The Role of Complex Translocations Associated with TP53 Somatic Mutations for Aiding Prognosis of Canine Diffuse Large B cell Lymphoma

Lymphoma accounts for up to 24% of all cancers diagnosed in pet dogs. Among these cases diffuse large B-cell lymphoma (DLBCL) is the most common subtype. Despite continued advances in veterinary medicine, the response to treatment for canine lymphoma remains highly variable with no reliable means to predict response.

Studies of lymphoma in people have identified characteristic genome changes that have both diagnostic and prognostic significance. In human DLBCL, mutations in the TP53 gene, and genome rearrangements involving the MYC, BCL2 and BCL6 genes have been shown to confer particularly poor prognosis in cases treated with standard of care multi-agent (CHOP-based) chemotherapy.

The investigator’s previous CHF-funded studies have shown that canine cancers, including lymphoma, exhibit genomic changes that are conserved with those observed in the corresponding human cancers, and have identified MYC and BCL2 rearrangements and a high frequency of TP53 mutation in canine DLBCL.

This research will screen a well-defined collection of over 450 pre-treatment, canine DLBCL samples to determine accurate frequencies of these genome changes. The researchers will investigate the correlation of these target aberrations with duration of first remission, and identify key genomic signatures that may aid prognosis of prospective canine lymphoma cases.

The data generated should assist owners and veterinarians with decisions regarding treatment with CHOP. Patients with signatures predictive of poor response to conventional CHOP chemotherapy may benefit from more aggressive treatment at the outset to improve outcome.

Co-sponsored with the AKC Canine Health Foundation, Grant Number: 02317

RESEARCHERS

Dr. Matthew Breen, PhD
North Carolina State University
Amount: $8,000

Disease Risks Associated with Spay and Neuter: A Breed-Specific, Gender Specific

This study extends the investigator’s recently completed AKC Canine Health Foundation-funded project studying 12 dog breeds to identify major differences in the degree to which spay or neuter may be related to an increase in joint disorders (hip dysplasia; cranial cruciate ligament tear) and/or cancers (lymphoma; hemangiosarcoma; and mast cell tumor).

The original breeds studied were: Labrador Retriever, Golden Retriever, German Shepherd Dog, Rottweiler, Boxer, Bulldog, Doberman Pinscher, Dachshund, Corgi (both breeds), Chihuahua, Yorkshire Terrier and Shih Tzu. Findings did not associate an increase in disease association in the small breeds with spaying or neutering, while in larger breeds disease risk was dependent upon gender, and whether the spay or neuter procedure was performed before or after one year of age (Hart, B.L., L.A. Hart, A.P. Thigpen and N. H. Willits. 2014. Long-term health effects of neutering dogs: Comparison of Labrador Retrievers and Golden Retrievers. PLoS ONE 9(7): 10.1371/journal.pone.0102241).

In this second phase, the following breeds have been added to the study: Great Dane, Australian Shepherd, Bernese Mountain Dog, Cocker Spaniel, Border Collie, Beagle, St. Bernard, Irish Wolfhound, Jack Russell Terrier, Pug, Maltese, Pomeranian, Miniature Schnauzer, Boston Terrier, Australian Cattle Dog, Shetland Sheepdog, English Springer Spaniel, Cavalier King Charles Spaniel, and West Highland White Terrier. Upon completion of the study, the major publisher, Wiley, has agreed to place the total data set of all 31 breeds on an open access website as a resource for breeders, dogs owners, researchers and veterinarians.

Co-sponsored with the AKC Canine Health Foundation, Grant Number: 02275

RESEARCHERS

Dr. Benjamin L Hart, DVM, PhD
University of California, Davis
Amount: $3,000 in 2016 with an additional $2,000 in 2017

Veterinary Oncology Fellowship-University of Missouri

Shirley Chu, DVM; University of Missouri

Dr. Chu of the University of Missouri College of Veterinary Medicine is a veterinary oncology resident and graduate student pursuing her PhD. Dr. Jeffrey Bryan serves as Dr. Chu’s mentor for her project on “Examination of the Methylome of Golden Retriever B cell lymphoma”.

Project Abstract: Lymphoma is one of the most common cancers in people and in dogs, and Diffuse Large B cell Lymphoma (DLBCL) is the most common aggressive lymphoma in these species. Dr. Chu’s project is to further understand the effect of DNA methylation on cancer, using epigenetics, the study of potentially reversible changes to nuclear material that ultimately determine DNA expression.

DNA methylation is the most permanent epigenetic mark and has been the most widely studied. DLBCL is the subject of this \study to elucidate the first methylome in the canine species (specifically in Golden Retrievers). MIRAseq (methylated CpG island recovery assay) is an enrichment technique that was used to collect genome wide DNA methylation informion. The methylomes will be analyzed to determine if a distinct fingerprint can be seen in DLBCL in Golden Retrievers, if this fingerprint models human DLBCL, and if a diagnostic panel can be produced for early diagnosis and aid in prognostication.

Other future projects to understand the genetic landscape of cancer in dogs include a parallel whole genome, exome and RNA‐sequencing of DLBCL for the identification of actionable somatic mutations, biomarkers of minimal residual disease, sub-typing, tumor heterogeneity, structural variants and breed related susceptibility.

Co-sponsored with the AKC Canine Health Foundation

RESEARCHERS

Dr. Shirley Chu DVM (mentored by Dr. Jeffrey Bryan)
University of Missouri
Amount: $1,000

Capturing Tumor Cells in Canine Blood

This pilot study’s aim is to develop a blood assay to count tumor cells circulating in a dog’s blood. This technique has proven effective in telling the clinician how aggressive a cancer is and provide information as to treatment and outcome for the patient. This assay is available for humans but currently not for our canine companions.

Just like their human owners, many dogs suffer from cancer, which is often malignant, spreading through the body via blood. Once tumors have spread, they usually result in a poor outcome, including death. The tumor cells in circulation (CTCs) can be counted in the blood of people with cancer using immunocapture devices. The number of CTCs in blood can tell the clinician how aggressive the tumor is, its potential to spread, and how long a patient might survive.

There is currently no such way of detecting CTCs in our canine companions. Development of an assay for counting CTCs in canine blood would be of tremendous benefit to our canine patients because, from a simple blood test, we could detect hidden tumors and gather information on tumor severity and the likelihood of spread or metastasis.

The investigators will test a novel immunocapture microdevice ‐ the GEDI ‐ for counting tumor cells in canine blood. This device can capture CTCs from blood in human patients with various cancers. This study will test its potential to do the same for dogs. In this pilot study, blood samples from healthy dogs will be manipulated to test the ability to count how many added tumor cells are captured by the GEDI device. If the GEDI does capture the tumor cells, the next step will be to determine if the device can capture CTCs from the blood of dogs that are known to have cancer, paving a path to early detection of cancer in dogs.

Co-sponsored with the AKC Canine Health Foundation, Grant Number: 02237-A

RESEARCHERS

Dr. Tracy Stokol, Ph.D.
Cornell University
Amount: $2,000

A Novel Approach for Prevention of Canine Hemangiosarcoma

Hemangiosarcoma, an aggressive form of cancer in dogs, is the cause of death for one out of every five Golden Retrievers in the United States. Portuguese Water Dogs and Boxers also have an especially high risk for this disease which is devastating for all dogs. Hemangiosarcoma is incurable partly because the cancer is detected at a very advanced stage when it is resistant to conventional therapies. Thus, an unconventional approach to improve outcomes for hemangiosarcoma patients will involve effective methods for early detection and for disease prevention.

This project will pair two novel technologies consisting of a patented test to detect hemangiosarcoma cells in blood samples, and a treatment that attacks the cells that establish and maintain the disease. Three milestones will be met: first, will be to expand understanding of the performance and utility of the blood test for cancer in dogs with active disease; second will be to confirm the utility of the test to predict disease progression in treated dogs; and third, will be to establish the performance of the test in the “early detection” setting (dogs at high risk without evidence of active cancer), and thus measure hemangiosarcoma prevention through eradication of the tumor initiating cells with the targeted, investigational drug.

This project will create tools to guide further development, licensing and deployment of these paired technologies against cancer, specifically hemangiosarcoma, with an ultimate goal for disease prevention in all dogs.

Co-sponsored through the collaborative efforts and generosity of the Golden Retriever Foundation and American Boxer Charitable Foundation. The AKC Canine Health Foundation supports the funding of this effort, and will oversee administration of funds and scientific progress reporting, Grant Number: 02234‐MOU

For details and logistics of this project, to include eligibility criteria and how to enroll, please visit Dr. Modiano’s page for the Shine On project.

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Canine Hemangiosarcoma – 2005 Research
Canine Hemangiosarcoma – The Road from Despair to Hope

RESEARCHERS

Jaime F. Modiano, VMD, PhD
University of Minnesota
Amount: $432,000
Administrative Amount: $40,000

Measuring Chemotherapy Drug Resistance in Dogs with T-cell Lymphoma

Quantitative Assessment of Minimal Residual Disease Kinetics during CHOP Chemotherapy of Canine T-cell Lymphoma via Next-Generation TCRVß Sequencing

Human cancer treatment is becoming more and more tailored to each individual case and tumor characteristics. This study will be conducted by researchers at North Carolina State University and will follow a small number of cells that evade chemotherapy agents, providing information on the effectiveness of particular agents against T-cell lymphoma. The data from this study will be used to personalize treatment protocols for dogs on an individual basis in hopes of improving outcomes, survival rates, and quality of life.

Description: Since the use of combination chemotherapy was first reported in 1968, little progress has been made in improving the survival of dogs with T-cell lymphoma. Effectively monitoring chemosensitivity – the number of tumor cells killed by chemotherapy– of individual lymphoma cells exposed to multiple agents over many treatments remains a challenge. Following the small numbers of cells that evade chemotherapy would provide information on the effectiveness of a particular chemotherapy agent.

In this clinical trial, researchers will use state-of-the-art DNA technology to measure changes in this small population of resistant cancerous T-cells in client-owned dogs with lymphoma. Data will be used to personalize treatment protocols for individual dogs in hope of improving survival and quality of life.

Co-sponsored with the Morris Animal Foundation, Grant Number: D16CA-056

RESEARCHERS

Dr. Paul R. Hess, DVM, PhD
North Carolina State University
Amount: $10,000

Discovery of Biomarkers to Detect Lymphoma Risk, Classify For Treatment, and Predict Outcome in Goldens

Lymphoma is a predominant cancer affecting Portuguese Water Dogs as well as numerous other breeds of dogs. Epigenetics is one growing area of research into the behavior of cancer in both medicine and veterinary medicine.

One significant area of epigenetic research involves the investigation of changes to the genome that do not involve a change in the nucleotide sequence. Examples of mechanisms that produce such changes are DNA methylation and histone modification, each of which alters how genes are expressed without altering the underlying DNA sequence. These are important areas of human oncology research that have not been as extensively investigated in veterinary medicine.

This study has the goal of identifying epigenetic markers of lymphoma that can be used to identify dogs at risk, assist with early diagnosis, and aid in the development of individualized treatment plans for affected dogs that can eventually applied to any breed.

Research Objective

Lymphoma strikes 1 in 8 Golden Retrievers, approximately one-third of the cases being B-cell. While T-cell classifications currently inform therapy choices for dogs, B-cell classifications have been investigated little in Golden Retrievers.

Dr. Jeffrey Bryan, in collaboration with Drs. Anne Avery and Heather Wilson will focus their efforts on an area of emerging importance in cancer: epigenetics. Epigenetics is defined as stable and heritable patterns of gene expression that do not entail any alterations to the original DNA sequence. Epigenetic DNA methylation changes clearly underlie development of lymphoma in humans, but have been evaluated minimally in dogs.

Dr. Bryan and collaborators propose to improve diagnostic, classification, and prognostic ability using flow cytometry paired with biopsy to characterize the B-cell lymphomas of Golden Retrievers. They will identify DNA methylation changes in lymphoma cells not present in normal cells to develop biomarkers of each class of lymphoma and identify new therapy targets for affected Goldens.

More significantly, because DNA methylation changes occur so early in the process of cancer formation, they hypothesize that they could serve as biomarkers of risk, allowing medicine or diet to prevent lymphoma in Goldens before it develops. Finally, they propose to identify tumor initiating cells (TIC) in lymphoma biopsies to characterize stem-like cells by surface markers and DNA methylation changes. Identifying these cells will aid therapeutic strategy development. Each project advances a current frontier of research. By performing them in parallel, the markers from each can be combined, correlated, and translated into biomarkers of risk, diagnosis, and prognosis to advance the prevention and management of lymphoma in Golden Retrievers.

Co-sponsored with the AKC Canine Health Foundation, Grant Number: 1918-G

RESEARCHERS

Dr. Jeffery N. Bryan, DVM
University of Missouri, Columbia

Health Implications of Early Spay/Neuter on Canine Health

Most dogs in the United States are spayed or neutered, and the default recommendation has been to perform these elective surgeries prior to physical maturity. However, recent data suggest that early spay and neuter may adversely impact the health and well-being of dogs.

In preliminary studies funded by CHF, Dr. Ben Hart of the UC Davis College of Veterinary Medicine found that early spay or neuter, prior to 12 months of age, was related to a significant increase in risk in five diseases of concern: hip dysplasia; cranial cruciate ligament tear; lymphosarcoma; hemangiosarcoma; and mast cell tumor.

CHF has now funded the second phase of Dr. Hart’s research in which he will expand his work to consider breed differences in vulnerability to joint disorders and risks of various cancers after early or late spay/neuter. Breeds considered will include: Labrador Retrievers, German Shepherd Dogs, and Dachshunds. Rottweilers, Chihuahuas, Standard Poodles, and Miniature Poodles will be included if resources and patient data are available. The expectation is that by inclusion of multiple breeds in phase II Dr. Hart will be able to develop a generalized understanding of the impact of early spay and neuter on disease risk in dogs. This in turn will enable veterinarians and breeders to make data-driven recommendations regarding timing of spay/neuter procedures to reduce the risk of development of multiple devastating diseases.

Co-sponsored with the AKC Canine Health Foundation, Grant Number: 1840

Related News

Long-Term Health Effects of Neutering Dogs: Comparison of Labrador Retrievers with Golden Retrievers

RESEARCHERS

Dr. Benjamin L Hart, DVM, PhD
University of California, Davis

Developing a New Tool to Study Viral Infections and Cancer in Dogs

Development of an MHC Class I Tetramer to Study Virus- and Tumor-specific CD8+ T-cell Responses in Dogs

Summary: Researchers will develop a state-of-the-art molecular tool to track and study killer T-cell populations that are responsible for fighting viral infections and cancer in dogs.

Description: In humans, a powerful immunologic reagent called a tetramer is standardly used to visualize changes in the body’s killer T-cells. These cells respond to immunologic challenges and are critical to the body’s immune system.

Current knowledge of T-cell behavior in dogs could be significantly advanced with the development of a dog-specific tetramer. Researchers will work to construct the first canine tetramer, which would then be used in the development of vaccines for infectious diseases and cancer in dogs.

Co-sponsored with the Morris Animal Foundation, Grant Number: D15CA-015

RESEARCHERS

Dr. Paul R. Hess
North Carolina State University

Innovations in Prevention, Diagnosis, and Treatment of Cancer

Lymphoma and hemangiosarcoma are major health problems in Portuguese Water Dogs. This study collaborates with several top universities and researchers who are utilizing cutting edge technology to identify several regions of the genome that contain genetic, heritable, risk factors for lymphoma and hemangiosarcoma in Golden Retrievers. This information can then be applied to other breeds, including Portuguese Water Dogs. Tumor-specific mutations, when identified, can be clinically applied to reduce the incidence of these cancers, identify cancer earlier, and modify treatment plans to the individual patient.

Research Objective

Lymphoma and hemangiosarcoma are major health problems in Golden Retrievers, causing both suffering and premature death. Through ongoing collaboration, Drs. Jaime Modiano, Matthew Breen, and Kerstin Lindblad-Toh have identified several regions of the genome that contain genetic heritable risk factors for lymphoma and hemangiosarcoma in Golden Retrievers. They have tumor-specific mutations that occur recurrently in both cancers, some of which are linked to duration of remission when treated with standard of care. Their results indicate that a few heritable genetic risk factors account for as much as 50% of the risk for these cancers.

These findings offer the potential to develop tests and strategies for DNA tests that can predict risk for individual dogs, as well as to manage risk across the population as a whole. Indeed, both the inherited risk factors and tumor mutations point to pathways that have been implicated in the pathogenesis of lymphoma and hemangiosarcoma, and thus should inform the development of targeted therapies. In the current study, Drs. Modiano, Breen, and Lindblad-Toh will find the precise mutations for the heritable genetic risk factors and to validate markers (mutations) used to determine risk at the heritable loci in a larger independent population of Golden Retrievers from the United States and from Europe in order to develop robust risk prediction tools and an accompanying DNA test. Further, they will identify and characterize tumor mutations and study their relationship to the heritable risk factors, tumor pathogenetic mechanisms, and disease outcome.

Co-sponsored with the AKC Canine Health Foundation, Grant Number: 1889-G

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RESEARCHERS

Dr. Jaime F Modiano, VMD PhD
University of Minnesota

Matthew Breen, PhD, CBiol, FSB
North Carolina State University