Propionic Acidemia Foundation Research Grant – Richard

PAF Awards $33,082.12  Research Grant in 2019

PAF Awards $30,591  Continuation Grant in 2020

Eva Richard, PhD, Universidad Autonoma de Madrid, Spain

“Cardiomyocytes derived from induced pluripotent stem cells as a new model for therapy development in propionic acidemia”

Understanding the cellular and molecular mechanisms that occur in genetic diseases is essential for the investigation of new strategies for their prevention and treatment. In this context, induced pluripotent stem cells (iPSC) offer unprecedented opportunities for modeling human disease. One of the fundamental powers of iPSC technology lies in the competency of these cells to be directed to become any cell type in the body, thus allowing researchers to examine disease mechanisms and identify and test novel therapeutics in relevant cell types.

The main objective of this project is focused on the generation of human iPSC-derived cardiomyocytes (hiPSC-CMs) from propionic acidemia (PA) patients as a new human cellular model for the disease.In PA, cardiac symptoms, namely cardiac dysfunction and arrhythmias, have been recognized as progressive late-onset complications resulting in one of the major causes of disease mortality. Using hiPSC-CMs we will study cellular processes, such as mitochondrial function and oxidative stress which have been recognized as main contributors for PA pathophysiology. In addition, our aim is to unravel novel altered pathways using high-throughput techniques such as RNAseq and miRNA analysis. We will also examine the potential beneficial effects of an antioxidant and a mitochondrial biogenesis activator in PA cardiomyocytes. The results that derive from this project will be relevant for the disease providing insight into the affected biological processes, and thus providing tools and models for the identification of novel adjuvant treatments for PA.

Update April 2020 – Eva Richard PhD

Thanks to propionic acidemia (PA) foundation, we have developed a new cellular model of PA based on induced pluripotent stem cells (iPSC) with the goal of defining new PA pathomechanisms which could be potential therapeutical targets. Traditionally, disease pathophysiology has been studied in immortalized or human cell lines and in animal models. Unfortunately, immortalized cells often do not respond as primary cells and animal models do not exactly recapitulate patients‘ clinical symptoms. So far, patients-derived fibroblasts have been mainly used as cellular models in PA due to their availability and robustness, but they have important limitations. The ability to reprogram somatic cells to iPSCs has revolutionized the way of modeling human disease. To study rare diseases,
stem cell models carrying patient-specific mutations have become highly important as all cell types can be differentiated from iPSCs.

We have generated and characterized two iPSC lines from patients-derived fibroblasts with defects in the PCCA and PCCB genes; and an isogenic control in which the mutation of the PCCB patient was genetically corrected using CRISPR/Cas9 technology. These iPSC lines have been successfully differentiated into cardiomyocytes,
and their presence was easily established by visual observation of spontaneously contracting regions and by the expression of several cardiac markers. PCCA iPSC-derived cardiomyocytes exhibited reduced oxygen consumption, an accumulation of residual bodies and lipid droplets, and increased ribosomal biogenesis. Furthermore, we found increased protein levels of HERP, GRP78, GRP75, SIG-1R and MFN2 suggesting
endoplasmic reticulum stress and calcium perturbations in these cells. We also analysed a series of heart-enriched miRNAs previously found deregulated in heart tissue of a PA murine model and confirmed their altered expression.

The present study represents the first report of the characterization of cardiomyocytes derived from iPSCs generated by PA patients ́ fibroblasts reprogramming. Our results provide evidence that several pathomechanisms may have a relevant role in cardiac dysfunction, a common complication in PA disease. This new cellular PA model offers a powerful tool to unravel disease mechanism and, potentially, to enable drug
screening/drug testing. Despite improved therapy over the past few decades, the outcome of PA patients is still unsatisfactory, highlighting the requirement to evaluate new therapies aimed at preventing or alleviating the clinical symptoms. Additional research is required to determine the contribution of the mechanisms identified in this work to the cardiac phenotype and how this knowledge can help formulating better personalized therapeutic
strategies in the future.

We sincerely thank the Propionic Acidemia Foundation for supporting our investigation, which has resulted in a truly motivating experience for us, feeling we belong to the PA research family. The funding we received has led to important advances in PA pathophysiology, and our aim is to continue this research in the near future.

Update September 2019 – Eva Richard PhD

There is an unmet clinical need to develop effective therapies for propionic acidemia (PA). Advances in supportive treatment based on dietary restriction and carnitine supplementation have allowed patients to live beyond the neonatal period. However, the overall outcome remains poor in most patients, who suffer from numerous complications related to disease progression, among them cardiac alterations, a major cause of PA morbidity and mortality. In our research, we developed a new cellular model of PA based on induced pluripotent stem cells (iPSC) with the goal of defining new molecular pathways involved in the pathophysiology of PA which would be potential treatment targeting.

Traditionally, disease pathophysiology has been studied in immortalized or human cell lines and in animal models. Unfortunately, immortalizedcells often do not respond as primary cells and animal models do not exactly recapitulate patients‘ symptoms. So far, patients-derived fibroblasts have been mainly usedas cellular models in PAdue to theiravailability and robustness, but they have important limitations.

The ability to reprogram somatic cells to iPSCs has revolutionized the way of modeling human disease. To study rare diseases, stem cell models carrying patient-specific mutations have become highly important as all cell types can be differentiated from iPSCs. We have generated and characterized two iPSC lines from patients-derived fibroblasts with defects in PCCA and PCCB genes. These iPSC lines can be differentiated into cardiomyocytes that mimic the tissue-specific hallmarks of the disease. The presence of PA cardiomyocytes has been easily established by visual observation of spontaneously contracting regions, and the expression of several cardiac markers. We have observed that PCCA-deficient cardiomyocytes present an increase in degradation products and in lipid droplets, and exhibit mitochondrial dysfunction compared to control cells. We further discovered the down-regulation of several miRNAs in PCCA cardiomyocytes compared to control ones, and several miRNAs targets are currently being analyzed in order to investigate underlying cellular pathological mechanisms. Interestingly, we have performed several experiments to analyze the effect of the mitochondrial biogenesis activator, MIN-102 compound (PPAR agonist, derivative of pioglitazone) in cardiomyocytes.

Preliminary results showed an increase in the oxygen consumption rateof PCCA and control cells. In our next steps, we plan to complete the analysis in the PCCA cardiomyocyte line, characterize PCCB cardiomyocytes and to study in depth the therapeutic potential of MitoQ and MIN-102 compounds.

We would like to sincerely thank the Propionic Acidemia Foundation for supporting our research.

Update March 2020

 “Cardiomyocytes derived from induced pluripotent stem cells as a new model for therapy development in propionic acidemia.”

Eva Richard, Associate Professor

There is an unmet clinical need to develop effective therapies for propionic acidemia (PA). Advances in supportive treatment based on dietary restriction and carnitine supplementation have allowed patients to live beyond the neonatal period. However, the overall outcome remains poor in most patients, who suffer from numerous complications related to disease progression, among them cardiac alterations, a major cause of PA morbidity and mortality. In our research, we developed a new cellular model of PA based on induced pluripotent stem cells (iPSC) with the goal of defining new molecular pathways involved in the pathophysiology of PA which could be potential therapeutical targets.

Traditionally, disease pathophysiology has been studied in immortalized or human cell lines and in animal models. Unfortunately, immortalized cells often do not respond as primary cells and animal models do not exactly recapitulate patients‘ symptoms. So far, patients-derived fibroblasts have been mainly used as cellular models in PA due to their availability and robustness, but they have important limitations.

The ability to reprogram somatic cells to iPSCs has revolutionized the way of modeling human disease. To study rare diseases, stem cell models carrying patient-specific mutations have become highly important as all cell types can be differentiated from iPSCs. We have generated and characterized two iPSC lines from patients-derived fibroblasts with defects in the PCCA and PCCB genes. These iPSC lines can be differentiated into cardiomyocytes that mimic the tissue-specific hallmarks of the disease. The presence of cardiomyocytes has been easily established by visual observation of spontaneously contracting regions, and the expression of several cardiac markers. PCCA iPSC-derived cardiomyocytes exhibited an alteration of autophagy process with an accumulation of residual bodies and mitochondrial dysfunction characterized by reduced oxygen consumption and alteration of mitochondrial biogenesis due to a deregulation of PPARGC1A. We also evaluated the expression of heart-enriched miRNAs previously associated with cardiac dysfunction and several miRNAs were found deregulated. Furthermore, we found increased protein levels of Herp, Grp78, Grp75, sigma-1R and Mfn2 suggesting ER stress and calcium perturbations in these cells.

We are planning to analyze PCCB cardiomyocytes to compare the results with PCCA and control data. We are working to obtain mature cardiomyocytes in order to perform electrophysiology studies (K+ currents) using a whole-cell patch clamp method. We are interested in the study of the tissue-specific bioenergetic signature comparing cardiomyocytes derived from control and PA patients´ iPSCs by reverse phase protein microarrays (RPPMA). Future work also includes testing the effect of the mitochondrial biogenesis activator, MIN-102 compound (PPAR agonist, derivative of pioglitazone) and of the mitochondrial targeting antioxidant MitoQ in PA cardiomyocytes.

We would like to sincerely thank the Propionic Acidemia Foundation for supporting our research.

 

 

 

Propionic Acidemia Foundation Research Grant Guofang Zhang

PAF Awards $48,500 Research Grant

Guofang Zhang, PhD, Duke University

“Propionyl-CoA and propionylcarnitine mediate cardiac complications in patients with propionic acidemia”

Energy production is the central cardiac metabolism for continuous mechanical work. An average human adult heart consumes ~ 6 kg ATP/day. ATP storage in the heart is only sufficient to sustain the heart beat for a few seconds. A tightly coupled cardiac energy metabolism from various substrates is critical for sufficient ATP production required by normal heart function.

One molecule of palmitic acid (fatty acid) generates much more ATP than one molecule of glucose does after their complete metabolism.Fatty acids contribute ~70-90% cardiac energy production in normal condition. However, heart still maintains high flexibility of fuel switch in response to various available substrates. Acetyl-CoA is the first convergent metabolite derived from the diverse fuel substrates via different pathways and enters tricarboxylic acid cycle (TCAC) for energy production. Therefore, the level of acetyl-CoA or the ratio of acetyl-CoA/CoA tightly controls the metabolic fluxes from two major fuels, i.e.,glucose and fatty acid, in the heart. Acetyl-CoA or CoA level is also finely tuned by carnitine acetyltransferase (CrAT) that catalyzes the reversible interconversion between short-chain acyl-CoAs and acylcarnitines.Acetylcarnitine level is ~10-100 fold greater than that of acetyl-CoA in heart and is seen as the buffer of acetyl-CoA. CrAT is highly expressed in high energy demanding organs including heart and mediates fatty acid and glucose metabolism possibly by dynamically interconverting acetyl-CoA and acetylcarnitine into each other.The deficiency of CrAT has been shown to change cardiac fuel selection.

Propionic acidemia (PA) is often associated with cardiac complications. However, the pathological mechanism remains unknown. We have demonstrated that high exogenous propionate led to the propionyl-CoA accumulation and cardiac fuel switch from fatty acid to glucose in the perfused normal rat hearts (Am. J Physiol. Endocrinol. Metab.,2018,315:E622-E633). The deficiency of propionyl-CoA carboxylase in PA also induces the accumulation of propionyl-CoA. Next, we will attempt to understand whether and how the elevated propionyl-CoA in the Pcca-/- heart (collaboration with Dr. Michael Barry)could interrupt cardiac energy metabolism by investigating the fuel switch flexibility, CrAT mediated metabolism, and buffer capacity of acetylcarnitine using stable isotope-based metabolic flux analysis (J. Biol. Chem., 2015,290:8121-32). We hope that the outcome of this project will provide meaningful therapeutic recommendation for patients with PA, especially with the cardiac complication.

Donations – Talli

Donate in Loving Memory of Talli Smith

Your gift supports our mission to find improved treatments and a cure for Propionic Acidemia by funding research and providing information and support to families and medical professionals.   The Propionic Acidemia Foundation is a registered 501 (c) 3 non-profit organization.   Contributions to Propionic Acidemia Foundation are tax deductible; however, consult with your tax advisor for your particular circumstances. Your gift makes a big impact.

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Contact [email protected] if you would like to donate Stock to Propionic Acidemia Foundation.

Peter

Peter – updated 10/18/18

Peter

Hello, my name is Peter and I am 24 years old currently living in Rochester, New York. I have PA. During the first 4 weeks of my life, I was considered “fussy”, but nothing out of the ordinary. At 4 weeks, I experienced projectile vomiting, around the time my mom started supplementing breast milk with formula. I was admitted to the hospital and they believed it was due to my pyloric sphincter (the muscle at stomach opening) and they performed surgery. I stopped vomiting and my health improved for a few weeks, but hindsight suggests it is because I was placed on iv’s and was “cleaned out” during the surgical procedure.

At 6 weeks, I began having absence seizures and was re-admitted to the hospital. A diagnosis came two weeks later. The seizures were a result of extremely elevated glycine levels that crossed the blood/ brain barrier to spinal fluid. All they could do was start me on a non-offending diet and wait for the glycine to reduce. I was started on Propimex (my “special juice”), and 4 weeks later (still in the hospital) the seizures stopped and I was released from the hospital.

My early muscle tone was impacted and I did not walk until 18 months. Physically I seemed delayed but other development testing was favorable.  They monitored my physical and mental development over several years.

Since my hospitalization as an infant I have never had a related metabolic “episode”, or any additional seizures or hospital stays.

My diet was supplemented with Propimex formula until I was about 4 years old. I was a vegetarian until 10 years old when I had my first hot dog! My favorite food was and still is pasta. I had bi-annual appointments at the metabolic clinic at the University of Rochester Medical Centerin which a dietician would suggest the amount of protein I should be eating. I took my lunch through my high school years to help control protein amounts.  It was relatively easy to stay within the protein guidelines since I did not eat a lot of meat.

In high school, I began having rapid heart palpitations and sometimes struggled in gym class when we had to run long distances.I was sent to a pediatric cardiologist for a baseline EKG and had a slightly prolonged Q time.The doctors determined that I had metabolic induced cardiomyopathy. This has been noted in other PA patients.

I was treated with a low dose beta blocker and blood pressure medication to help manage blood flow and hopefully minimize tachycardia events.I have been monitored yearly and my Q time is now “high normal” along with a normal eco cardiogram for 2 years now. I have learned that if I exercise on a near empty stomach, I feel fine! I do have an occasional adrenaline induced tachycardia but I have learned to manage it bytaking deep breaths to stop it quickly.

I went away to college and graduated with both a Music Business / Vocal Performance Degree, and then followed with a second degree in Business Administration. I lived in the dorms, ate campus food, and had a great college experience! I really did not have any issue eating dormitory food as there were many vegetarian options available. I tend to self-regulate and really am not a big meat eater. I probably eat meat or fish2 to 3 times a week. By the way, I LOVE sushi.

I am currently employed as a National Sales Representative  for a company which sells HR, payroll and other services to local businesses. Since middle school, I have been involved in musicals, opera productions, and a cappella groups.

With respect to my current medications, I currently take Levo carnitine and it can be a struggle to keep my free carnitine in the normal range.  I have blood tests once per year and the only thing elevated is glycine and propionic acid levels. All other amino acids remain in the normal range. As mentioned I take a beta blocker and an ACE inhibitor.

I have yearly appointments at U of Rochester Med Center- Pediatric Genetics and also a yearly visit with the cardiologist. Over my lifetime U of R has struggled to keep a full time metabolic specialist on staff. My current physician covers both genetics and metabolics and is extremely busy.

I have never been genetically typed and I would love to support future research or disease understanding. My family and I are happy to share details to anyone who is interested.

That is my story, and I know that I am one of the lucky ones. I do hope that my story encourages parents and children learning to live with PA.

Thank you,

Liver Transplantation Part 2

Liver Transplantation

Part 2: Outcomes Following Liver Transplantation in Children with PA and MMA

James Squires, MD, MS

Dr. Squires is a liver disease specialist at UPMC Children’s Hospital of Pittsburgh and an assistant professor of pediatrics at the University of Pittsburgh School of Medicine.

Jodie M. Vento, MGC, LCGC

Jodie Vento is a genetic counselor and manager of the Center for Rare Disease Therapy at UPMC Children’s Hospital of Pittsburgh.

Part 1 of this article, published in the Spring 2018 issue, provided answers to questions that families may have about what to expect from a liver transplant for a child with Propionic Acidemia (PA). Here, in Part 2, the authors summarize and explain the findings of a recent study of outcomes in children with PA and methylmalonic acidemia (MMA) who received liver transplants at UPMC Children’s Hospital of Pittsburgh.

Why did you do this study?

Before we get to why we did this study, please allow us to back up a bit and briefly discuss the history of liver transplantation for PA and MMA, which was first proposed in the early 1990s. Because the enzyme deficiencies that cause PA and MMA exist throughout the body, not just in the liver, liver transplantation was never expected to be a cure for these diseases. The thinking was that by providing enough functional enzyme to minimize, if not eliminate, metabolic crises­––the most severe complications of PA and MMA for affected children, as well as one of the most frightening features of these diseases for families––a liver transplant could enhance stability and improve quality of life for affected children.

In recent years, policies on the allocation of donor livers in the United States have changed to give priority to patients with PA and MMA because of their risk of sudden, life-threatening metabolic crises. As a result, children with these disorders can now be listed for a liver transplant based on their diagnosis alone rather than on disease complications or severity.

A recent study, based on statistical analysis,found that liver transplantation for PA and MMA may increase both the length and quality of patients’ lives and decrease health care costs over a patient’s lifetime. However, because PA and MMA are rare disorders, it has been difficult to gather a strong body of evidence showing how well patients fare after undergoing a liver transplant.

The Pediatric Liver Transplant Program at UPMC Children’s Hospital of Pittsburgh was established in 1981 by world-renowned transplant surgeon Thomas E. Starzl, MD, PhD. Our Director of Pediatric Transplantation, George Mazariegos, MD, FACS, pioneered liver transplantation for children with metabolic diseases in 2004. Since then, UPMC Children’s has performed more than 330 liver transplants for children with metabolic diseases, more than any other transplant center. We’ve also performed more liver transplants in children than any other center in the United States and more living-donor transplants than any other pediatric center in the country. Our one-year survival rate for pediatric liver transplant patients is 98%, exceeding the national average of 95%, according to the Scientific Registry of Transplant Recipients (January 2018 release).

We decided to do this study because, given the breadth and depth of our experience in this field, we thought that we could make a useful contribution to medical knowledge by gathering and evaluating all of the information available to us on outcomes for all of the patients who have undergone a liver transplant for PA or MMA at our institution.

How did you do this study?

We searched our medical records database to identify all patients with a diagnosis of either PA or MMA who received either a liver transplant or a combined liver and kidney transplant between 2006 and 2017.To comply with patient privacy regulations, we first removed any and all information that could personally identify these patients. Then we examined data from their medical records and recorded information such as their age and family history, medical treatment received prior to the liver transplant, laboratory tests performed, and how they fared both immediately after the transplant and in the following months and years.

What did the study find?

We identified a total of nine patients with PA (three patients) or MMA (6 patients) who had undergone a liver or liver and kidney transplant at UPMC Children’s between 2006 and 2017. The age at which patients received their transplant ranged from one year old to 21 years old; the median, or midpoint, was nine years old. Five patients were female and four male. Eight of the nine patients had been diagnosed during their first week of life; one patient was diagnosed at age eight months.

Prior to the transplant, all of the patients had been treated with protein restriction and carnitine supplementation. Several were also receiving medication to reduce ammonia levels in the blood. Eight of the nine patients were being fed by a gastrostomy tube (also known as a “G-tube”). All were experiencing frequent metabolic crises that often required hospitalization. Additional disease-related complications included cardiomyopathy (damaged heart muscle), metabolic stroke, pancreatitis, and low blood cell counts.

Five of the six patients with MMA received combined liver and kidney transplants. One patient with MMA and all three patients with PA underwent liver transplants only. Patients’ median post-transplant length of stay in intensive care was just short of 30 days, while the total transplant-related hospital stay averaged 55 days. Patients were followed after their transplant for a median of 3.5 years (range one year to more than 11 years).

Six of the nine patients developed symptoms of liver rejection; one patient developed symptoms of kidney rejection. Rejection episodes were treated with steroids and higher doses of anti-rejection medication to suppress the immune system. None of the nine patients experienced transplant failure.

Two patients needed treatment for blood clots in the main artery that carries blood to the liver. A third patient needed treatment for a blockage in a vein that transports blood from the liver back to the heart.

Four patients experienced a build-up of bile in the liver that was caused by a blocked bile duct and required treatment with a biliary catheter. At the last follow-up, three of the four patients had been able to discontinue use of the biliary catheter.

Five patients developed viral infections that required treatment. No patients experienced a complication known as post-transplant lymphoproliferative disorder, a dangerous rapid increase in white blood cells that can sometimes occur in people who are taking medication to prevent rejection of a transplanted organ.

No patients have experienced metabolic crises since the transplant. All nine patients showed improved metabolic control––indicated by normal levels of lactic acid in the blood––during the first month after the transplant. Kidney function stabilized or improved in all patients with MMA. At the two-year post-transplant assessment, heart function had improved in a patient with PA and severe cardiomyopathy.

What conclusions can be drawn from the study’s findings?

In this study of nine children with PA or MMA who were followed for an average of 3.5 years, we show 100 percent survival for both patients and their transplanted organs.

For MMA, these findings are similar to those of other recently published reports. For PA, although our population is relatively small (three patients), our finding of 100 percent survival for both patients and transplanted organs stands in contrast to other published reports that found poor survival among patients with PA following a liver transplant.

Still, many patients experienced complications in the period immediately before, during, and after the transplant. The high rate of complications underscores the complexity of these metabolic diseases. The most common complications were those involving the blood vessels, including blood clotting in the main artery of the liver. This complication has been previously reported.

All patients had reduced levels of lactic acid in the blood, indicating improved metabolic control, both shortly after the transplant and at later postoperative follow-up. Complications such as kidney disease (in patients with MMA) and cardiomyopathy (in patients with PA) stabilized and improved after transplantation.

The fact that no patients experienced metabolic crises after transplantation indicates that partial enzyme replacement via a liver transplant enabled a “resetting” of patients’ metabolic fitness.

At UPMC Children’s our approach to nutritional support after a liver transplant has been to gradually ease protein restriction, with the goal of establishing a long-term individualized level of support for each patient. It is unlikely that protein restriction can ever be completely eliminated. However, the results of this study show that––with close monitoring by an experienced interdisciplinary team––protein restriction can safely be relaxed, in an individualized fashion, after a liver transplant.

What do the study results mean for children with PA and their families?

A liver transplant cannot cure PA. It can, however, reduce or eliminate metabolic crises and result in greater stability and better quality of life for children with PA. The decision as to whether a liver transplant is right for your child with PA is one that every family must make for themselves, based on their knowledge of their child and in consultation with a multidisciplinary team of experts who specialize in liver transplantation for metabolic diseases.

This study adds to the increasing body of evidence that liver transplantation can be performed safely and successfully in patients with severe, complex metabolic conditions such as PA and MMA, especially when performed at centers with broad and deep experience in the management of these highly challenging conditions.

Reference: Critelli K, McKiernan P, Vockley J, Mazariegos G, Squires RH, Soltys K, Squires JE. Liver Transplantation for Propionic Acidemia and Methylmalonic Acidemia: Peri-operative Management and Clinical Outcomes. In press, Liver Transplantation. Accepted for publication June 2018.

Novel therapies for Propionic acidemia – update Sept. 2018

Novel therapies for Propionic acidemia

Nicola Brunetti-Pierri, MD, Fondazione Telethon, Italy

This proposal was focused on the characterization of a fish model of propionic acidemia (PA) and on the development of novel therapies. The PA medaka fish model was found to recapitulate several clinical and biochemical features of the human disease, including reduced survival and locomotor activity, hepatic lipid accumulation, increased propionylcarnitine, methylcitrate, and propionate. Moreover, PA fishes showed better survival when fed with low-protein diet.

To gain insight into the disease pathogenesis and to search for potentially novel therapeutic targets, we performed an unbiased 3’-mRNA-Seq and NMR-based metabolome analyses. Both analyses showed global differences between PA and wild-type (wt) medaka. Interestingly, metabolism of glycine and serine resulted affected both at transcriptional and metabolites level and further studies are ongoing to investigate the role of these changes in the disease pathogenesis. Moreover, we found a marked increase in protein propionylation in PA fishes compared to wt controls. Protein propionylation is a post-translational modification occurring under normal conditions but its physiological role is unknown. Like protein acetylation, it is likely involved in regulation of gene expression, protein-protein interactions, and enzyme function. Interestingly, NAD-dependent sirtuins that are responsible of deacetylation of multiple proteins and have also de-propionylating activity, were significantly reduced in PA fishes. We speculated that aberrant protein propionylation in PA is toxic and proteomic studies are ongoing to reveal proteins with aberrant propionylation. With the support of this grant several drug candidates have been also investigated with the goal of developing new pharmacological approaches for PA.

In conclusion, we performed extensive phenotyping of the PA fish model that can be useful to unravel novel disease mechanisms and therapeutic targets.

updated September 2018

Targeting Serine and Thiol Metabolism in Propionic Acidemia

Targeting Serine and Thiol Metabolism in Propionic Acidemia

Hilary Vernon, MD PhD, Johns Hopkins University

While it has been known for several decades that dysfunction of the enzyme propionyl-CoA carboxylase underlies propionic acidemia (PA), many key downstream metabolic adaptions to this primary defect are not well defined. In our research, we developed and studied a new cellular model of PA, with the goals of understanding how the cell is affected in PA, and to identify new pathways for potential treatment targeting.

We initially studied both protein expression in fibroblasts (skin cells) from individuals with PA, and metabolites in urine from individuals with PA, and discovered changes in pathways related to serine metabolism. Serine is an important amino acid that is involved in the synthesis of folate intermediates, glutathione, and other important cellular metabolites. Serine metabolism is of particular interest because it has also been shown recently to be dysregulated in other mitochondrial diseases, and there is a growing interest in how to target this pathway for therapeutic intervention.

In order to more closely study these findings, we developed a new cellular model of propionyl-CoA carboxylase deficiency, where we used CRISPR technology to mutate the PCCA gene in a kidney cell line called HEK293. This new model cell line has important biochemical hallmarks of PA, including absence of the PCCA protein, elevated propionyl-carnitine, very low methylmalonyl-carnitine, and elevated glycine. We discovered that when these cells are in the growth phase, they express genes involved in serine synthesis at higher levels than cells that have normal propionyl-CoA carboxylase activity. We further discovered that the PA cells are very sensitive to deprivation of serine in their culture media, and grow slower than cells with intact propionyl-CoA carboxylase activity. This growth abnormality is not seen when the cells are grown in media that contains serine. Interestingly, we looked at these same pathways in a CRISPR model of methylmalonic acidemia, a closely related disorder to PA, and while we found some overlap in sensitivity to serine, the gene expression patterns we different. This highlights the biochemical uniqueness of PA.  Currently, we are completing flux metabolomics studies in these cells, which will determine exactly what this serine is being metabolized to, and we expect these experiments to be completed by the end of August. In our next steps, we plan to study how treating the cells with different metabolites may alleviate this serine growth defect.

We would like to sincerely thank the Propionic Acidemia Foundation for supporting our research. The funding we received has led to important breakthroughs in our work, and we are excited to continue to move this research forward in the coming years.

updated September 2018

 

Delima Page

Fundraiser for Propionic Acidemia Foundation (PAF) in memory of Lauren and in honour of Jenna

Help Aubrey reach her goal!

Goal:  $2500    Raised as of 11/12:   $1167

Aubrey, mom to Jenna and Lauren will run the Vancouver Fall Classic Half Marathon on Nov. 4th, 2018… at the same time as other PAF parents are running the NY Marathon!

We are fundraising for PAF, as our adult daughter Jenna continues to live with this disorder.  More research and funding arerequired to get closer to finding a cure!  We are optimistic that in Jenna’s lifetime, a cure will be found.

Jenna is now an adult.  She is turning 20 on November 18th!  She graduated high-school life skills and is transitioned to a program called Gateway To Adulthood (GTA).  Jenna’s metabolic status has been stable.  However, last year when Jenna turned 19 she suddenly had her first seizure.  It was a scary time for us as we didn’t understand why she developed epilepsy.  It was happening often.  With a metabolic crisis, we knew our protocol.  Yet, with seizures we had to be alert and constantly in Jenna’s presence, as it could happen at any time.

As with any “normal” teen, Jenna is longing for her independence and seeks the love of a boy.  She admits to being a romantic and wants her prince charming to come one day and sweep her off her feet!  Jenna is quite the fashionista, too.  She wants to (one day) start her own clothing line that she designed.  In her free time, she likes to create stories:  Love stories, to be exact.  She will ask her friends to act out her story. Like a boss director, Jenna knows what she wants and tells everyone their rolls!  We are extremely proud of our daughter.  Once a baby we thought we would not see to live past age of 3, is now a thriving adult and living a beautiful life.

Help us  reach our goal of raising $2500 for PAF!

Below are details of previous year’s fundraising results & photos:

  1. https://laurendelima.weebly.com/2011-10-13-first-anniversary-paf.html
  2. https://laurendelima.weebly.com/2012-10-13-second-anniversary-starlight-foundation.html
  3. https://laurendelima.weebly.com/2013-06-23-run-half-marathon-childrens-wish.html
  4. https://laurendelima.weebly.com/2014-06-21-tough-mudder-canuck-place.html
  5. https://laurendelima.weebly.com/2015-06-14-seek-the-peak-laurens-guardians.html
  6. https://laurendelima.weebly.com/2016-11-18-jennas-princess-ball.html
  7. https://laurendelima.weebly.com/2017-07-23-prospera-valley-mediofondo.html

This year marks the 8th year of the DELIMA family campaign in memory of Lauren.

Thank you for your continued support.

Love, the DELIMA Family




PAF sponsors research on propionic acidemia by minority students – Summer 2017

PAF sponsors research on propionic acidemia by minority students – Summer 2017

Last summer, Propionic Acidemia Foundation (PAF) established a collaboration with Dr. Patricia Schneider from the department of Biology at Queensborough Community College (QCC, Queens, New York) to sponsor a project on the impact of propionic acid in the incidence of autism in Propionic Acidemia (PA) affected individuals. The project was part of the research initiative “Bridges to the Baccaularate”, a National Institute of Health (NIH) funded project that provides resources for a summer research project for minority students. Designed and mentored by Dr. Marisa Cotrina, herself the mother of a PA child, this work investigated the incidence of autism in the propionic acidemia population and the validity of mouse models of autism to study the impact of propionic acid in brain. A unique asset of the project was the utilization of the data collected by the PAF PA International Patient Registry. The authors of the study are currently preparing a manuscript for publication of the results found.

At the end of the project, our student, Sindy Ferreiras, had the opportunity to present her research in the area of Neuroscience at the Annual Biomedical Research Conference for minority students (ABRCAMS) that took place in Phoenix, Arizona last November. Well done, Sindy!

PAF Awards grant for Dr. Oleg Shchelochkov and Dr. Charles P. Venditti for $32,912

PAF awarded a  $32,912 research grant to Oleg Shchelochkov, M.D. and Charles P. Venditti MD, PhD at National Human Genome Research Institute, National Institutes of Health  – 2018

“Diversion of Isoleucine and Valine Oxidative Pathway to Reduce the Propionogenic Load in Propionic Acidemia.”

Patients with propionic acidemia require lifelong protein restriction. In addition to taking a protein restricted diet, many propionic acidemia patients are also prescribed medical formulas. This dietary approach aims to decrease the intake of four amino acids that can become propionic acid. These four amino acids – isoleucine, valine, threonine, and methionine – are called essential, because they cannot be made in the human body and need to be supplied from foods. Too much protein intake creates a situation where excess can lead to a buildup of propionic acid in the body. On the other hand, limiting these four amino acids too much can lead to poor growth. Therefore, patients’ diets are optimized to minimize propionic acid production while encouraging good growth. We wonder whether it is possible to increase dietary protein intake while minimizing the risk of propionic acid buildup.

To answer this question, we are planning to do a series of experiments in zebrafish. Why use zebrafish? Zebrafish share significant similarity to humans in how they process propionic acid. In addition, zebrafish reproduce and mature quickly, which are very important qualities to help search for new drugs that could be used to treat propionic acidemia. Our zebrafish are kept in a special building where the animals are being cared for by a dedicated team that includes scientists, veterinarians, engineers, aquatic specialists, and many others. They check on fish and feed them several times a day, maintain fish tanks, and keep their water very clean.

This type of facility is unique and had enabled our studies of metabolic diseases in zebrafish. Our ongoing studies have shown that zebrafish affected by metabolic diseases have symptoms that are very similar to patients. Even with treatment, affected fish have difficulty growing, get tired easily, have poor appetites and sometimes perish before adulthood. Using special genomic tools, we are planning to change in how the fish processes protein to direct it away from becoming propionic acid. As we make these changes to the biochemical pathways of propionic acidemia zebrafish, we will be carefully watching how these treatments improve their growth, development, appetite and survival. These experiments will help us understand how we can potentially reduce propionic acid toxicity while helping patients achieve a less restrictive diet.

Interview with Joel Pardo  – Summer  2020

Can you tell me about yourself and how you became interested in science?

I was always interested in the sciences. I think ultimately what propelled me towards a career in science was my research experience at the University of California, San Diego. The mentorship I received from Dr. Joshua Bloomekatz helped me develop the ability to reason scientifically and appreciate the opportunities to grow professionally. I learned from him how to design experiments to answer important scientific questions. We often had lengthy discussions about the direction of my project. He helped me make sense of the collection of observations coming from different sources and nurtured my own independent thinking.In gaining an appreciation for his analytical method of thinking, I began to see myself as someday contributing to scientific thinking as a physician-scientist.

During your training at NIH, you worked on a project to find new treatments using zebrafish. What did you find exciting and challenging about studying zebrafish?

Most people are familiar with mice, which are often used in science to find and test new drugs. Working with mice requires a lot of work to have enough animals needed for an experiment. Zebrafish, on the other hand, can produce hundreds of offspring after one breeding cycle. Zebrafish lay eggs directly into water, which also makes it easier to study them soon after they hatch. Somewhat surprisingly,the zebrafish enzymes that handle propionic acid are very similar to the enzymes in humans. These two properties of zebrafish make them an exciting model to study a disease like propionic acidemia.

One of the most challenging parts of my research in zebrafish was their size. Zebrafish offspring are very small, measuring less than a quarter of an inch. I had to spend a lot of time looking at zebrafish under the microscope and learn how to move them around without hurting them. This can be difficult as these small animals are fragile at this young age.

Can you tell us about your PA project?

Earlier in my work, we were able to get zebrafish, which had mutations in the genes linked to propionic acidemia. I needed to understand what propionic acidemia does to zebrafish. We were able to show that propionic acidemia in zebrafish looks a lot like the disease we see in patients. Fish with propionic acidemia had poor appetite, did not grow well, and had difficulty moving. Using special genetic tools, we then attempted to change how zebrafish processed propionic acid and helped them survive longer. Our preliminary results are proving promising, but more work is still needed.

What are your plans after you complete your training at NIH?

The NIH postbac program is a full-time research award for students that have recently completed a bachelor’s degree and are considering a career in science or medicine. I was fortunate enough to join Dr. Charles Venditti’s lab 2 years ago to work on the zebrafish project under Dr. Oleg Shchelochkov. I thoroughly enjoyed my post-bac experience. Looking back on the past 2 years, I feel the lab, and in particular the mentorship of Dr. Shchelochkov, has facilitated and nurtured my growth as a future physician-scientist with roots in propionic acidemia research. In 2019 I applied to MD/PhD programs at several US universities. After having traveled to over half a dozen states and interviewing at many fantastic universities, I ultimately decided upon the physician-scientist training program at University of Minnesota. As I plan my transition to the program, I am currently looking for winter coats.