The NIH Undiagnosed Diseases Program: Expansion to National and International NetworksGMT20240403-160016_Recording_avo_1280x720

April 04, 2024

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00:11Yes,
00:23I'm just trying to give it back too.
00:24So at the end, I have questions
00:27and there's just about online that
00:29I'll get the questions too. But,
00:36but I'm going to pop down and
00:39I'm going to go through. OK,
00:53good afternoon, everybody.
00:55We'll get started.
00:56Welcome to Grand Rounds in
00:58the Department of Pediatrics.
00:59My name is Cliff Bogue.
01:00I'm the chair of the department and
01:02pleased to have all of you here and
01:05especially our special guest speaker.
01:06So I just have a few announcements
01:08beforehand and then I'm going
01:09to turn it over to Young *****
01:12to introduce today's speaker.
01:14So our upcoming next two grand rounds
01:17next week we have what we call cared
01:20care rounds and this is talking
01:23about collateral damage orphaned by
01:25trauma and we have Matthew Hornick
01:28in peed surgery as long as Jessica
01:31and Heather who are going to be here
01:33to talk about a case presentation.
01:36And then the next week,
01:38April 17th,
01:39we have Kristen Schroeder who's
01:41coming from Duke University to talk
01:44about global oncology development
01:45of a comprehensive pediatric
01:47cancer program in Tanzania.
01:49So that should be really interesting.
01:53Also for those of you who are going
01:54to the pediatric academic societies
01:56meeting in a few weeks,
01:59please come out on Saturday night
02:01to our dessert reception will be
02:03at the Fairmont Royal York at.
02:04You're welcome. Bring your friends,
02:07people that you know alumni.
02:08We'd love to see folks and have a
02:10chance to connect and enjoy one
02:12another at that meeting in Toronto.
02:16Also we have, as you know we're getting near
02:19the end of our strategic planning process
02:22that's been going on for several months.
02:25We're going to have a several one
02:28hour virtual strategic plan feedback
02:31sessions as an opportunity to give some
02:33final feedback and most importantly
02:35think about how do we move forward,
02:37how do we get a broad buy in for some of the
02:40exciting things we have in the in the plan.
02:42So you'll be these
02:44invitations will be going out,
02:46but you can see we have them focused in
02:50various pillars of of our department and
02:53and so please encourage you if you're
02:56available come to those and participate.
02:59They'll be really one of the last
03:02chances for that kind of impact
03:04input before we sort of finalize the
03:06plan and then begin to implement it.
03:09All of our grand rounds are
03:11available for CME.
03:13We don't have any commercial support.
03:16The number to text is here in
03:18good old whiteboard and also
03:19we'll be in the Zoom chat,
03:22but you can get CME credit
03:25for participation today.
03:27And with that,
03:27I'm going to turn it over to Yang
03:29Hui from the Department of Genetics,
03:31but also a close colleague in Pediatrics
03:34to introduce today's speaker.
03:39Thank you, Cliff. Indeed,
03:42I'm the professor for genetics and Pediatrics
03:44and Chief for the clinical genetics service.
03:48Since last year we started in this
03:50called the pediatric genetics,
03:51joined Ron and last year was a great success.
03:54And then we're going to keep doing
03:56that every year and hope maybe we'll
03:58do more and then one time per year
04:02today is the best special speaker.
04:04It's my prefer to kind of
04:07introduce Doctor William GAO.
04:10We should call Beer GAO.
04:11It's much easier.
04:13So for the people like Beer
04:16clearly take a lecture to tell
04:19his career great career for sure.
04:21I can spend our to talking to the
04:25his career or he can tell the story.
04:28Bill is good storyteller.
04:29He said OK I can tell story all the time.
04:32So very quickly some is on the fly
04:34this introduction for that and which
04:37very quickly highlight something.
04:38I will not touch UDMUDP because
04:40he's going to tell the story just
04:42put more not in the description
04:44in the fly a few highlights.
04:46So Bill Current is a senior
04:50investigator and the director for UDP,
04:53the head of a biochemical genetics.
04:55He usually said he given a lot of
04:58title last maybe 10 years and then
05:01he was the clinical director for the
05:04NHGI for almost longer than 15 years.
05:07A long very very long than many
05:09other title section chief.
05:11Also he declined a lot of exciting
05:14offer to he could be the and I and
05:18I see actually director at some
05:19point he said I don't want that one
05:22or department chair some school and
05:24all together but he clearly want to
05:27dedicate his time to working on rare
05:30disease undiagnosed disease that's
05:32what he's the signature program he created.
05:36He's a funding director 2008 and
05:39that led to the UDN Undiagnosed
05:42Disease Network 2017 right
05:472017. Now he remained
05:48active for that program.
05:50He gave up many titles but he keep
05:52this one for long term so quickly
05:56be a graduate from MIT and then it
06:00was probably missing opportunity
06:02for Harvard did not take him to the
06:05Harvard Medical School and he went
06:07to the Wisconsin finished MDPHD and
06:09did a pediatric residency and he was
06:13a chief resident and and also come to
06:16the NIH finishing clinical genetics
06:19and clinical biochemical genetics.
06:22His was funding ACMG American College
06:25of medical genetics fellow he was
06:28just what dinner tonight last night
06:30he was telling a story at that
06:32time doing board exam for medical
06:34genetics like half people write
06:36an exam for other half trainees.
06:38So that's what how how the history goes.
06:41So maybe it's easy then today,
06:43so you don't have to go to
06:45database to pick that one.
06:46He's researched clearly he's
06:49a physician scientist and it's
06:51a role model for me for sure.
06:54And he covered dedicated time to
06:56discover the gene for rare disease
06:59primarily by chemical genetic disorder.
07:02Now he went ahead and did a lot of
07:05basic science study understanding
07:07what's the mechanism for that disease
07:10then continue for developer treatment,
07:12engage FDA.
07:13He developed a multiple treatment
07:15for rare disease and a lot of
07:17the signature program.
07:18If I look at the CV that's a list
07:20of engagement with FDA and make
07:22sure the patient in the clinical
07:25gather that hope to after diagnosis.
07:28And this clearly is a proliferix kind of Pi.
07:32He published 650 paper and a lot
07:37of clinical protocol and a lot of
07:39product treatment FDA approval there.
07:42Then he also discovered the signature
07:45program is he discovered the molecular
07:48basis for the cystinosis or sciatic
07:51acid disorder which is pretty rare for
07:53many of you probably have not heard
07:55about and he probably will tell the
07:57story too and the many other rare
07:59disease most in the metabolic related.
08:02Then his group after study UDP
08:06discovered almost 300 disease gene and
08:10the disorder and the more then as a
08:15mentor I should mention Bill Trader
08:19train 42 clinical biochemical genetic
08:23fellow almost ten 110th of anti people
08:27board certified in my specialty.
08:30He was the first one push for
08:33the new specialty called medical
08:35biochemical genetics,
08:36which I was the beneficiary.
08:39I was the first founding fellow 2009 five
08:44for because he's a vision for that program.
08:48So you can see many of people he
08:51trained including here and many
08:53other institution is a leader in
08:55the biochemical genetics.
08:57As I said,
08:58I would leave the UDPUT not touch that part.
09:00He would tell the story.
09:02Because of that you can clearly see
09:06how much accomplishment and reward
09:08that he has got. It's a long list.
09:10I just pick a few.
09:12I I I actually copy from his CV
09:14make sure I cover some major one.
09:17He received multiple time for
09:19NIH director award many years
09:21throughout his career.
09:23He,
09:23as NICHD Hall of the honor only 15 scientists
09:28was recognized, would recognize
09:322013, including three or four Nobel Laureate.
09:36In that list he was the president of
09:39a Society of Embroiler metabolism.
09:42He received the Nathan Davis Award,
09:44outstanding government service
09:46from AMA Euris Lifetime Chiefman
09:50Award and 2019 he was elected for
09:54National Academy for Medicine.
09:57With that I guess I should
09:59say a few personal notes.
10:01As you know I I'm clinical genetics
10:03also clinical I think I call them
10:05medical biochemical genetics,
10:07not clinical biochemical genetics
10:08that's more lab to direct it.
10:10So I definitely know Bill's all works before
10:13I know him in person so we can know him.
10:16It's when the UDN when I was at Duke.
10:20We are part of a clinical side of the UDN
10:24funding like the first seven side name.
10:26So we met her before the COVID.
10:29We met her regularly quarterly
10:31at DC or some other House Hotel.
10:35Now we're starting to engage with a
10:38Bill and learn he's a he's a great
10:42visionary leader for this field.
10:44One thing I like very much is
10:45every time you notice Consortium,
10:47so many people in a meeting and he
10:49often time opened up the talk at
10:51the beginning for the Consortium
10:52as you know to try to get all these
10:54smart people in the same room to
10:56talking some challenging topics,
10:58a very,
10:59very sometimes very challenging.
11:00So Pierre usually opened up
11:04sometimes kind of light moment joke
11:061st and make everyone laugh first.
11:08So as you can see it when you laugh,
11:11everything can synchronize very well.
11:13After that initial sort of first
11:15few minutes you always can find
11:17a way to make everyone laugh.
11:19And before we get in serious
11:21about our serious topic so that
11:23I was fair bit impressed,
11:25I'm missing out a lot of things
11:27are really good to hear.
11:28I hope we can join you back and
11:31enjoy that part of learn how you
11:33are leading this consortium and and
11:36visionary leader for this very,
11:38very challenging topic for
11:40often time in the medicine.
11:43And also I think 2015 was a Project
11:47B and I have made a trip to the
11:50Shanghai to kind of disseminate all
11:54Nash international outreach for UDA P
11:58program that was great fun to when together.
12:01I really still remembers a lot of people
12:03also almost try to show one picture here.
12:06We will definitely show
12:07next time with that Bill,
12:08thank you so much for coming
12:10from your busy schedule and we're
12:11looking forward to your talk.
12:17Thanks very much.
12:18I'll try to get people to laugh
12:20with me rather than laugh at me.
12:22And also some of the jokes that I tell here,
12:25I can't actually tell in Shanghai,
12:28but where's this all? Here we go.
12:30So and I want to tell stories today,
12:33stories about unusual diseases and mechanisms
12:36of disease that you're not so likely to see,
12:40but that still interest us and may have
12:43applications to common disease because
12:45that's what happens with some rare diseases.
12:48And I want to mention that Cindy Tift is the
12:51Director of the pediatric portion of the
12:54UDP and David Adams does the bioinformatics.
12:56And we have two great neurologists
12:58I mentioned here and two great
13:00internists and a psych coordinator.
13:01And this program is supported by the
13:05volunteer efforts of a huge number of
13:07experts within the intro program of
13:09the NIH experts in rare diseases there.
13:12This program was established in and
13:15announced in May of 2008 with two goals.
13:18One to help people reach a diagnosis
13:20when they've sought A diagnosis
13:22and haven't been able to get one,
13:24and the other is to discover new
13:26things about biochemistry and cell
13:28biology and mechanisms of disease.
13:29So to contribute to medicine.
13:31And the way it works is that the
13:34applicants submit their medical records
13:35and I for the adults and doctor TIFF for
13:38the children will look them over and
13:40triage them to different experts to say,
13:43offer an opinion about whether this
13:46is reasonable or not as something
13:47that we should study.
13:49We only accept about 1/3 or so of the
13:52people who apply and we offer some advice
13:55to the others and the ones that we see,
13:57we see for a week at the NIH free of charge.
14:00So we don't charge any third parties either.
14:03Over the course of the last 15 or 16 years,
14:05we've seen over 6000 medical records
14:07and seen over 1600 at the NIH.
14:10A lot of kids and more than half of
14:12our cases are neurological cases,
14:14a lot of exomes and especially family exomes.
14:17We get a skin biopsy for fibroblast
14:20culture to do some gene function
14:22studies on about 70% of the patients.
14:25We see a lot of diagnosis and
14:28publications as well.
14:29And for the genetics,
14:32we can do customized
14:35personalized phenotyping,
14:36but also some of the genetics
14:39that's available commercially.
14:40We also do snip arrays on many of our
14:44patients and excellence and genomes
14:45as I mentioned and for some of them
14:48when we have multiple different
14:49candidate genes and variants and genes,
14:52we'll do functional studies in
14:54fibroblasts and there's a model organisms
14:56core as well that we can employ.
14:59I wanted to give you some examples
15:01of discovery and this is an early
15:03discovery of ours.
15:04We saw five adults from the Kentucky,
15:08Ohio region who are all siblings and
15:11they had claudication in their lower
15:13extremities because of ischemic pain.
15:15So vascular insufficiency is what they had.
15:18Their coronaries were largely
15:21spirit, but these are their arteries
15:25and this is there's no contrast here.
15:28So this is all calcification of their
15:30femoral and popliteal arteries.
15:31And here it is on PA.
15:34And see the Der Salus petis calcified here.
15:38No wonder they had pain and they were
15:42surviving off their collaterals.
15:44And they also had in the metacarpal
15:46phalangeal joints some calcification.
15:48There you can see that as well.
15:51Well, it turns out that their
15:53parents were third cousins,
15:55and we know that the first
15:56cousins share 1/8 of their genes.
15:58Second cousins share 132nd of their genes.
16:01Third cousins share one 128th of their genes.
16:04So if we're going to consider this a
16:07recessive disorder that may be caused
16:10by the consanguinity that they have,
16:12we can look in one 128th of their genes,
16:16that is to say,
16:17the regions in which they are homozygous.
16:20And the reason that that's important is
16:21because in that region of homozygosity,
16:23if you have one variant,
16:25you're going to have two variants,
16:26and this could have caused
16:28a recessive disease.
16:29Well, it turns out that there
16:31were such regions on a SNP array,
16:33single nucleotide polymorphisms.
16:35That disarray contains a million of those,
16:39meaning that since they're 3.2 billion bases,
16:42these SNPs are about 3000 bases apart.
16:47And every one of these little
16:49blue dots is a SNP.
16:50And so the those blue dots,
16:55the separation between them
16:56represents 3000 bases.
16:58So this is a region of chromosome 6,
17:00and what's shown those dots
17:03are only the heterozygous SNPs.
17:05So we've eliminated the top ones
17:07which are the AAS and the bottom
17:09ones which are the BBS and this,
17:11these are the ABS that you see.
17:13And you can see that for the siblings here,
17:16that's children 12345.
17:18They all have a region with
17:21no heterozygosity.
17:21So they're homozygous in this region,
17:23meaning that this if there's a variant here,
17:26they're going to have it on
17:27both of their alleles.
17:29And that was a region of 22
17:31mega bases with 92 genes.
17:32And our heart,
17:33lung and blood associates picked out
17:36anti 5E as a candidate for causing
17:38this disease and that encodes CD 73,
17:43an enzyme in the vascular endothelium
17:46that converts AMP to adenosine
17:49and inorganic phosphate.
17:51And in fact these five individuals
17:54you see here had all homozygous
17:58nonsense mutation shown here we found
18:01another family with three affected
18:04individuals with a different mutation,
18:06also homozygous and then a compound
18:09heterozygous family as well.
18:11So those individuals in three
18:13different families.
18:14And it turns out that the
18:16fibroblasts expressed this gene.
18:17So here's NT 5E expression in normals.
18:20And then in two of the affected individuals,
18:23the enzyme activity was also
18:24decreased in the fibroblast,
18:26and this enzyme activity could
18:28be rescued by transduction with
18:30a vector that contains CD 73,
18:32the missing enzyme.
18:33So we're trying to prove fusality here,
18:36and that's pretty much how we get it.
18:38Furthermore,
18:38the fibroblasts express not only the
18:41genotype but a phenotype and the
18:45phenotype was increased alkaline
18:47phosphatase activity.
18:49You'll see why this is important,
18:50but this is the affected patients
18:54cultures of fibroblast stained
18:56for alkaline phosphatase.
18:58This is the control and when
19:00you treat with adenosine,
19:01which is the missing product of the CD 73,
19:06you mitigate the alkaline phosphatase
19:09excess and in other words, it rescues it.
19:12It not only rescues that,
19:14but it rescues calcification.
19:17Alizarin red staining is a
19:20reflection of calcium accumulation.
19:23So here's the affected
19:24compared to the normal.
19:25And here is the cell culture of the
19:30affected individuals transduced
19:32with a lentivirus containing CD73.
19:34You see it corrects the calcification.
19:37So does adenosine,
19:39again the product that's
19:40missing and so does levamisol,
19:42which is an inhibitor
19:44of alkaline phosphatase.
19:45So alkaline phosphatase is very important,
19:48has a very important role in this.
19:51And that that role is shown here.
19:53Ordinarily on the vascular endothelium
19:56you have CD 73 converting AMP to
20:00adenosine and then adenosine interacts
20:02with the vascular cell receptors.
20:05To trophically inhibit tissue non
20:08specific alkaline phosphatase.
20:10When that doesn't happen and and you
20:12see that in the patient's fibroblasts
20:14that didn't happen because the patients
20:17had increased alkaline phosphatase,
20:19then the alkaline phosphatase
20:20which is supposed to go to the
20:23surface of the cells and convert
20:26pyrophosphate into inorganic phosphate,
20:28that doesn't happen and instead you
20:31have too much alkaline phosphatase and
20:34the inorganic phosphate is formed and
20:37it enhances mineralization whereas the
20:40pyrophosphate normally inhibits and
20:42that accounts for the calcification
20:44in the vessels of these individuals.
20:48Here's another case,
20:51an 18 month old little girl who had
20:53failure to thrive and some intestinal
20:56problems with TTP and dependent etcetera.
20:58But her sort of claimed the fame for
21:03this particular disorder was she had
21:05hypopigmentation and the poor visual
21:07acuity that associates with it.
21:08Also had organomegaly, liver,
21:10spleen, kidney and storage there,
21:13and also had developmental delay with
21:17poor myelination and some infections too.
21:22But I'll mention this before telling you
21:25why she had no osteopatrosis and we'll
21:29keep that in mind when we see what she had.
21:33So here she is.
21:36She has cutaneous albinism
21:38and also white hair.
21:41She actually has some pigment in her iris,
21:43which is very unusual and
21:45delayed myelination here.
21:46And here's her storage in the liver,
21:49these big storage cells and in the
21:52duodenum and in the PMMS etcetera.
21:54And even in the fibroblast,
21:55she has bacols here.
21:57Bacols are better seen here as well.
22:00And then we found another patient who
22:03had essentially the same phenotype.
22:05So again, hypopigmentation,
22:07large liver, spleen,
22:09kidney and storage,
22:11developmental delay for myelination
22:14and no osteoporosis.
22:17Here's the fella and his dad,
22:19and here's the poor myelination.
22:22And here's the storage.
22:24And the storage and the storage fibroblast.
22:28Well,
22:28it turns out we found a de Novo
22:31mutation in a gene called CLCN 7,
22:35and CLCN 7 has a particular
22:38function in lysosomes.
22:40When the proton pump pumps hydrogen
22:43ions into a lysosome it does so
22:45for a long time and and then the
22:48hydrogen ions accumulate on the
22:49inner membrane of the lysosome
22:51and create an electric chemical
22:54gradient against which the next
22:56proton has a hard time getting in.
23:00So in order to dissipate that gradient
23:04God created CLCN 7 to put a counter
23:08ion chloride into the lysosome.
23:11So you if you don't have CLCN 7 you can't
23:14acidify the lysosome very well at at all.
23:18So chloride,
23:19This CLCN 7 provides the
23:21counter ion for this.
23:23And in fact there's a disease
23:25associated with loss of function
23:27by allelic mutations in CLCN 7.
23:30That is a disease called osteopatrosis.
23:32So in other words,
23:33the bone doesn't get broken down
23:35because the osteoclasts can't create
23:39the lacunae of acidic lysosomes to
23:43use the hydrolases to breakdown the
23:45bone so they have osteopetrosis.
23:48Again, loss of function bileelic.
23:50This was instead a mutation
23:52in a different spot which was
23:55monolelic and de Novo.
23:57So we posited that this
23:58is a gain of function.
24:00And our collaborator Joe Mendel and
24:02NANDS did patch clamp studies of
24:05xenoposol sites to demonstrate the
24:07chloride channel and the chloride
24:09movement across the membrane
24:12of these oocytes.
24:13When he put in the wild type,
24:15he got this much current.
24:16When he put in the mutant,
24:17he got this much current.
24:18And that increase in current was
24:21associated down here with more acid.
24:26In other words,
24:27hyperacidosis of the lysosome.
24:30Used fluorescent markers to demonstrate that.
24:33But you can see the difference
24:34in fluorescence in the
24:35pro band, the other pro band and
24:38the consequent decrease in pH
24:41really only a .2 or .3 units of pH,
24:45but that's it's a log scale.
24:48So that's a lot of more acid in
24:51these lysosomes of these individuals.
24:54So we're saying that this is a new
24:56disease associated with hyper acidity
24:58of the lysosomes and you can see why
25:01that would cause storage because the
25:03lysosomal hydrolases not only need acid,
25:06they needed the the right pH.
25:09Isn't a an optimal pH occur for these things,
25:14So it can't be too acid.
25:15It can't be not enough.
25:16Not only that,
25:17but this was a dominant disorder and we
25:21proved that by transfecting the mutant
25:25CLC on 7 gene into normal fibroblasts.
25:30So remember these normal fibroblasts
25:31is a normal contingent of CLC on 7.
25:33Now they've also got the the mutant
25:35and the mutant causes the accumulation
25:37of vesicles that you see here.
25:39So dominant disorder and then our
25:43people Rallu and May created a mouse
25:47knock in of the mouse paralogue of
25:51it's called CLCN 7 and those mice are
25:55a little bit hypo pigmented and they
25:58have the back rules and they have
26:00the storage in their liver etcetera.
26:02Well,
26:03one interesting issue is that you
26:05can actually alkalinize lysosomes.
26:07We knew this from the early studies
26:09of new fell etcetera and the way you
26:11can do it is by giving chloroquine.
26:13So we fed these folks fed chloroquine
26:18and lysotrac or red is an indication
26:21of lysosomal acidity.
26:23So all this red here and then you
26:26add more higher concentrations of
26:27chloroquine and the red goes away,
26:30meaning that you're now offering
26:32some alkalinization to the lysosomes
26:34and you can see that the pH actually
26:36goes up with increased chloroquine.
26:38Well,
26:38one the the physician for the patient
26:41from Ghana was Doctor Deborah de
26:44Salvatore in New Brunswick and
26:46she wrote a protocol to treat her
26:49patient with chloroquine.
26:50And when she did that,
26:52his kidney size decreased.
26:53He had more energy.
26:55He rolled over for the
26:57first time in his life.
26:58Both of our patients died their disease,
27:01but a number of other patients
27:03have appeared and we're trying to
27:07establish a protocol to treat them
27:09with the legal specific Aligos
27:10because this is a gain of function
27:12that maybe you could knock down.
27:17Another case of a couple of
27:18brothers who we saw early and
27:21they had lost some milestones,
27:22became a toxic and myoclonic, had seizures.
27:26One of them died and they had an
27:28MRI that showed a small cerebellum
27:30and we didn't know what they had.
27:33So here's the small cerebellum
27:34which you can see in the pro band
27:36compared to the mom and the dad.
27:37So we did an exome sequencing on
27:40the family and it turns out there
27:42are six members of the family.
27:44So we had the parents and we had
27:46two affected and two unaffected.
27:48And when you compare 1 exome
27:50with another exome,
27:51there'll usually be about 20,000 variants.
27:55So among all these family of six,
27:56there are 120,000 variants that were
28:00different and so we had to filter
28:02that down and we finally filtered it
28:04down to considering that this was
28:06going to be a a recessive disease
28:08because the parents were first cousins
28:09and shared 1/8 of their genes.
28:11So we're looking for a recessive
28:13disorder with the homozygous variant
28:15that we would call a mutation
28:18eventually found one in AFG 3L2.
28:20So homozygous for that and AFGL 3L2.
28:25AFG 3L2 is a very interesting
28:28mitochondrial protease that is important
28:31for the formation of axons in nerves,
28:35and this protein does two things.
28:39It forms a heterodimer with a
28:42protein called paraplegian and then
28:44it forms a homodimer with itself.
28:47And there were already diseases
28:49associated with AFG 3O2 and paraplegia.
28:53The paraplegian gene was a recessive
28:57hereditary ******* paraplegia
28:59and the AFG 3L2
29:03disorder was SCA 28 was a
29:07dominant spinal cerebellar ataxia,
29:09but this was the first occasion in which
29:12there were bioelic mutations in AFG 3L2.
29:14In other words, complete loss of function.
29:16So this protein could no
29:19longer react with itself.
29:21There were no good copies to form the
29:25homodimer and it can no longer react with
29:28paraplegion and form the heterodimer.
29:29So these patients,
29:31these boys had both diseases,
29:33they had both SBG 7 and SCA 28
29:36along with myoclinic epilepsy only
29:39patients in the world with that.
29:42So over the course of our work in the
29:46undiagnosed these program which is
29:49within the NIH intramural program,
29:51we have discovered 30 new disease
29:54gene associations and some of
29:56them are listed here.
29:57So the phenotype on the left and then
30:00the gene associated with it on the right.
30:02And obviously in order to prove
30:04that you have this association,
30:07you need to publish it,
30:08which means you need at least
30:09two cases to demonstrate it.
30:11And here are 15 other ones.
30:16Now I want to show you a phenotypic
30:18expansion because sometimes we discover
30:20new diseases and sometimes we discover a
30:23different expression of a known disease,
30:25and that's what we're going to show you here.
30:27This is a tube B4B tubulopathy.
30:31So you know, the microtubules transport
30:35things like small vesicles from near the
30:39nucleus to the plasma membrane, etcetera.
30:41So here's a little girl who's got some
30:43eye findings and some dysmorphisms
30:46and she's got hypophosphatemic
30:48rickets along with nephrocalcinosis.
30:51In fact, we documented renal tubular
30:54Fanconi syndrome in here along with
30:56hearing loss and her hypotonia and
30:59we found a de Novo heterozygous
31:01mutation in tube B4B and we eliminated
31:05all other causes of that we knew of
31:09renal tubular Franconi syndrome.
31:10Incidentally, a lot of this work
31:12was done by Jason McFadden here,
31:14who is matriculating into Yale
31:16Medical School next year.
31:17So we're all proud of him.
31:20And so the known diseases of tube B4B,
31:23the at least two phenotypes associated,
31:25one was up here with this mutation,
31:27the C1171 and this the 1172 etcetera
31:31and they all had the auditory
31:33dysfunction and some eye findings
31:36etcetera along with various other stuff.
31:39But none of the patients associated with
31:42these mutations had renal tubular Franconi
31:45syndrome or hypophosphatemic rickets.
31:47Ours was a different mutation.
31:51In order to understand how
31:53this might be occurring,
31:55we got help and collaboration from
31:58an expert in this in child health,
32:02and he taught us about microtubules.
32:06They start out as dimers of alpha
32:08and beta tubulin,
32:09and then they form lines,
32:11and then the lines line up and form
32:14essentially circles,
32:15which are really cylinders.
32:18And there's an edge that is growing.
32:22I'll show you that here the edge
32:24grows and then it recedes.
32:27And this growth and recession
32:29is critical for movement of
32:34vesicles along these microtubules.
32:36If you don't have this ability
32:38to grow and to recede,
32:39you can't move things and the
32:42microtubules don't form properly.
32:45So it turns out that there's a site on
32:49the tube B4B that is responsible for the
32:54microtubule assembly and disassembly.
32:57And that site is a site that binds
33:01GTP and has a GTPA that hydrolyzes
33:08the GTP to GDP.
33:10And that site is here and the
33:15mutations in previous cases are
33:17here and here having nothing to do
33:20with that Gtpa's activity site.
33:23But our patients mutation is right
33:26nearby and likely affected that Gtpa's
33:29activity and site and therefore impaired
33:33the disassembly of these microtubules.
33:36And there's evidence for that,
33:38biochemical evidence because when
33:40microtubules don't disassemble,
33:42they stay around longer and
33:45therefore they are modified and
33:47they're modified by acetylation.
33:49So when you measure total
33:52turbulent by a western blot,
33:54you see that the control and the probe
33:56band have the same amount roughly,
33:57but when you measure accelerated tubulin,
34:00the probe band has much more
34:02because the stuff is sitting
34:04around not being disassembled.
34:06This is a little bit to me like
34:08collagen being over modified.
34:09When there are variants and it stays,
34:11it has more time to be modified anyway.
34:16This indicates that the disassembly
34:18did not occur properly and leads
34:20us to the hypothesis for the
34:22renal tubular Franconi syndrome,
34:24namely that these tubules in renal
34:29tubular cells in proximal tubular
34:31cells move vesicles which contain
34:36phosphate transporters like SLC 34A3.
34:41And those transporters in the
34:42vesicles need to be moved to the
34:45plasma membrane of the tubules
34:46in order for them to function to
34:49reabsorb phosphate back into the body.
34:51And when that doesn't occur because there's
34:54no disassembly of the microtubules,
34:57the stuff the vesicles don't move,
34:59the transporter is not moved to the
35:01plasma membrane and these individuals cannot,
35:05cannot reabsorb their phosphate.
35:07So we told the family about this.
35:11It had been 10 years.
35:12We'd seen the family, but kept working
35:14on it and they were very pleased.
35:16And that's Jason. OK.
35:18So I would say that first of all,
35:21collaboration is important.
35:22This is possibly a mechanism for
35:25hypophosphatemic rickets and it was a great
35:27learning experience for for everybody.
35:29I'm going to quickly go through some
35:33diagnosis to demonstrate the unusual
35:35nature of the diagnosis that we make.
35:38Here are some that are we would call rare
35:41and I have like several slides of this.
35:43So I'm going through these fast.
35:44You don't have to actually look at any
35:47of these unless you find it of interest.
35:49But you see five in the world,
35:50six families in the world,
35:5120 families in the world,
35:52etcetera.
35:53Really unusual stuff because we get our
35:55patients from major medical centers that
35:57have already worked up the patients,
35:59you know really a lot and more diagnosis.
36:04I generally say that if you know
36:07all or almost all these diagnosis,
36:09you should get a life.
36:14Even geneticists will not know most,
36:16most of these in general and
36:18some the recent diagnosis.
36:20Yeah, I don't want to waste a lot of time
36:22on this but a lot of and then a couple
36:25of cases of personalized treatment.
36:27This is a 12 year old girl who had
36:29these exostoses and you can see it's
36:32not pleasant and you can see it with
36:34the arrow here and turns out she
36:38had familial tumoral calcinosis,
36:40which I know you know about time,
36:42but this is an FGF deficiency
36:45because FGF is a hormone that causes
36:49phosphate to be excreted in the urine.
36:52And if you don't have FGF you you
36:56instead reabsorb the phosphate and you
36:58have a lot of phosphate inside you.
37:00So an FGF deficiency will cause a high
37:04tubular reabsorption of phosphate.
37:05But you see here in the fourth line,
37:07fifth line or something,
37:09it's 96% or so and it it should
37:12be probably less than 80% or so.
37:15So we know that mutations in this
37:18Henacetyl galactosamineal transferase
37:22which puts an anaceto group on to FTF 23,
37:26see here.
37:27These are the anaceto
37:30galactosamineal residues
37:32along with sugars on FTF 23.
37:35These protect FTF 23 from being
37:38broken down and if you don't have
37:40that enzyme to put on then you
37:43don't have the protection and then
37:44the FTF 23 gets broken down and
37:47becomes inactive and essentially
37:49that's this is the mutation just
37:52showing that and showing that there
37:54was a lot of C terminal meaning
37:58broken down FTF 23 that caused this.
38:01So we're able to treat with debulking
38:04and a low phosphate diet etcetera
38:06and also an anti-inflammatory.
38:08So there was treatment associated with this.
38:12A second example of taking a rare disease,
38:15making a diagnosis and being able
38:17to treat is this 14 year old from
38:19Nigeria whom we didn't see because
38:21we couldn't get her over here.
38:22But also we we got her DNA etcetera
38:25and she had a lot of fractures
38:27and rickets and was treated with
38:30surgery and vitamin D here her,
38:33her X-rays and here she is spending
38:37most of her life in casts etcetera.
38:40And so we knew that there was
38:43a differential for the rickets
38:45including vitamin D deficiency,
38:46hypophosphatemia and metabolic acidosis.
38:48We got our labs showed the low phosphorus,
38:52the high up phos, it's a bone,
38:54bone breakdown,
38:55but you also had low serum bicarb
38:58and low serum potassium.
38:59So we got our DNA and we found
39:03the mutation in SLC 4A1 which is
39:08a transporter for chloride and
39:11bicarbonate in the distal renal tubule
39:13and it means that the treatment for
39:16her instead is alkali replacement
39:18and potassium and not more vitamin
39:21D and surgeries all the time.
39:23I think this is the final case.
39:26I'm going to show you a 22 year
39:28old woman with dystonia.
39:30And when she was in grade school,
39:33her teacher, she would grip her hand like,
39:35like a pen,
39:36like like this because she
39:38had dystonia in her fingers.
39:39Teachers, you know, holler at her,
39:41that's not the way to hold a pen, etcetera.
39:42She couldn't help it.
39:44And later she had trouble with her
39:46gait because of dysonia, you know,
39:48muscles clenching like this.
39:49But mainly she had problem with her tongue.
39:51She couldn't eat properly.
39:53She lost weight down to 80 lbs.
39:54She couldn't speak properly, etcetera.
39:57And we found a monolithic mutation in KMT 2B,
40:00which is a histone lysine methyl
40:03transferase and we didn't know
40:05that there was an association with
40:07this disease at the time, but.
40:09Because we were sharing,
40:11we put this on a website that
40:13other people could see.
40:14One of those people was Doctor Manju Korean,
40:18who ran a dystonia clinic in London.
40:20And she called me up one day and said,
40:24you know,
40:25I have 20 patients with KMT 2B mutations,
40:28and five of them we've treated
40:31with deep brain stimulation.
40:33So we work together somewhat.
40:35And then found another patient.
40:37Doctor Soldadas is a neurologist
40:39that saw this 20 year old,
40:41very similar history to the one
40:42that I just showed you.
40:43This 20 year old had clumsiness,
40:45poor Gait, couldn't speak properly
40:47because of dystonia on her tongue.
40:50She couldn't write properly,
40:52treated with Baclofen, etcetera,
40:55etcetera. And here she is. Yes,
41:02the normal cognition, normal adultion,
41:06can't move properly. Lying in bed,
41:09able to signal,
41:13able to learn by signaling,
41:35Doing that with our
41:51I don't know who this Taylor Swift is,
41:53but she apparently likes a song of hers.
41:57In any event, we found that this young
41:59lady also had a de Novo KMT 2B mutation,
42:02therefore recommended treatment
42:04with deep brain stimulator.
42:06And here she is after the
42:10deep brain stimulation.
42:14And I I I guess maybe I don't have
42:16to sort of emphasize how important
42:18small increments and this maybe
42:20not as even a small increment.
42:23But when you have someone who's so
42:25devastated that the activities of
42:26daily living are so difficult for not
42:29only the patient but for the family,
42:31this was transformational for this family.
42:36And why did it occur?
42:38I mean, she can walk steps.
42:43You know, I'm going to go on here.
42:45But it occurred because we shared
42:50something and someone else shared with us.
42:54OK, so a couple other things we do.
42:56We have rounds on Thursday mornings
42:58and that for basically people between
43:01college and medical school or Graduate
43:03School and present cases in person,
43:06journal club and stuff like that.
43:08In 2014, as Yahoo we mentioned,
43:12we expanded to the Undiagnosed
43:14Diseases Network, which is a national
43:18consortium with 7 clinical sites,
43:20a coordinating center, sequencing cores,
43:23a metabolomics core model,
43:26orchism screening center that does largely
43:29Drosophila and Zebrafish and a repository.
43:33I'm Pi of the protocol because
43:35it's the research protocol.
43:36Every patient enrolled as a research patient.
43:39So we do the genetics especially
43:42and expanded to 11 sites,
43:45extramural sites,
43:45the UDP and the interim program
43:48is part of that as well.
43:50And that group saw almost 7000 applications
43:55and almost 2800 evaluated almost 2400
44:00individuals made over 700 diagnosis
44:03etcetera published a lot of papers in 2014.
44:06We also expanded to UDP sites around
44:11the world where the NIHUDP would
44:14served as a model and established the
44:18Undiagnosed Disease Network International,
44:20which has a website,
44:22a charter committees etcetera.
44:24We've had 12 meetings,
44:25Last one was in Tbilisi,
44:27Georgia,
44:28next one is in Seoul,
44:30Korea and have new initiatives
44:34including a diagnostics working
44:36group and a low and middle income
44:38countries working group that has
44:40representatives all around the world.
44:42And that working group is collaborating
44:45with the Wilhelm Foundation.
44:47Wilhelm Foundation is dedicated
44:49to the promotion of undiagnosed
44:54diseases programs throughout the
44:56world because Elaine Cedaroth,
44:58who founded this with her
45:01husband along with the UDP.
45:03In 2014,
45:03she came to my laboratory and wanted
45:06to have this Wilhelm Foundation
45:08founded because she had three children
45:11who all died of an undiagnosed
45:14neurological disease in childhood.
45:17So the Wilhelm Foundation and the UD
45:20and I working groups got together
45:23to establish the Champions program
45:25which I have so far identified
45:28individual physicians in the Congo,
45:32Ghana,
45:32Pakistan and Mali to establish
45:35undiagnosed disease programs there.
45:37And so the UD and I and the Willem
45:40Foundation can provide resources
45:42in terms of collaborations,
45:44access to sequencing, some teaching.
45:47In other words, people coming over to,
45:49for example,
45:50United States and Olaf Beaudemar's lab at
45:54Harvard and some financial support that,
45:57for example,
45:59came to the Wilhelm Foundation from
46:01the Chan Zuckerberg Initiative and
46:03is being funneled them to these
46:05champions in other countries.
46:10Yeah, this describes it.
46:11And this is one of the beautiful young
46:14patients in one of those countries.
46:18So I saw this leave you with the
46:22fact that there are many ways
46:23that we could work together.
46:25And I think you know there are referral
46:28of patients if you're interested,
46:30but you know we do have plenty of patients.
46:33But for example we have 1600 patients
46:36that we've seen and probably about
46:38800 of them have not been solved
46:40and we have genetics on them and
46:43really good phenotyping on them and
46:45we have fiberglass on most of them.
46:47So if you had a favorite gene and maybe
46:50there was a variance in that favorite
46:52gene that was associated with a phenotype,
46:55but it was only one patient and
46:57you didn't know if this variant and
47:00this gene was causal to the disease.
47:03You can tell me what the gene is and
47:04I'll tell you if we have a patient
47:06in our database that has a variance
47:08in that gene and then you can decide
47:09what you want to do with that Also.
47:12I think maybe I've already provided
47:13A protocol and consent and manual
47:15of operations here,
47:16but that's certainly available.
47:18I think our goal and maybe the
47:21goal of all of the physicians is to
47:24lend a helping hand and these are
47:29particularly needy group because
47:33not only isn't there a treatment,
47:34but there isn't a diagnosis and
47:37maybe especially there isn't even
47:39a community for them because they
47:41can't say what they have.
47:43So thank you for your attention.
48:00It's
48:02beautiful talk, beautiful stories.
48:05The story that is the most amazing
48:08is the story of the centre.
48:09In your story I had a question
48:13related to the tubulopathy that
48:17caused the Rick. It's not the Nigerian
48:19one, but the tubular
48:20mutation, and you describe
48:24the Fanconi syndromes.
48:27You couldn't tell whether it was
48:28isolated to phosphorus or other solutes
48:30were part of that phenotype as being.
48:36And I wondered if this might tell
48:40us if that protein has a specific
48:43role in intracellular
48:45trafficking of phosphate transporters
48:47versus other solute transporters.
48:51No, you you, you're you're right.
48:53And I failed to mention that the pancoli
48:56syndrome was, I would say isolated.
48:58In other words, she did not have, you know,
49:01Asturia or small molecular
49:03reporter. Yeah, she said that's why
49:05the hypothesis was that it was really
49:07the SLC or yeah, whatever it was. But I
49:10I think you're you're right that
49:14the microtubules made the important
49:18in other cells of the tubules of the
49:22kidney for transport of transporters,
49:26for movement of transporters to the membrane.
49:28And that has not been investigated in in fact
49:33the scene that I showed you was hypothetical.
49:35So we haven't actually demonstrated that that
49:37transporter didn't get there and that that
49:39was the cause that's this bottom
49:46doctor. Thank you for the
49:48inspirational background.
49:50The question I had was about your
49:52story about phenotype expansion.
49:55And as clinicians and diagnosticians,
50:00we often come across symbol gene
50:03disorder and it is actually able to
50:05talk about phenotype attention whenever
50:08there are any news from stations.
50:10So in this group of patients,
50:13are they two genetic diseases and
50:15some work by Jennifer Posey have
50:18shown that up to 5% may be higher.
50:21Individuals have two diseases coming together
50:25at bending or penotrive.
50:27So my question to you is in the
50:30undiagnosed disease program, are you
50:32recognizing that history? The answer
50:35is yes. We look for it.
50:37I I think that the expansion of
50:39phenotype was a little different
50:40because we could explain
50:42you know the other phenotypes findings,
50:45but we have a number of cases
50:47that we think that there's another
50:49gene involved and it's not just
50:52that it's a modifying gene,
50:54it's another monogenic disease
50:57that we haven't figured out.
50:59But when you have two of them,
51:01it's really and and and
51:02when they're both new,
51:03it's really difficult to distinguish the
51:06which of the phenotypes is associated
51:08with one variant and which might be
51:10associated with another variant.
51:12And I I guess I was talking to
51:13some of the people to hear that
51:17the pursuit of those really difficult
51:20cases takes this much effort and
51:22the pursuit of a new case that
51:25we see takes this much effort to
51:27get solution. Where should we spend our money
51:30and our resources and our time?
51:32So in a way we have to go for those
51:35really unusual cases that do match
51:37it that could be digenic and that
51:43will give us a good reputation and
51:45a big paper and stuff like that.
51:48But in that, in the time that we spend
51:51for that, we could see three new cases and
51:54diagnose two of them or whatever, you know.
51:57So really it's a prioritization issue,
51:59but the answer is yes.
52:01We think that we see number
52:03of diagenic disorders.
52:06I remember when I was working on cystinosis,
52:09we had a young man with cystinosis
52:12and when he ran the first base playing
52:14ball as an adolescent, his fever.
52:18Turns out he also had the braces,
52:20and we actually published that as an example.
52:24But you know,
52:26just because you have water energy
52:28doesn't need not to get it on.
52:50OK, let's just
52:53first she says thank you.
52:58My question is of hundreds
52:59of candidates Snips and then
53:02Glenbar and Polygon,
53:04DAD later than that has been
53:06invariant significance.
53:08How do you think of all
53:09topical pathogenic variants?
53:15Well, you can. You can
53:20see that
53:24one thing is the punitive
53:26inheritance pattern.
53:28So you pretty much can't single
53:32out those paths with the periods.
53:33If you just do a single Excel,
53:35you don't have the first thing.
53:37So if you have the parents,
53:38you can tell if it's inherited and if
53:43the IT segregates with the disease.
53:45So extremely important to have the parents,
53:49the truth, the trios are critical and
53:52then a quartet is helpful as well.
53:55So another sibling that's
53:56either affected or not
53:57affected and all that that's also
53:59beneficial, not as beneficial
54:01as having the parents alone. And then
54:04to have a fifth member of the family
54:06isn't quite so important.
54:08It's the quartet and the trio
54:11says for, But the the point is
54:13that the inheritance pattern
54:15and knowing who is affected by doing
54:18extensive phenotyping will help
54:20you to eliminate a huge number of
54:22the variants that have not
54:25already been eliminated by being
54:27associated with a benign phenotype.
54:31But a lot of times we'll end up
54:34with 510 or 20 variants that are
54:36candidates for causing the disease
54:38and then depending upon how specific
54:41the disease is determines how much we'll
54:44invest in the gene function studies.
54:48You know, because if there's now
54:49specificity in the phenotype,
54:51we're not going to spend
54:52a lot of time looking at
54:54such a very causing dysfunction.
55:03Anything else
55:25and diagnose?
55:29Thanks for popping
55:43up.