Equine disease model for herpesvirus neurologic disease and uses thereof

Abstract

The present invention relates to an in vivo equine disease model for equine herpesvirus-1 neurological disease comprising a horse having a low pre-exposure level of herpesvirus-specific CTL precursors and / or is approximately 20 years of age or older wherein the horse is experimentally infected with a neuropathogenic strain of equine herpesvirus or a mutant thereof. The invention includes a method of preparing an in vivo equine disease model for equine herpesvirus-1 neurological disease comprising obtaining a horse that possesses low pre-infection levels of EHV-1 specific CTL precursors and / or is approximately 20 years of age or older and inoculating the horse intranasally with an effective infecting amount of a neuropathogenic strain of EHV-1. A particularly preferred method involves the advanced neurological disease stage when the experimental horse presents clinical signs of myeloencephalopathy. Additionally, the invention concerns a method of quantifying the risk factors and predicting the development of clinical neurologic signs of equine herpesvirus-1 neurological disease in a horse comprising the steps of (a) determining the pre-infection CTLp frequency to be less than approximately 40 EHV-1 specific CTLp per 106 PBMC; and (b) determining the post-infection viremic load following exposure to EHV-1 to be approximately 10-fold or more over the viremic load present in horses following exposure to a non-neuropathogenic strain of EHV-1. Also described in the invention is the determination of the risk of developing the clinical neurologic signs by use of an equation y=a+bx wherein y is the peak viremic load, a=2.97, b=−0.027 and the variable x is the pre-infection CTLp frequency. Lastly, the invention deals with a new live, attenuated vaccine formulation that is effective against neurologic disease due to equine herpesvirus-1.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS[0001]This application is a continuation of U.S. application Ser. No. 13 / 054,608, now U.S. Pat. No. 9,642,908, which application was accepted on Jun. 2, 2011 under 35 U.S.C. §371 based on PCT International Application No. PCT / US2009 / 004409, filed on Jul. 29, 2009, which claims the priority benefit under 35 U.S.C. §119(e) of U.S. Provisional Application No. 61 / 137,412, filed on Jul. 30, 2008, abandoned. All prior disclosures are hereby incorporated by reference in their entirety into the present disclosure.STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT[0002]Not ApplicableNAMES OF PARTIES TO A JOINT RESEARCH AGREEMENT[0003]Not ApplicableREFERENCE TO A “Sequence Listing”[0004]A Sequence Listing provided in this application as a text file identified as “2712-06C1_SeqList_PCT_AM103059_ST25” is incorporated herein by reference thereto. The Sequence Listing was created on Sep. 16, 2009 and the size of the text file is 1,380 bytes.BACK...

AI Technical Summary

Benefits of technology

The present invention provides a new and effective way to evaluate the efficacy of vaccines against equine herpesvirus-1 (EHV-1) and protect horses from developing neurological disease. The use of a live, non-neuropathogenic strain of EHV-1 as a challenge agent in horses with low levels of antiviral cellular immunity is a unique and validated model for testing the protective efficacy of vaccines against EHV-1 myeloencephalopathy. The invention also includes the development of live, attenuated viral vaccines containing a non-neuropathogenic strain of EHV-1 and the use of various routes of administration for the vaccine. The invention has practical benefits for the horse industry as it provides a valuable tool for the development and validation of new vaccines against EHV-1.

Problems solved by technology

Characterized by high neurologic morbidity and case fatality rates, resistance to prevention by vaccination, and the ability to affect horses of all breeds and ages, EHV-1 myeloencephalopathy has the potential for causing catastrophic losses to both the welfare of horses and the economy of equine-based businesses (M. T. Donaldson and C. R. Sweeney, “Equine herpes myeloencephalopathy,”Compend Contin Edu Pract Vet 19:864-871 (1997); W. D. Wilson, “Equine herpesvirus 1 myeloencephalopathy,”Vet Clin North Am Equine Pract 13:53-72 (1997); R. W. Henninger et al., “Epidemic neurologic disease due to equine herpesvirus-1 at a university equestrian center,”J Vet Intern Med 21:157-165 (2007); C. van Maanen et al., “Neurological disease associated with EHV-1 infection in a riding school: clinical and virological characteristics,”Equine Vet J 33:191-196 (2001); M. J. Studdert et al., “Outbreak of equine herpesvirus type 1 myeloencephalitis: new insights from virus identification by PCR and the application of an EHV-1-specific antibody detection ELISA,”Vet Rec 153:417-423 (2003); S. M. Reed and R. E. Toribio, “Equine herpesvirus 1 and 4,”Vet Clin North Am Equine Pract 20:631-642 (2004); T. A. Jackson and J. W. Kendrick, “Paralysis of horses associated with equine herpesvirus 1 infection,”J Amer Vet Med Assoc 158:1351-1357 (1971); K. M. Charlton et al., “Meningoencephalomyelitis in horses associated with equine herpesvirus 1 infection,”Vet Pathol 13:59-68 (1976); P. B. Little and J. Thorsen, “Disseminated necrotizing myeloencephalitis: a herpes-associated neurological disease of horses,”Vet Pathol 13:161-171 (1976); H. Platt et al., “Pathological observations on an outbreak of paralysis in broodmares,”Equine Vet J12:118-126 (1980); R. S. Greenwood and A. B. Simpson, “Clinical report of a paralytic syndrome affecting stallions, mares and foals on a Thoroughbred studfarm,”Equine Vet J 12:113-117 (1980); K. E. Whitwell and A. S. Blunden, “Pathological findings in horses dying during an outbreak of the paralytic form of Equid herpesvirus type 1 (EHV-1) infection,”Equine Vet J24:13-19 (1992); C. W. Kohn and W. R. Fenner, “Equine herpes myeloencephalopathy,”Vet Clin N Am:Equine Pract 3:405-419 (1987)).

However, the Kydd et al. study dealt solely with abortion as a disease outcome of EHV-1 infection and did not address the neurological manifestation of EHV-1 infection.

Of particular alarm to the equine industry is its recent targeting of horses in riding / boarding stables and of young horses assembled at race track venues for training and racing, with consequent high mortality rates and severe economic losses to the boarding and racing sectors of the industry.

Although efforts to develop a more effective, second-generation vaccine against the neurologic herpesvirus disease are underway by several vaccine manufacturers, no equine disease model exists for assessing the effectiveness of such experimental vaccines.

In fact, a practical neurological disease model has never been created, either in connection with standard laboratory mice, rats or guinea pigs where the neurological effects of EHV-1 infection can never develop, or even from prior equine studies of EHV-1 in horses or ponies.

Furthermore, the ponies never reached the point where they presented any neurological signs.

Their blood analysis and physical characteristics are of limited value, consequently, in determining the factors giving rise to neurological disease, the traits of a neurological disease model or the criteria useful in predicting development of neurological disease.

First of all, the EHV-1 infection in the mouse never can progress to express the same breadth of neurological signs that plague the natural host, namely, horses, thus failing to provide a complete picture as a viral vaccine candidate model to permit adequate vaccination strategies in horses.

Secondly, a major adverse effect of infection in pregnant mares is the induction of spontaneous infectious abortions, which cannot be duplicated or investigated in the vaccination studies run in the classic murine model.

Thirdly, without a solid understanding of the clinical signs and progression of the EHV-1 disease in horses, the vaccine candidates will not have broad applicability and activity to prevent more serious neurologic signs than simple infections caused by the single etiological agent in the mouse.

Fourthly, the detection of new mutant equine herpesviruses has further limited the application of the murine model as a practical animal model for vaccine purposes.

The patentees note the limitations of using the standard murine model to confirm protective activity in horses.

Patentees further complain that the level of protection against challenge infection and ultimately in preventing abortion in pregnant mares can only be established in the target animal and, thus, the initial murine testing is of limited value in the final analysis.

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