. I met her in February of 1983. When I was an intern at the children's hospital, Philadelphia I met her. She was lying in bed. The history I had gotten from the emergency room is that she came in with a hemoglobin of eight. And failure to thrive. That's why she was in the emergency room.

[00:31:44] Bruce Cohen: They diagnosed her as having a urine infection and she was coming in for IV amoxicillin. I went to see her as I walked into the room, she was lying in bed moribund. I remember we, called the code and I paged my mentor, Richard to come see her. We got her out of that and this is not a picture of her muscle biopsy.

[00:32:11] Bruce Cohen: These are images taken off of Google. The top is off Google. The bottom is off another patient I had, but this is what her muscle biopsy looks like even at this age there were some ragged red fibers and she had clumps of mitochondria on the electromicroscopy and the sub sarcolemmal membrane.

[00:32:34] Bruce Cohen: And as she began to grow up, she had moderate intellectual disability, a clear cut myopathy, ptosis and then later on in her life progressive external ophthalmoplegia. Progressive hearing loss, hepatopathy cardiomyopathy. And we really did not have a genetic label on her until 1993. When a research lab found the common Kearns-Sayre deletion in her.

[00:33:04] Bruce Cohen: Now this is Pearson syndrome and most patients with Pearson syndrome don't have the common Kearns-Sayre deletion but have other types of deletions, some larger and some smaller in the mitochondrial DNA. And this is generally caused by non inherited deletions in large portions of the DNA. I run across a Pearson patient probably every three to four years.

[00:33:36] Bruce Cohen: I think we've seen. Two new patients. We've seen two new patients in my medical center since I got there 11 years ago. So it is a pretty rare disorder. She passed away of her cardiomyopathy in 1993. I want to talk a little bit about MELAS, mitochondrial encephalopathy lactic acidosis, and stroke-like syndrome.

[00:33:58] Bruce Cohen: This was described in a paper written by Steve, Peter Phillips, Darryl De Vivo and Billy DiMauro. I think I've got all the authors it was published in 1984, interestingly a screenplay writer picked up on this paper. And use it as the basis of the 1992 academy award winner, film of the year, A Few Good Men.

[00:34:27] Bruce Cohen: And so I'm not going to go into the story of A Few Good Men, but the character that we really never meet in the movie died of what was MELAS. And this is a typical MRI picture of what a stroke would look like in MELAS. It's not so clear to me, whether MELAS would be labeled MELAS if we were to describe it today.

[00:34:51] Bruce Cohen: Because a lot of our patients don't get strokes that have the mutation, the main mutation that caused MELAS, but that's an hour long [00:35:00] discussion as to how we really describe phenotypes in mitochondrial medicine. I'm going to describe two patients with MELAS and, most MELAS occurs due to pathologic mutations and the transfer RNA leucine gene.

[00:35:18] Bruce Cohen: And at the 3243 allele that describes 80%. But not all MELAS patients have mutations in this particular gene. So the first I'm going to give you an a, childhood presentation and adult presentation. I met a girl I actually still take care of her mother and younger brother. But I met her when she was five.

[00:35:40] Bruce Cohen: She had a normal birth weight. Her mother at the time was 33. Mum had short stature type two diabetes, bipolar disease, fibromyalgia, and hearing loss. This little girl had some slight, motor delay, but cognitively was normal. She had failure to thrive and had a g-tube placed when she was three. Developed

[00:36:00] Bruce Cohen: Wolff Parkinson white syndrome when she was three. And interestingly, this is a typical presentation of early MELAS. As we see a lot of Wolf Parkinson white, and she had a successful ablation performed, she went into status epilepticus at age four and an MRI scan that looked like the picture I showed you before.

[00:36:21] Bruce Cohen: Doug Nordli, for those of you who know him who was at the time at Lurie children's hospital, at university of Chicago. Hospitals picked up on this . the, Doug trained two years after I did at Columbia. So we were all really hypervigilant about MELAS found ,did a lactic acid level.

[00:36:42] Bruce Cohen: And then quickly found this mutation the 3243 mutation in this girl, and it fit very well with a pattern of illness that we described as MELAS, explained her mother's subclinical presentation and this little girl unfortunately went on to die of status epilepticus a couple of years after I met her.

[00:37:06] Bruce Cohen: She had a younger brother who also has a mutation. He is now entering college and has migraines. He's had one event that was either a complex migraine or a stroke that did not appear on. MRI testing. There was no restricted diffusion, so it was we, have him under observation.

[00:37:32] Bruce Cohen: The next patient was a 47 year old man. When he presented, he worked as an electrical engineer described as a very nice man. And very smart. His mother died in her early fifties of "Presenile Dementia". Obviously it was MELAS. He then developed a personality change and became this really angry bitter.

[00:37:56] Bruce Cohen: Man, it is a sign of his early side of his dementia, developed hearing loss. He had very high lactic acid level, his MRI didn't show a stroke It showed global atrophy and he too had the 3243 mutation. I'm going to skip because I'm running out of time. I'm going to skip through the presentation of this patient except that it occurred over a five-year period.

[00:38:19] Bruce Cohen: Very quickly. She had developed a movement disorder. Then later on, she developed seizures. Then later on, she was put on Depakote for her seizures and developed liver failure. And this was 1997 that all this occurred in. She had a muscle biopsy because someone's suspected she had a liver transplant and did fine.

[00:38:39] Bruce Cohen: She had a muscle biopsy that was read out as it's consistent with mitochondrial disease. I followed her and followed her and then she came in 2005 with a whole new set of problems. She had developed PEO , dysarthria, ataxia. She actually had retinitis pigmentosa hearing loss, memory difficulties.

[00:39:01] Bruce Cohen: None of these were present before. And then the diagnosis was obvious because and I last saw her on a zoom visit last. But our diagnosis is obvious in 2005. It wouldn't have been able to make it in 2004 because a whole history had about 70 years of work had laid itself out very quickly.

[00:39:26] Bruce Cohen: And since most of you have heard about Alpers-Huttenlocher Syndrome, I'm just going to skip through the details because I'm sure most of you are familiar with it. I, didn't play on my timing. And I'm running out of time. So I'm just going to advance through this , it ends up the gene involved wasn't discovered until 1996.

[00:39:46] Bruce Cohen: And it was basically just another human polymerase. There is 17 human polymerase genes. And this one, gamma and, Bill Copeland described it and discovered it [00:40:00] and he thought it was interesting, but didn't use it, at the NIH. Bob Naviaux, who's an internist, a mitochondrial doc picked up on this gene and said, this must somehow be involved in mitochondrial disease as it ends up five years later, Van Goethem.

[00:40:18] Bruce Cohen: And European collaborators described that mutations in this gene caused Progressive Ophthalmoplegia. And and Bob was able to describe in , 2004 that other mutations in this gene caused Alpers' syndrome. And then what happened mainly in Europe over the next year, over the next year between 2004, when Bob Naviux described.

[00:40:45] Bruce Cohen: Alpers. In 2006, when the first papers were really starting to come out over a hundred different mutations in this gene were described as causing Alpers' syndrome PEO and everything in between. And this is the gene map I pulled off yesterday off the web of all the different allelic mutations in POLG that.

[00:41:14] Bruce Cohen: caused this wide spectrum of disease. I'm going to talk a little bit about the North American Mitochondrial Disease Consortium. It's a group of about 17 different hospitals in North America that are basically trying to curate in, in doing natural history exploration, as well as some other research projects in patients with mitochondrial disease.

[00:41:38] Bruce Cohen: Proud to be a big contributor to patients to this consortium. But we basically were able to find if we just look at phenotypes is that Leigh syndrome accounts for 13% of all our mitochondrial patients in this database, there's a syndrome that's labeled as multi-systemic. And these are patients that are proven to have mitochondrial disease, but don't fit in any particular phenotype.

[00:42:03] Bruce Cohen: Those are called multi-systemic MELAS about 10% CPEOplus about 5%.

[00:42:36] Bruce Cohen: How do we do a lab evaluation for patients? In 2022, I do a history and physical exam. If they seem fit the phenotype, I do blood for lactate, amino acids and carnitine, urine organic acids. And if I really feel that they fit the phenotype, I go to genetic testing. I X'd out what I had written before, which is a microarray and whole exome.

[00:43:03] Bruce Cohen: And now we're going to Whole Genome. This is more of the NAMDC database. And again, now this is the overlapping phenotypes. It's not as pure as I described them.

[00:43:36] Bruce Cohen: There's no FDA approved therapies. The EMA has approved Elamipretide for Barth syndrome and I believe Idebenone for Leber's hereditary optic neuropathy. There's some protocols in development. I'm just going to move to the last slide. So there's no FDA treatments. We really, the treatment is the best take care of the patient and do organ system by organ system involvement.

[00:44:04] Bruce Cohen: One of the most important things we do in the United States is get the kids on proper ventilatory care. Usually it is a matter of BiPAP. So for those of you who have access to whole genome or whole exome, that's really important, but if you don't have access to, this is the most important slide. If you work on this and you stick to the phenotypes, you're going to probably identify 90% of the patients that are out there.

[00:44:32] Bruce Cohen: So I'm going to wrap it up and apologize for running late. And let's go to the questions.

[00:44:42] Samantha Marin: Thank you so much, Dr. Cohen for a fantastic talk. We're going to do the questions in rapid fire if that's okay for you. And I'm just going to go in order and hopefully we have time to answer most of the questions that have been asked. So the first one is given the ATP [00:45:00] production is central to most.

[00:45:01] Samantha Marin: Cellular functioning. Is it possible that we've missed phenotypes by not checking for milder biochemical changes like mild kidney function.

[00:45:12] Bruce Cohen: So we're not going to really know the answer to that question until we have millions of people screened for in whole genome. And it's that answer's probably going to come out of the computer, not out of the clinic.

[00:45:28] Bruce Cohen: Yeah, good question though.

[00:45:31] Samantha Marin: Is it possible? And this is a loaded question to change the course of mitochondrial disease. And the question was specifically about Leigh syndrome using CoQ10 and carnitine or Carglumic acid.

[00:45:46] Bruce Cohen: Patrick Chinnery who now is at Cambridge, wrote a Cochrane review in 2012.

[00:45:53] Bruce Cohen: That's been updated several times. And the Cochrane review looked at the entire medical literature. And the conclusion was that there's no systematic tests that have been or trials that have been done that lead us to think that any of these vitamins are providing benefit to the patients tested in the trials.

[00:46:19] Bruce Cohen: It doesn't say that a patient doesn't benefit. A single patient may not benefit from any of these. But there's no systemic evidence of there being a benefit.

[00:46:32] Samantha Marin: Great. So in the second case of Leigh syndrome, the adult onset case, the MRI findings were slightly asymmetric. So do we find asymmetric MRI findings in mitochondrial diseases?

[00:46:45] Bruce Cohen: So in Leigh Syndrome, we talk about the, lesions being symmetric and again, for the most part, they tend to be symmetric, but we have plenty of patients where there, is some mild asymmetry in MELAS you'll have global atrophy as you is a potential finding, but this man may have had several strokes and back in the day that the imaging wasn't as fine as it is today.

[00:47:18] Bruce Cohen: Couldn't see. The evidence of those strokes in the past,

[00:47:25] Samantha Marin: and just keeping on that same topic, there was a question about Leigh syndrome specifically and why that one tends to be more asymmetric than other conditions.

[00:47:36] Bruce Cohen: I don't think Leigh Syndrome is more asymmetric.

[00:47:38] Samantha Marin: My apologies. It was MELAS syndrome

[00:47:41] Bruce Cohen: Oh, MELAS don't know. One of the newer theories, is that what we're seeing as strokes is really the MRI evidence of a unrecognized status epilepticus. So partial status, but subclinical. And we start gently seeing that a couple of patients that come in there, they come in confused.

[00:48:07] Bruce Cohen: We hook them up to an EEG machine. We find that they're in focal status. Yet we don't see any motor manifestations of that status. And their MRIs show what looks like a restricted diffusion in a stroke. But once we get the status under control, the MRI improves and in some cases really back to normal.

[00:48:29] Bruce Cohen: So that would push us into the status. That was not my theory. I didn't come up with it. But I found that to be true in some cases,

[00:48:41] Samantha Marin: How true is it that exercise in the individual can augment the number and size of mitochondria in a given.

[00:48:52] Bruce Cohen: There's a lot of evidence at Mark Tarnopolsky's lab where he's done exercise mice, mouse models, and POLG disease.

[00:49:00] Bruce Cohen: And it's clear cut if you force exercise mouse with POLG. And again, it's a Knock-in model of, POLG. So it's not a human model, but if you exercise them a mouse. Compared to the, unexercised litter mates. They don't develop the same myopathy through the, several month trial. And he's also done human work where he takes both healthy humans ,couch potato humans, 80 year olds that are just uplifted by ADB and 80, and then mitochondrial patients.

[00:49:37] Bruce Cohen: And he puts them on treadmills. And in every single group their mitochondria are more robust biochemically there's more mitochondria biochemically. And in fact that's the model for some of the new drugs that we're testing in the clinic in clinical trials that [00:50:00] mimic what exercise would do to a cell.

[00:50:04] Bruce Cohen: Now, whether these drugs work is yet to be seen and certainly not all mitochondrial patients can get on a treadmill.

[00:50:14] Samantha Marin: Thank you. And just for the sake of time, I'm going to ask some of the more broad questions and I can answer some of the questions in the chat. I think the great question is in a resource deficient setting how would you diagnose mitochondrial disease?

[00:50:33] Samantha Marin: If the MRI and blood and CSF lactate is normal.

[00:50:38] Bruce Cohen: I would say if they fit the mitochondrial phenotype, you're in pretty good shape. I just diagnosed a girl after 20 years who had clear evidence, biochemically of a mitochondrial disease. She has genetic evidence and again, we've been looking for mutations in Coffin-Siris genes now for seven years, but she had, we just found it two to two or three days ago.

[00:51:03] Bruce Cohen: She has a 3000 base deletion that was missed by, good microarray and missed by whole exome and just found by whole genome. And the best that I can tell is that Coffin-Siris syndrome is not a mitochondrial disease. But it certainly looks like it. She had lactic acidosis .She had high frequency hearing loss, developmental delay, seizures.

[00:51:26] Bruce Cohen: She fit the phenotype, but you're not going to get it right all the time. But I would say that in a low resource area if you've got an MRI scan that I that's, almost a high resource area. I, would think that would be, but I think you just, if you just do the best that you can, and I don't think you're going to go wrong using your clinical judgment.

[00:51:52] Samantha Marin: And I'm going to give two more questions and then I think we'll end there leading from that.

[00:51:57] Samantha Marin: Is there any role of doing a muscle biopsy in the era of genetics?

[00:52:04] Bruce Cohen: I was at the peak of my muscle biopsy career I was doing, I think the biggest year was 52 muscle biopsies in that year. I've done no muscle biopsies in the last two years, partially because of COVID. And I do about two muscle biopsies a year now, and sometimes I'm doing the mostly.

[00:52:27] Bruce Cohen: to, back up the genetic findings. So if you have, availability to the whole genome , you're, I think you're going to be pretty, pretty good. So I would say on rare occasion, a muscle biopsy can be helpful.

[00:52:45] Samantha Marin: And I think the last question, which is a fantastic question. Are, there clinical pointers that differentiate mitochondrial DNA variants versus nuclear DNA variants causing mitochondrial disease?

[00:52:59] Bruce Cohen: I don't know if I understand that question.

[00:53:03] Samantha Marin: So I guess in a patient who has a mitochondrial disease from a nuclear DNA mutation or a mitochondrial disease from a mitochondrial DNA mutation, is there clinical differences between those two patients?

[00:53:19] Bruce Cohen: Other than the phenotypic description? I don't think so.

[00:53:27] Bruce Cohen: When I think about my Complex 1 patients with mutations in the mitochondrial DNA genes. And in my Complex 1 patients with the nuclear DNA genes, there's so much overlap. There was a paper by out of CHOP and MRI paper that got into the MRI findings and the difference. And there may have been some differences.

[00:53:50] Bruce Cohen: I don't know, Samantha if you're aware of it. You see a difference? It's really interesting. If you get five or six mitochondrial doctors together, put them in a bar, give them enough beer. There'll be fighting over these very questions as to what the right answers are with within five minutes.

[00:54:07] Samantha Marin: I think that my fast answer and what I've learned in my career, although this is blurring, is that children who present with mitochondrial disease more often have a nuclear DNA variant, whereas adults who present more often have a mitochondrial DNA variant.

[00:54:20] Samantha Marin: So in the pediatric population, I tend to do nuclear DNA first if I have to choose one of the two, however, I have definitely seen patients where I'm surprised and in the last year alone, I had many more mitochondrial DNA variants compared to what I had expected. So I'm not sure that is truly a differentiating factor.

[00:54:40] Bruce Cohen: Yeah. That may be a fact that most of the evaluations get done in kids by the time patients are adults and get to adult. So that may just be a, I would agree. That's an interesting phenomenon, but I think that may just be, who's getting worked up.

[00:54:57] Samantha Marin: Okay. Great. I'd like to thank everybody for the [00:55:00] questions.

[00:55:00] Samantha Marin: I hope I answered some of the questions in the chat. If they couldn't be answered by Dr. Cohen. And we hope that we see you join us next time.

[00:55:09] Bruce Cohen: Thank you. Thanks everybody. Appreciate you sticking around. .

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