Neurological Research Update, 2013.
Thank you for the opportunity to update everyone on our collaborative genetic studies of polyneuropathy (LPN) and other neurological disorders in the Leonberger breed.
1. We can report some additional work surrounding the first causative LPN mutation now known as the LPN1 mutation. Four Saint Bernards affected with similar clinical signs and identical nerve biopsy alterations as Leonbergers were each found to have two copies of the LPN1 mutation, and therefore have an identical basis for their disease as do Leonbergers with LPN1. So, the Leonberger clubs have helped to find a cause for a genetic problem in a related breed! We are now working to determine the frequency of the LPN1 gene in a population of healthy Saint Bernards to determine its overall effect on the health of this breed might be. Lastly, we are currently editing the LPN1 manuscript and I can see no reason why this will not be submitted by the end of summer to a scientific journal for peer review.
2. Although it is clear that two copies of the LPN1 mutation cause early-onset severe LPN, the effects of a single copy of the LPN1 mutation on disease susceptibility is not completely understood. This is because many dogs with a single LPN1 copy have no LPN signs throughout their lives, while others with a single copy have LPN. To add to the confusion, dogs without the LPN1 mutation at all can also have LPN disease. Although, the average age of onset of LPN signs for dogs with either a single or no LPN1 gene copy is around 6 years old, there can be earlier onset in some of these dogs. To make a long story short, this very much suggests that multiple genetic bases for LPN-like disease exist in Leonbergers.
Figures from the Minnesota sample collections indicate that if two copies of the LPN1 mutation are always required for disease signs, this would explain approximately 18% of all LPN cases. And, if there is indeed an influence of a single copy of the LPN1 mutation on disease then it could contribute to as much as 40% of all LPN cases.
3. Minnesota and Bern have therefore, with your support, invested considerable time and resources trying to nail down chromosomal locations of other LPN genes. Much of the focus has been on a region of canine chromosome 7, found in our original gene-mapping study that found the LPN1 mutation. This chromosome 7 region is borderline significant for association with older onset LPN, depending on which sample sets of affected and control dogs are utilized in the analyses. Nevertheless, we have performed quite a bit of sequencing of genes in that region hoping to find a profound genetic alteration that could perhaps explain some of the LPN cases, but no such mutations have been found.
Our other gene-mapping studies to find additional LPN gene locations have attempted to consider effects of different ages of onset, gender, and co-inheritance of single LPN1 alleles, on LPN in the entire Leonberger cohort or subsets of it. The result thus far is that despite including 121 cases and 72 controls in these analyses, we have not found additional significantly associated LPN loci that withstand statistical scrutiny. That is not to say that other potential LPN loci sometimes appear that have borderline hints of association, and that we examine these regions very thoroughly for possible neural genes of interest that, if mutated could cause a neurological disease, but they do not prove to be statistically and reproducibly significant when appropriate corrections for the overall population relatedness are taken into account.
Consequently, we are left with the possibility that there are multiple genes and mutations in Leonbergers that can contribute to LPN-like diseases (LPN1, as well as LPN2, perhaps LPN3, etc) with similar sorts of clinical signs due to mutations in different genes. A related possibility is that other LPN-like diseases are more genetically complex, and result from combinations of mutations in multiple genes at the same time, that makes it difficult to find the chromosomal locations unambiguously with the current sample population. We will seriously need to consider whether more whole genome gene-mapping data must be gathered to improve our statistical power to identify these additional loci and enable more productive gene sequencing and mutation discovery work.
4. In collaboration with the University of Utrecht, we have made progress on the LEMP nervous system disorder (leukomyeloencephalopathy), and have a very solid chromosomal locus for this condition that was obtained by gene-mapping of only 12 LEMP cases (the large population of control dogs for LPN could be used as controls for this study as well). We are now at the stage with Bern of trying to find a LEMP mutation from whole genome sequencing data. One interesting note is that several dogs classified as LPN on our submission forms actually have LEMP as determined from the genetic marker data.
We have only about a dozen LEMP cases in our collection and we are quite sure it is a young onset recessive disease. We do not know the extent to which true LEMP cases are being reported and accurately diagnosed. At this time without a mutation test we cannot say for certain how prevalent it truly is in the breed. We think we are in a pretty good position, with a bit more funding, to find this mutation and we can then go from there to see how prevalent it really is. It may turn out to not be a major problem as a whole, or the most important problem, but it also might be something that can be dealt with before it becomes so.
We do think it possible that some true cases of LEMP are being lumped in by the public and inexperienced veterinarians with neurological conditions such as polyneuropathy. This was evidenced by the example mentioned above where the paperwork information led us to include the case in our attempts to identify an LPN2 gene location. But when we compared the genetic data from the 10 or so true LEMP cases with this dog it was seen that this dog clearly had the LEMP genetic profile. It could also very well have a form of LPN as well I suppose.
It brings up the issue, again, that there are probably multiple disorders present in Leonbergers, even several neurological disorders. Multiple genetic issues due to founder effects is the case with all breeds of course and maybe just a little more of a problem in Leonbergers due to their history.
Clustering would be expected with a relatively infrequent severe recessive disease because both parents must be carriers at minimum and then one quarter of the pups would be affected. It is unlikely that many LEMP cases are being bred due to the early and severe signs, but if cases are mated with carriers half of the pups would be affected. If a clear dog is one of the mating pair it would prevent any LEMP cases from being produced.
Your group has been very generous and supportive to get us all this far, but, perhaps it is not a surprise that additional financial support will be needed to advance this cause much further.
Sincerely,
Jim Mickelson, PhD
Professor Veterinary Biochemistry and Genetics
University of Minnesota
Thank you for the opportunity to update everyone on our collaborative genetic studies of polyneuropathy (LPN) and other neurological disorders in the Leonberger breed.
1. We can report some additional work surrounding the first causative LPN mutation now known as the LPN1 mutation. Four Saint Bernards affected with similar clinical signs and identical nerve biopsy alterations as Leonbergers were each found to have two copies of the LPN1 mutation, and therefore have an identical basis for their disease as do Leonbergers with LPN1. So, the Leonberger clubs have helped to find a cause for a genetic problem in a related breed! We are now working to determine the frequency of the LPN1 gene in a population of healthy Saint Bernards to determine its overall effect on the health of this breed might be. Lastly, we are currently editing the LPN1 manuscript and I can see no reason why this will not be submitted by the end of summer to a scientific journal for peer review.
2. Although it is clear that two copies of the LPN1 mutation cause early-onset severe LPN, the effects of a single copy of the LPN1 mutation on disease susceptibility is not completely understood. This is because many dogs with a single LPN1 copy have no LPN signs throughout their lives, while others with a single copy have LPN. To add to the confusion, dogs without the LPN1 mutation at all can also have LPN disease. Although, the average age of onset of LPN signs for dogs with either a single or no LPN1 gene copy is around 6 years old, there can be earlier onset in some of these dogs. To make a long story short, this very much suggests that multiple genetic bases for LPN-like disease exist in Leonbergers.
Figures from the Minnesota sample collections indicate that if two copies of the LPN1 mutation are always required for disease signs, this would explain approximately 18% of all LPN cases. And, if there is indeed an influence of a single copy of the LPN1 mutation on disease then it could contribute to as much as 40% of all LPN cases.
3. Minnesota and Bern have therefore, with your support, invested considerable time and resources trying to nail down chromosomal locations of other LPN genes. Much of the focus has been on a region of canine chromosome 7, found in our original gene-mapping study that found the LPN1 mutation. This chromosome 7 region is borderline significant for association with older onset LPN, depending on which sample sets of affected and control dogs are utilized in the analyses. Nevertheless, we have performed quite a bit of sequencing of genes in that region hoping to find a profound genetic alteration that could perhaps explain some of the LPN cases, but no such mutations have been found.
Our other gene-mapping studies to find additional LPN gene locations have attempted to consider effects of different ages of onset, gender, and co-inheritance of single LPN1 alleles, on LPN in the entire Leonberger cohort or subsets of it. The result thus far is that despite including 121 cases and 72 controls in these analyses, we have not found additional significantly associated LPN loci that withstand statistical scrutiny. That is not to say that other potential LPN loci sometimes appear that have borderline hints of association, and that we examine these regions very thoroughly for possible neural genes of interest that, if mutated could cause a neurological disease, but they do not prove to be statistically and reproducibly significant when appropriate corrections for the overall population relatedness are taken into account.
Consequently, we are left with the possibility that there are multiple genes and mutations in Leonbergers that can contribute to LPN-like diseases (LPN1, as well as LPN2, perhaps LPN3, etc) with similar sorts of clinical signs due to mutations in different genes. A related possibility is that other LPN-like diseases are more genetically complex, and result from combinations of mutations in multiple genes at the same time, that makes it difficult to find the chromosomal locations unambiguously with the current sample population. We will seriously need to consider whether more whole genome gene-mapping data must be gathered to improve our statistical power to identify these additional loci and enable more productive gene sequencing and mutation discovery work.
4. In collaboration with the University of Utrecht, we have made progress on the LEMP nervous system disorder (leukomyeloencephalopathy), and have a very solid chromosomal locus for this condition that was obtained by gene-mapping of only 12 LEMP cases (the large population of control dogs for LPN could be used as controls for this study as well). We are now at the stage with Bern of trying to find a LEMP mutation from whole genome sequencing data. One interesting note is that several dogs classified as LPN on our submission forms actually have LEMP as determined from the genetic marker data.
We have only about a dozen LEMP cases in our collection and we are quite sure it is a young onset recessive disease. We do not know the extent to which true LEMP cases are being reported and accurately diagnosed. At this time without a mutation test we cannot say for certain how prevalent it truly is in the breed. We think we are in a pretty good position, with a bit more funding, to find this mutation and we can then go from there to see how prevalent it really is. It may turn out to not be a major problem as a whole, or the most important problem, but it also might be something that can be dealt with before it becomes so.
We do think it possible that some true cases of LEMP are being lumped in by the public and inexperienced veterinarians with neurological conditions such as polyneuropathy. This was evidenced by the example mentioned above where the paperwork information led us to include the case in our attempts to identify an LPN2 gene location. But when we compared the genetic data from the 10 or so true LEMP cases with this dog it was seen that this dog clearly had the LEMP genetic profile. It could also very well have a form of LPN as well I suppose.
It brings up the issue, again, that there are probably multiple disorders present in Leonbergers, even several neurological disorders. Multiple genetic issues due to founder effects is the case with all breeds of course and maybe just a little more of a problem in Leonbergers due to their history.
Clustering would be expected with a relatively infrequent severe recessive disease because both parents must be carriers at minimum and then one quarter of the pups would be affected. It is unlikely that many LEMP cases are being bred due to the early and severe signs, but if cases are mated with carriers half of the pups would be affected. If a clear dog is one of the mating pair it would prevent any LEMP cases from being produced.
Your group has been very generous and supportive to get us all this far, but, perhaps it is not a surprise that additional financial support will be needed to advance this cause much further.
Sincerely,
Jim Mickelson, PhD
Professor Veterinary Biochemistry and Genetics
University of Minnesota
