Drug – bio-affecting and body treating compositions – Antigen – epitope – or other immunospecific immunoeffector – Amino acid sequence disclosed in whole or in part; or...
Reexamination Certificate
1999-07-30
2001-04-03
Smith, Lynette R. F. (Department: 1645)
Drug, bio-affecting and body treating compositions
Antigen, epitope, or other immunospecific immunoeffector
Amino acid sequence disclosed in whole or in part; or...
C424S234100, C424S282100, C424S184100, C424S828000, C530S300000, C530S324000, C530S350000, C530S825000, C435S007100, C435S975000
Reexamination Certificate
active
06210676
ABSTRACT:
BIBLIOGRAPHY
Complete bibliographic citations of the references referred to herein by number in parentheses can be found in the Bibliography section, immediately preceding the claims.
FIELD OF THE INVENTION
This invention relates to compositions and methods useful for the prevention, treatment and early diagnosis of Lyme disease in humans and other animals. More particularly, this invention relates to outer surface protein (Osp) polypeptides which are able to elicit in a patient the formation of a specific immune response which is effective to diagnose, predict successful eradication of infection or protect against Lyme disease in a mammalian host. This invention also relates to a screening method to detect anti-Osp borreliacidal antibody activity, and antibodies which react with a protein fragment encoded by a DraI-SmaI DNA fragment of OspC. Also within the scope of this invention are antibodies directed against Osp polypeptides, diagnostic kits comprising the antibodies or the polypeptides, and vaccines using borreliacidal epitopes of OspA, OspB or OspC or a conserved DNA sequence fragment together with or without a vaccine carrier.
DESCRIPTION OF THE PRIOR ART
Lyme disease (
Lyme borreliosis
) is spread by a bite from an infected tick, and is the most commonly reported tick-borne infection in Europe and North America (1). This multi-system disorder has caused significant morbidity worldwide.
Lyme disease is caused by the spirochete
Borrelia burgdorferi
(
B.b.
), which is primarily transmitted during the blood feeding of
Ixodes ssp
ticks. Initially, spirochetes infect the skin and, in the majority of early cases, cause an erythema migrans lesion (2). Those stricken may not become ill for weeks, and nervous system symptoms (headaches, dizziness, hearing problems, tingling and trouble concentrating) may not occur for weeks or months. It is now known that infection can spread to the nervous system or joints, and the risk of neurological or joint complications increases the longer the disease goes untreated. Infection can be asymptomatic, or have a range of clinical presentations, depending on the tissues affected, the duration of infection, host factors such as the vulnerability of the immune system and immunogenetic factors which could predispose a patient to the development of certain complications.
The treatment of symptomatic patients is currently with a range of antibiotics, e.g., tetracyclines, penicillin and cephalosporins, but studies show mixed results. Left untreated, the bacteria can spread to the central nervous system, heart, brain, or joints, causing arthritis, cardiac infections and neurologic problems, and in rare cases, death (3-6).
Upon infection with borrelia, B cells in the body begin producing antibodies which recognize the foreign organism. There are at least two functional types of antibodies produced in response to a borrelia infection. One response is a nonspecific binding/opsonizing (coating) response which “marks” the antigen and may result in ingestion of
B.b.
by phagocytic cells. These non-specific antibodies are produced against proteins common among several bacterial species (viz. 41 kDa proteins to many bacterial flagella). Thus, these antibodies will recognize and attach to similar antigens on other bacteria. Because of this, diagnostic tests which detect these non-specific binding/opsonizing antibodies are generally nonspecific.
A second functional antibody response is the production of borreliacidal (lethal) antibodies which specifically recognize epitopes on some individual proteins of the
B.b.
organisms. After attachment of these antibodies to the
B.b.
organisms, complement interacts with the antibodies to form a membrane attack complex which kills the
B.b.
organism without the necessity of scavenging by phagocytic cells. This highly specific borreliacidal antibody response is often detectable within the first 2 weeks of infection. The successful detection and induction of borreliacidal antibodies is gaining importance in the Lyme disease diagnostic and prevention armamentarium.
Shortly after the discovery of
Lyme borreliosis
, researchers determined that vaccination of experimental animals with whole
B.b.
provided protection against challenge (7,8). Additional studies established the role of antibody-mediated protection and confirmed the ability of vaccination with
B.b.
to induce antibodies which provide protection against
B.b.
infection (9-11). To date, vaccination of animals with Osps of
B.b.
, especially OspA (12-14), OspB (12,14), and OspC (14-16), have provided protection against infection with the Lyme spirochete. Protection after vaccination with OspA and OspB have been shown to be due to the induction of borreliacidal antibodies which specifically killed the
B.b.
organisms (13,14,17-24). In contrast, anti-OspC borreliacidal antibodies have not been detected after vaccination (14-16) and investigators have postulated that protection after vaccination with OspC is due to other mechanisms (16).
Most of the efforts to date have focused on developing an OspA vaccine primarily because of the large amounts of OspA expressed on the surface of many
B.b.
laboratory isolates. To date, most borrelia spirochetes have had outer surfaces comprised mainly of OspA. Therefore, it has been believed that inducing borreliacidal antibodies against OspA would provide protection against the spirochetes. SmithKline Beecham (Philadelphia, Pa.) and Pasteur Merieux Connaught (Lyon, France) have developed vaccines based on the generation of borreliacidal antibodies to OspA. The SmithKline Beecham vaccine has been approved for general use, and the Pasteur Merieux Connaught vaccine is currently being assessed by the U.S. Food and Drug Administration. As one would expect, OspA vaccines have been shown to be effective in animal models when the animals have been needle challenged. In addition, OspA vaccination has provided protection against ticks infected with
B.b.
However, protection against a tick challenge has been dependent on the presence of high levels of anti-OspA borreliacidal antibodies. Schwan et al. (28) recently demonstrated that spirochetes in infected ticks downregulate OspA on their surface during ingestion of a blood meal. Thus, OspA vaccines must induce high titers of anti-OspA borreliacidal antibodies to destroy the spirochetes in the midgut of infected ticks before they downregulate OspA. Therefore, the duration of high titers of anti-OspA borreliacidal antibodies is a critical determinant of the long-term efficacy of an OspA vaccine.
The applicants recently demonstrated the inability of a commercial OspA vaccination to sustain adequate levels of anti-OspA borreliacidal antibodies in humans (23). It is also unlikely that an anamnestic response will occur quickly enough to eliminate
B.b.
organisms from infected ticks. In support, infection with
B.b.
has been documented in OspA-vaccinated humans and dogs (26,27). These results highlight the need to evaluate other
Lyme borreliosis
vaccine components.
In addition, Lyme disease is ordinarily diagnosed by detecting antibodies in the blood or cerebral spinal fluid, but the most commonly used tests are often inaccurate. False-negative, and more commonly, false-positive results continue to plague the serodiagnosis of Lyme disease. Several schemes using conventional diagnostic assays have been developed to more accurately detect Lyme disease. Unfortunately, little improvement has occurred and misdiagnosis continues to cause significant economic and health effects. In addition, the recent approval of an OspA Lyme disease vaccine will further confound conventional diagnostic testing. Thus, a sensitive and specific Lyme disease test which can be made widely available as a commercial kit and can discriminate between vaccinated individuals and patients with Lyme disease is still needed.
Detection of borreliacidal antibodies may also solve this problem. Borreliacidal antibodies have been shown to serve as the basis for a sensitive and highly specific serodiagnostic test (17,25-27
Callister Steven M.
Jobe Dean A.
Lovrich Steven D.
Schell Ronald F.
Devi S.
DeWitt Ross & Stevens SC
Gundersen Lutheran Medical Foundation
Smith Lynette R. F.
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