2021 July Mid Year Report
University of Pennsylvania Cataract Research
The current contribution of the breeders and owners is helping the research to acquire a sufficiently informative dataset that will allow the University team to move the project forward and develop a DNA-based test. Supported by Foundation’s special funding, the DNA and Eye Exam contributions of breeders have greatly increased and this is instrumental for this research moving forward. So good news on that front. Money continues to be a challenge as the Eye Support specific funding is currently low but we were able to cover this last payment from the general fund. Your continued support on all fronts is appreciated by the Foundation Board.
Three documents are below. Two are the report – one a 10-page summary – the other the full 18-page report. The summary report covers research since the last report. The full report covers past history as well. Your choice. Additionally – a NEW cataract testing form to be used for ALL tests moving forward. And finally, a presentation by Dr. Aguirre on details of the research with slide material will be forthcoming in the near future and will be announced when available.
Summary Aguirre Research Report July 2021 (10 pages)
Full Aguirre Research Report July 2021 (18 pages)
Presentation on current research is available on YouTube
2020 Year-End Annual Report
University of Pennsylvania Cataract Research
The Foundation Board is pleased to share with you a full report from Dr. Aguirre’s team that presents a synopsis of the research in 2020. The report is in PDF format below. Please also continue to avail yourself of the information from the July 2020 report and the recording of the August webinar which you will find below as well.
A shout out from Dr. Aguirre and the team for the wonderful support from the breeders in sending in DNA and updating eye exams. And kudos to the Indiana Cocker specialties clinic from which 50 new DNA and new/updated eye exams were gathered for study consideration. As Dr. Aguirre has said- we are setting the bar for other breed clubs. As well, you are reminded to continue to send in DNA samples and updated eye exams on dogs already in the study – whether clear or not.
There is a NEW eye exam form – the only difference is the contact information – and you will find that form below as well. Use that for 2021.
In closing, a reminder that in January a contract extension was completed for 18 months which will take the research into 2022. Fundraising need continues. Thanks for your continued support in all things Eye Support related – DNA, updates, and financial contributions. Keep them coming!
CLICK TO OPEN
August 14, 2020, Dr. Aguirre Cataract Research Presentation – Video Recording
You can access the presentation recording at the link above.
The presentation is 90 minutes long. Times and sections are outlined below for ease of search. Some of the material presented is repeated from the July written report. Starting with GWAS Research at 38 minutes there is ‘color’ commentary that presents information additional to the written report. This continues through to the end of the presentation and especially at the one-hour mark when it breaks into new material and thoughts not shared in the written report. Note that there is a small live screen of Dr. Aguirre presenting from his office that the software captured as well. You can move that screen to the side so it does not impact your viewing of the slides.
Start Introduction and updates on research team / COVID Impact and status
12:22 State of Research Data for 2020 – Updates to Research Approach
38:04 GWAS Research – Details on Analysis of 7 batches of tests
59:13 Results and ongoing analysis
1:05 Q&A Considerations and further actions to end of 2020
Suggested Research Approach Beyond 2020 into 2021
Current thoughts on approaches to 1 or 2 marker tests and why
Overview of requirements for putting marker tests into production
July 2020 Written Research Update
(Companion to Video above)
Below you will find a synopsis of the 20-page report. The full report can be found on this page (scroll down here and select the PDF) and the ASC Foundation website.
We will be recording a presentation from Dr. Aguirre explaining the research progress and hope to have it available to you no later than August 21. The presentation will include updates since this written report and Dr. Aguirre will answer submitted questions as well. To submit any questions you may have, use the email AskTheFoundation@gmail.com and submit questions by August 9.
SYNOPSIS OF PAST & CURRENT WORK
In previously reported work, some candidate genes were selected, and pedigree analyses was done resulting in linking most of the affected dogs to three common ancestors.
An autosomal recessive inheritance model was hypothesized that would explain at least a significant part of the cases. But a deeper analysis of the data suggested that a set of common shared genetic variants causing ALL genetic cataracts in the ACS population was unlikely. It was concluded that some of the cataracts seen may appear similar, but the underlying genetic cause was different. The search to identify the underlying genetic causes continues.
Since the last report, the number of dogs participating in the study increased despite a slowdown in receiving samples during the COVID-19 lockdown and closure of the University. We thank the fancy for their support and ask that they continue as the more samples we receive the stronger the analysis results.
It was previously reported the ACS seems to exhibit distinct sub-phenotypes (sub-groups) of inherited cataracts. Age of onset was one sub-group and is the primary area of concentration. Two age groups emerged, ages 2-5 years and ages 5-9. The second sub-group is where one eye develops a cataract at an early age and several years later a second cataract appears in the other eye. The third sub-group addresses the location of the cataract at the onset. These two latter sub-groups may be analyzed later when a higher number of samples may be available.
To proceed with deeper analysis, 60 of the best samples were divided into 26 affected cases and 34 controls. The number of controls was later increased to 70. Genome-Wide Association Studies (GWAS) with the larger sample size was done on the two age groups which have confirmed the specific chromosomal regions as discussed in previous reports.
All of this led to Whole Genome Sequencing (WGS) for selected cases and controls. Four cases of affected dogs and 4 controls were to be sent to a facility in Switzerland for WGS. Unfortunately, the work was delayed due to the impact of the COVID-19. Results have now been received but the filtering analysis is still underway with the hope of having some of the data available for the August webinar.
Given how complex this disease is in ACS, we hypothesize the occurrence of cataracts in ACS is likely a complex of 2 or more diseases. The selection of the appropriate sub-populations of cases and controls has moved the analyses and project forward. With the candidate region understood and having implemented the Whole Genome Sequencing with the results still pending, the immediate and future objectives are as follows:
- Increase the sample size of the database; a greater number of cases means we enrich the specific sub-populations, and a greater number of controls allowing us to avoid false positives. We believe we have reached a point that allows us to proceed with our plan, but we still need to keep updating our data and gathering new samples, at the very least for validation.
- In this current phase of the project, we are confident we have the candidate regions detected through GWAS that will focus the research using the newly acquired Whole Genome Sequence results.
- The preliminary cross-reference of the data did not point out any specific correlation between unilateral or bilateral, and age of onset. However, once a smaller pool of markers will be available, this relationship will be revisited.
- Whole Genome Sequencing data analysis will be the focus of our next steps. Through the data generated, we will select suitable markers to be tested within the population. Validation will then happen in two steps – through further sequencing, investigating the segregation of candidate variants within the population, and/or conducting further experiments confirming a supposed effect of the variant on gene development.
July 2020 Report – Download PDF here
END OF YEAR
2019 RESEARCH UPDATE REPORT
Click on URL Link below and you will find the complete 2019 annual research report from Dr. Aguirre and University of Pennsylvania Eye Support Cataract Marker research. The key points of this report are:
• As has been stated previously, this research has been more complex than originally thought.
• Since our report in July, research has focused on 2 age groups – 2-5 years and 5-9 years and 3 ancestors in common. Within those groups, research has identified a candidate region within the genome.
• It is believed there are at least 2 genes and possibly a 3rd gene modifier involved
• Moving forward, research will focus on 8 dogs ( 2 cases 2-5 years: 2 cases 5-9 years; and 4 controls)
• The research team will be applying new technology that is expensive but will provide 30% more data in the same amount of time.
• Although the report does not address the likelihood of research completion this year other than to state “…..a lot of work ahead”, if significant progress is made early this year then it might be possible to have a test by year-end. But, this is unlikely.
• Understand that even once the genetics are identified it will be 6 months or more before a test is developed and validated.
• It is the assumption of the Board that fundraising will need to continue at least another year beyond 2020.
The bottom line is we are narrowing the field of research which is real progress. Yet, there remains a lot of work to complete. Dr. Aguirre extends his thank you to everyone for the support given by the breeders and owners for sending in DNA samples, updated eye exams, and other information as required. This is essential for research to be successful and Dr. Aguirre and his research team feel the cocker fancy has really stepped up. Although progress is good, we must not relax on DNA and Eye Exams. The Foundation will be sponsoring another clinic in St. Louis in July at the National and our fundraising chairs will be continuing efforts in that regard as well.
MOLECULAR GENETIC STUDIES OF INHERITED CATARACTS
IN THE AMERICAN COCKER SPANIEL
July 2019 Webinar Update from Dr. Aguirre
The unedited recording and visuals will take approximately 60 minutes to listen to. Click on the link to listen to the webinar and watch the slide content Dr. Aguirre is speaking to. If you would like to see a copy of the presentation slides, see the PDF link below. That is followed by a short synopsis of the presentation and finally, there is a PDF of the entire report in written format (16 pages). Please feel free to utilize these materials as needed and share freely.
PDF of Webinar Slides – 2019 Dr. Aguirre Webinar PDF
Synopsis of Dr. Aguirre’s July 2019 Webinar
Advisory – To fully understand the study status it is strongly suggested you listen to Dr. Aguirre speak to the genetic study process and results as he adds color to the findings and to the techniques. For a quick review, the following summary is offered.
As of June 2019, the study has collected 793 blood samples/records/pedigrees. Dr. Aguirre sends congratulations to the fancy for the participation the past year. While participation was great concern the first two years, it is now excellent and needs to continue. This has allowed the study to find samples that are of interest for further detailed study.
Reminder – the original hypothesis has changed. The working hypothesis had been that the mode of inheritance is simple autosomal recessive based on Dr. Yakely’s 1971 studies. This had directed the initial phase of the research. We now know that is not the case and that inheritance is far more complex. This redirected our efforts to examine this new interpretation of the heritability of cataracts in the American cocker spaniel.
Our Research Approach
• Collect samples from phenotype ascertained dogs: normal (controls) and affected with cataracts (cases).
• Establish what is the minimum age when the dog is considered a control.
• Group cases into ‘reasonable phenotype groupings.
• Reconsider groupings as a number of cases come in. Example: -Initial cases were bilateral ant/post cortical cataracts in 2-5 yr range.-after looking at >200 dogs, and receiving close to 800 records, the early grouping strategy was revised to:
* Cataracts had to be progressive even if they ”looked” inherited.
* Young (2-5 yr) and older (5-8 yrs) cases need to be included.
* Unilateral or asymmetric cataract cases need to be included.
New approach evolved not from wishful thinking but based on data, and the approach was revised based on the study results.
With these groups, the study carried out Genome-Wide Association (GWAS) to identify the chromosomal region of interest and do Whole Genome Sequencing (WGS). We are at this stage now for the first time and that is great news and exciting for the future results.
Genome-Wide Association Study (GWAS)
For those not familiar, GWAS is an observational study of a genome-wide set of genetic variants [aka single nucleotide polymorphisms (SNPs)] in different individuals to see if any variant is associated with a trait like cataracts. It is expensive testing so we are careful to select proper examples. The webinar goes into great detail on this study and should be listened to for more details. GWAS typically focus on associations between SNPs and inherited traits (e.g. coat color, length of hair, defects-CATARACTS).
As long as the trait can be scored accurately in a sufficiently large population of cases and controls, the position of the trait in the genome can be localized. Then the gene/specific defect can be identified. That is our goal and we can now begin to do this type of analysis.
What has been found and what will be done
• Cataracts in the ACS is more complicated than originally suspected
• It is not a single gene (i.e. monogenic) disorder (otherwise the Manhattan plot would have given a single sharp peak shown in the presentation), but cataracts are caused by at least two different genes that result in cataracts that are clinically indistinguishable. There is likely to be a 3rd gene that is a disease modifier.
• Although efforts will focus on dogs in the >2 – 5< yr range (to simplify our effort), GWAS does not distinguish between these dogs and those that are older (>5 – 8<) or with those that have symmetrical bilateral cataracts or unilateral cataracts that over the span of several years develop cataracts in the second eye.
• Now that the study finally has zeroed in on the chromosomal regions of interest, analysis is being done in 4 controls and 4 cases for the chromosomes being studied. More will be done on an as-needed basis. Genomic regions of interest will be screened for associated variants, which will help develop future tools/tests for identifying carriers. Whole-genome sequencing data will be analyzed for variants individually present in cases. We will test the association of identified variants with the disease by sequencing cases and controls
• Our end of year update should prove to be enlightening and we are more hopeful than ever that we are on the right track although with scientific caution the study is not able to determine when the end is in sight at this time.
Full Written Summary of the Webinar – Mid Year July 2019 Report by Dr. Aguirre = Aguirre ACS_Report_15_July_2019
July 2018 Summary by ASC FOUNDATION Scientific Research Chair Doug McFarlane of Mid-Year 2018 Progress Report from Dr. Aguirre at University of Pennsylvania
· Purpose: This report was pulled together two months AFTER the June 2018 podcast with Dr. Aguirre so it contains additional progress information that Doug has distilled. If you want, you can also read the full 13 page PDF of the report found under FILES on the ASC Yahoo Group page. To revisit the podcast – click here: https://fccdl.in/7tjHSukqVp
· Eye Exams Need to Continue: While significant progress has been made in recent months, they still need YOU and YOUR COCKERS – any and all with pedigrees. At this point, Cockers ALREADY INCLUDED in the study are the most important because the true key for the assessment of a phenotype is the observation of the progression – or lack of – overtime. So be sure to bring in your already-included Cockers for re-checks and have your eye vet complete the research paper linked below. http://www.optigen.com/doc/
· The need for funding is never-ending. While DNA contributions and eye exams are finally strong enough, financial contributions have slowed and we need to make up the difference. You will be hearing from the Eye Support Cockers fundraising team shortly but no time like the present to consider a memorial donation for a lost loved one, family member, friend, or fur baby. A wonderful way to support their legacy and the future of our breed. Checks made out to ASC Foundation are tax-deductible too.
On behalf of the ASC Foundation Board of Directors, thank you for your continued support and interest in this important research. Please feel free to share this information on social media, etc. // Charles Born, ASCF Communications Chair.
Summary by ASC FOUNDATION Scientific Research Chair Doug McFarlane of the Mid-Year Progress Report from Dr. Aguirre at University Of Pennsylvania
The Project continues to focus on the identification of the genes and genetic variations responsible for inherited cataracts in American Cocker Spaniels (ACS). The goal is to develop a genetic test that can identify genetically normal, affected and carrier dogs. ACS dogs with inherited cataracts are born with normal lenses, which then proceed to opacify over time, leading to blindness by 2-10 years of age.
The mechanism of inheritance in ACS has been previously proposed as being autosomal recessive, but our recent observations suggests a more complex than one predicted. We have been able to advance our analysis by updating cases and control samples by working with breeders/owners which has become a critical step in our work. The need to keep the database updated has allowed us to pinpoint specific areas of the genome associated in various ways with the condition.
With the enhanced gathering of information about each sample, we were able to select candidate cases for cataracts predictable as having a genetic cause. Specifically, inherited cataracts in ACS are thought to appear sometime around 2-5 years of age and progress. Although now we have found a subset of cases where cataracts presumably inherited begin between 5-9 years of age.
Research going forward will be based on three main principles which are described in more detail in the full report. These principles are (1) Construction of a suitable dataset obtained through the identification of cases (2) mapping of the mutation(s) and (3) validation through sequencing. The ideal result in the study using these principles involves mapping the disease to a specific chromosomal region and sequencing a presumed candidate gene for the validation of the data once the genomic region is identified.
PROGRESS TO DATE
Candidate Genes: As previously reported the selection of candidate genes based on prior data about the same or similar diseases in other dogs and species proved to be negative. There was no association with the cataract physical characteristics. This was more thoroughly discussed in previous progress reports.
Pedigree Analysis: In the previous report we described the analysis of pedigrees to determine a probable mode of inheritance. The work showed no mode of inheritance based on pedigrees alone. However, we were able to link most of our affected subjects to three common ancestors. Therefore, we hypothesized that an autosomal recessive inheritance is at play and that such a model would explain a significant part of our cases. A deeper analysis of the that then suggested that a common, shared mutation causing ALL genetic cataracts in ACS is unlikely.
Samples Received: A significant part of the progress we are making is the increased number of dogs participating to 750 from the 675 previous number. This is a significant increase and is most welcome.
Phenotype Reassessment: With the aid of more samples and the use of the standardized Exam Forms we discovered two distinct Phenotypes (physical characteristics) of inherited cataracts. (1) the possibility of separating in regard to age and onset, and (2) we noted the emergence of a second phenotype, where one eye develops a cataract at an early age and several years later a second cataract appears in the other eye.
Genotyping and Data Analysis: Since our last report, and because of the work carried out in the phenotype reassessment, we were able to increase the suitable sample size for the research to 136 dogs. Dogs were divided into 63 cases and 73 controls. Each cases and controls was further classified by age of onset, type, and location of the cataract and reliability of the sample. With this expanded and better set of data further analysis was done. Again, the aim of this phase of the work was to associate a specific genomic region and its markers to a group of study cases.
Various statistical study techniques were again conducted with this improved data set. The results indicated that roughly 80% of all samples would fall within one of the two sub-populations. Further analysis was done on each of these two populations searching for a specific region in the chromosome where the indications were the strongest. Two specific areas on the chromosome were statistically significant showing a higher indication for the affected area.
Additional analysis continues on these two sub-populations. If these regions detected through this upcoming analysis shows variants causative for cataracts, or at least increasing the dog’s vulnerability to the development of one, this may confirm our hypothesis that not all cases can be explained as inherited from a single parent. The specific strategy going forward is then with more certainty to look for a complex disease which is consistent with our previous report that the occurrence of cataracts in ACS is a probable complex of diseases.
Even though we cannot currently show a simple and complete association of a single marker with the ACS cataract, we have found we can trace and identify trends and associations both under the assumption of a recessive disease, and under the assumption of a disease associated with an area of vulnerability on the chromosome not necessarily inherited in a recessive manner. This might imply the co-existence of multiple factors, not necessarily all of them genetic.
Our work continues as we continue to expand the set of data by working with breeders/owners to gather new and confirm existing data. This is extremely important as we have shown that as the quality of data improves so then does the opportunity to discover and confirm results.
The full report is available, and it describes these results in more detail. I have attempted to summarize 13 pages of a more technical discussion into 3 pages with less technical explanations. I sincerely hope I have properly conveyed the positive research progress that has occurred.
ASCF Scientific Research Chair