Periodontal disease is one of the oldest and most common diseases of man. It is apparent in human fossil remains and occurs in otherwise healthy individuals. Today, periodontal disease represents a major worldwide health problem. The disease is a result of the accumulation of dental plaque at the gingival margin. There are two broad classes of periodontal disease which roughly approximates the degree or severity of the pathology: gingivitis and periodontitis.
Gingivitis is an inflammation of the marginal gingival tissue due to the accumulation of dental plaque. For the most part, gingivitis is characterized by redness, swelling and bleeding of the gingival tissue. The extent and severity of these characteristics indicate the degree to which the disease has progressed. Periodontitis is characterized not only by the inflammation of the marginal gingivae, but also by loss of the attachment of the periodontal ligament, loss of alveolar bone and loss of the epithelial attachment due to apical migration. The pathological consequences of these physiological losses is the formation of a periodontal pocket, which can become infected, and thus be the source of bacterial infiltration into the host. The progression of established gingivitis to an advanced lesion may well lay the foundation for periodontitis.
The literature indicates that there are significant microbial population shifts from sites of gingival inflammations to subgingival pockets. Certain identified and specific bacterial organisms are known to be responsible for periodontal disease in humans; however, other organisms may also contribute to the severity of the disease. In addition, results from clinical studies show a correlation between the presence of certain microbial species and different types and degrees of severity of periodontal disease. There is a cause-and-effect relationship between the presence and quantity of plaque, containing a wide variety of colonized bacterial strains, and periodontal disease. It therefore follows that, by limiting plaque, the extent and severity of periodontal disease can be controlled.
Both chronic gingivitis and chronic periodontitis share two important characteristics which may be the clue to their sequential relationship. Both conditions are usually painless until their more advanced stages and both pathologies have an absolute requirement for bacterial plaque before the sequence of these conditions progress and develop into advanced periodontal disease. While there are secondary systemic and external factors which affect the extent the disease, the most important factor, and one that provides the greatest promise of being controllable, is the relationship between bacterial plaque and periodontal disease.
The disease begins its progression through an accumulation of bacterial plaque at the gingival margin. As the pathology progresses, there is chronic inflammation of the gingiva and periodontal ligament, with subsequent degeneration of various gingiva-tooth structures. The chronic inflammation is exacerbated by calculus formed from mineralized plaque at the various tissue interfaces and in the periodontal pocket. Epithelial tissue migration into inflamed and necrotic areas can engulf plaque structures, resulting in abscesses accompanied by purulent exudate. The final and most severe stage of periodontal disease is the resorption of alveolar bone and the eventual exfoliation of the tooth.
Plaque is a heterogeneous mixture of bacterial aggregations embedded in a sticky matrix. While bacterial composition of plaque ranges from 50 to 70 percent, the matrix is derived from dead cells, salivary glycoproteins and serum proteins that are laid on a polysaccharide backbone. The bacteria synthesize the polysaccharides for the plaque backbone as a step in their own colonization process. In addition to the viable bacteria and the matrix, plaque also contains food debris, small numbers of epithelial cells, white blood cells and various other components which are derived from the host and the host's activities.
The formation and development or proliferation of plaque occurs in two stages. The first step may require a base layer of salivary glycoproteins on the tooth's surface as well as on the soft tissue in the oral cavity. This base organic layer, derived from saliva, is adsorbed onto the surface and forms an acquired pellicle. This insoluble acquired pellicle serves as the foundation for supragingival plaque. The second step is the bacterial colonization by "pioneering" bacteria of the acquired pellicle. Once the bacteria have attached to the surface of a structure, they aggregate, develop colonies and plaque begins to form.
There are well over 100 different bacterial species in various dental plaques. This variation in the types of bacteria is influenced by diet, salivary components and bacterial interactions, to name a few. The location of the plaque in the oral cavity, the time of the day, age of the patient and the status of the general oral hygiene of the patient all contribute to the implications and consequences of dental plaque and periodontal disease. Consequently, it is not surprising that plaque is a heterogeneous collection of bacterial communities attached to the tooth providing a vast array of biochemical and physiological consequences. Two major pathological conditions as consequences are periodontal disease and dental caries.
Enzymes as therapeutic agents present unique possibilities. However, some of the early oral pathology research using enzymes was based on the assumption that they would be bactericidal to colonies of organisms found in plaque and therefore would act as "disinfectants". This approach, however, was not fruitful. Recently, it was shown that treatment of buccal epithelial cells with protease altered bacterial adhesion; however, this treatment also distorted the ratios of various bacterial populations. More promising results were obtained when the focus was shifted from bactericidal action to altering plaque formation. These latter results were seen in vitro and in vivo as well as in animal models and human in clinical trials. However, these approaches also fell short of desired therapeutic effectiveness most likely because the required time for an effective action exceeded the retention time of the enzyme in the oral cavity. In short, salivary flow, other fluid and food movement and normal mechanical agitation in the oral cavity reduced the retention time of the enzyme(s). These factors shortened the residence time of the enzymes, resulting in less than desirable clinical efficacy.
When enzymes were tested in vitro, the importance of residence time within the oral cavity was not identified as an important issue. There is no indication that the design of these in vitro studies even identified this important variable. These in vitro systems, that demonstrated activity of enzymes in reducing plaque, did, however, identify other important factors. These other factors included: (1) possibly more than one enzyme may be necessary; (2) greater specific activity of the enzyme may be required; (3) a more appropriate enzyme may be required; or (4) a combination of enzymes may be more effective.
Plaque itself is an extremely complex mixture of various components, namely, macromolecules, living and dead cells (whole bacteria and sloughed epithelial cells from the host), cell fragments and various other contributions of material from both the host and the bacterial flora. The pioneering work on the chemical aspects of plaque focused on the carbohydrate or polysaccharide (PS) backbone of plaque. This was an ideal place to start because the PS backbone not only served as a structural element for the plaque matrix, but it also served as a carbohydrate food-store for the growing colonies of bacteria. Most of the research on PS was centered around determining the properties and structure of glucans; however, there are many other components that form the composition of plaque. In reviewing the scientific literature describing previous dental therapeutic research involving enzymes, certain patterns emerge. Most of the enzyme research to control plaque was conducted under the aegis of caries prevention; however, it is well established that plaque control is a fundamental issue related to both caries prevention and the prevention of periodontal disease. The types of investigations carried out included in vitro examination of bactericidal effects, animal studies and clinical investigations involving human experimentation. Furthermore, most of the clinical studies used a mouthwash as the vehicle to deliver the enzymes, while fewer studies used chewing gum.
U.S. Pat. No. 4,138,476 (Simonson) teaches of plaque dispersing enzymes as oral therapeutic agents by molecular alteration. A glucanohydrolase is combined with a phosphate carrier group such that the enzyme itself has increased affinity for the surfaces of the teeth. The modified glucanohydrolase enzyme covalently crosslinks with the carrier, in the presence of a reacting agent such as ethyl chloroformate, and has an increased binding capacity to hydroxyapatite components of the teeth.
U.S. Pat. No. 5,490,988 (Beggs) relates to the delivery of therapeutic agents to a target site. The patent teaches a highly specific process whereby an antibody fragment is able to bind to a target site through antigen-antibody binding, and provides for a therapeutic agent to be connected onto the antibody fragment through an additional peptide appended to the antibody fragment. The product is thus constituted by the antibody fragment, the peptide and the agent.
Examination of the published clinical protocols for evaluating enzymes shows that there were two reasons why the selected enzymes did not completely exert their desired effects, even though limited clinical efficacy was seen:
a. the enzymes were not modified so that they would be held in the oral cavity for an extended period of time; and PA1 b. the oral rinsing was done for various durations and various selected times during the day without particular attention to dosing just prior to a time of limited oral activity (swallowing, chewing and saliva generation, etc.) like sleeping.
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