SBIR-STTR Award

Staphylococcus is the most common cause of infections worldwide. The pathogen persists asymptomatically on human skin from which it may spread to ocular tissues and cause corneal infection. When this happens, corneal bacterial keratitis can occur, and if not treated effectively, it may lead to blindness. Multi Drug Resistant (MDR) variants of S. aureus (Sa) are increasingly prevalent and are of major clinical concern, as the arsenal of effective antibiotics available for treatment is becoming depleted. The problem of antibiotic resistance is compounded by the ability of Sa to grow, not only in its familiar planktonc form, but also in the form of compact biofilms against which current antibiotics have limited effect. This creates a need to discover new and effective modalities against MDR pathogens. But it also creates a commercial opportunity, particularly if it is possible to reduce the likelihod that MDR pathogens will become resistant to these modalities. In this phase I STTR proposal, AmebaGone, LLC, in collaboration with the academic partner at the School of Medicine and Public Health of University of Wisconsin-Madison, will carry out proof-of principle experiments to test whether treating bacterial keratitis with Dictyostelid Cells (DC) can mitigate ocular Sa infections. DC are microscopic unicellular professional phagocytes of group Conosa that devour bacteria as their normal food source. DC can engulf and digest bacteria at a prodigious rate; and they exponentially multiply, until the bacterial prey is gone. Only when the supply of the surface exposed bacteria is large and the oxygen is abundant they become multi-cellular and differentiate into dormant spores. Genomic analyses of these organisms suggest that DC were among the earliest multi-cellular organisms to arrive on dry land. Because the sporulation cannot occur when DC feed on submerged bacteria; their evolutionary split from the aquatic amoebic cells (group Lobosa), some of which are pathogenic, had occurred over billion years ago. Humans and animals encounter DC in soil worldwide - with no adverse consequences. It is highly significant to this project that AmebaGone, LLC, has already acquired several Dictyostelid strains and has shown that these organisms can kill planktonic and biofilm-encased S. aureus - MRSA (USA300). These strains are samplings from a large archive of over 3,000 natural isolates of DC from diverse locations. Our aims are: (1) Screen a preselected group of DC for their ability to feed on planktonic and biofilm-encased cells of S. aureus 8325-4, a common pathogen of ocular tissues, (2) Determine the toxicity of DC when applied topically to the mouse cornea and determine the residence time of DC on the cornea following a single application, and, (3) Test the efficacy of DC in treating murine corneal infections with the strain of S. aureus If DC have no adverse effects and are efficacious against S. aureus in the ocular tissues of the mouse, the treatment will be tested in Phase II in a rabbit model of keratitis. The issued patents, the company owns, are to protect commercialization of DC as therapeutics in medicine and agriculture.

Public Health Relevance Statement:


Public Health Relevance:
Staphylococcus aureus (Sa) persists harmlessly on human skin from which it may spread to the eye and cause an infection that may lead to blindness, if not treated effectively. The pathogen is evolving resistance to almost all available antibiotics tht have been used for infection treatment. The research project we propose will test whether treating infected eyes with microscopic organisms that devour bacteria as their food source is a safe treatment for ocular Sa infections.

Project Terms:
A/J Mouse; Address; Adverse effects; adverse outcome; Agriculture; Animal Experiments; animal resource; Animals; Anti-Bacterial Agents; Antibiotic Resistance; Antibiotics; Archives; Bacteria; Bacterial Infections; Bacterial Model; Bacteriology; base; Biological; Biological Models; Biological Response Modifier Therapy; Blindness; cell killing; Cell Therapy; Cells; Clinical; Collaborations; commercialization; Consultations; Contracts; Cornea; Data; design; Development; Disease; Dose; Eating; efficacy testing; Exhibits; Eye; feeding; Food; Genomics; Genus staphylococcus; Goals; Health; high risk; Human; Immune; Infection; interest; Keratitis; Killings; Laboratories; Lead; Legal patent; Letters; Life; Location; Mann-Whitney U Test; Measures; medical schools; Medicine; methicillin resistant Staphylococcus aureus (organism); Microbial Biofilms; Microscopic; Modality; Modeling; Multi-Drug Resistance; multi-drug resistant pathogen; Mus; Names; novel; Organism; Oryctolagus cuniculus; Oxygen; pathogen; pathogenic bacteria; Phagocytes; Phase; phase 1 study; Polymers; professor; prototype; Pseudomonas aeruginosa; public health medicine (field); Reproduction spores; Research; Research Personnel; Research Project Grants; research study; residence; Resistance; Resistance development; Rodent; Rodent Model; Safety; Sampling; Schedule; Services; Skin; Small Business Technology Transfer Research; Soil; Source; Staging; Staphylococcus aureus; Streptococcus pneumoniae; Surface; Technology; Temperature; Testing; Therapeutic; Therapeutic Effect; Time; TimeLine; Tissues; Topical application; Toxic effect; Universities; Variant; Wisconsin; Work

Bacterial keratitis (BK) is an infection of the cornea that, if left untreated, can cause blindness. Staphylococcus aureus (Sa), Streptococcus pneumoniae (Sp), and Pseudomonas aeruginosa (Pa) are three major causes of BK in North America. Symptoms of BK include pain, redness, inflammation, and opacity of the affected cornea and loss of visual acuity (1). AmebaGone Inc., (AG) partnered with UW-Madison Department of Ophthalmology to develop a novel biocontrol method to treat BK. AG holds issued patents and has expert knowledge related to use of Dictyostelid cells (DC), a benign, soil- dwelling organism known to destroy pathogenic bacteria through the phagocytosis. Issued patents broadly cover the use of DC to eat biofilm-enmeshed and free-living (planktonic) bacteria (“Therapeutic Ameba and Uses Thereof”; USPTO granted patent 8,551,471 and follow up patent US 8,551,671). A patent covering countries of the European Union has been filed and the firm has licensed rights to more than 3,000 DC strains from 3 worldwide collections occupying diverse natural habitats. Benign DC diverged from aquatic amoeba, some of which are pathogenic, over 1 billion years ago. In Phase I work, AG identified 36 Dicty strains capable of efficiently killing (> 4 log10 reduction in bacterial titers) Sa at eye temperature. Of these strains, AG identified 4 DC strains that reproducibly germinated to high levels and developed a gel formulation that increased the maximum recovery time from 1 to 16 hours. Additionally, AG tested safety of these DC strains and found no increase in corneal pathology or discomfort compared to the vehicle control. In the Phase II project AG proposes to: 1) Expand the target of DC from singular action against Sa to additional causative agents of BK- Pa and Sp, 2). Develop effective formulation and assess DC stability therein. 3) Conduct in vivo safety and basic biology testing of DC treatment in the eye, and 4). Quantify efficacy of DC cocktails in treating murine cornea infected with Sa, Pa, or Sp in vivo. The data generated in these studies will position us to prepare an Investigational New Drug (IND) application for the Food and Drug Administration (FDA).

Public Health Relevance Statement:
Narrative Bacterial keratitis (BK) is an infection of the cornea that can cause pain, redness, inflammation, decreased visual acuity and in severe cases blindness. AmebaGone Inc. proposes to develop a prototype biotherapeutic for BK using microorganisms that naturally prey on bacteria - even those enmeshed in biofilms which antibiotics cannot kill. This novel approach has far- reaching implications to treat many more infectious diseases, providing an alternative to conventional antibiotics.

Project Terms:
Affect; Amoeba genus; Animal Experiments; Antibiotic Resistance; Antibiotics; antimicrobial; Asses; Awareness; Bacteria; Bacterial Infections; Benign; Biocontrols; Biological Response Modifier Therapy; Biology; Blindness; cell mediated immune response; Cells; Collection; Communicable Diseases; Consultations; Cornea; Corneal Injury; Country; Data; design; Dose; Eating; European Union; Eye; Eye Infections; follow-up; Formulation; Gel; Genome; Grant; Habitats; Half-Life; healing; Hour; IACUC; Immune response; In Vitro; in vitro testing; in vivo; Infection; Inflammation; Investigational New Drug Application; Keratitis; Killings; Knowledge; Lead; Left; Legal patent; Letters; Measures; Methods; Microbial Biofilms; microorganism; Mus; North America; novel; novel strategies; Ophthalmology; Organism; Pain; pathogen; pathogenic bacteria; Pathogenicity; Pathology; Phagocytosis; Phase; phase 2 study; Positioning Attribute; Predisposition; professor; prototype; Pseudomonas aeruginosa; Recovery; Redness; Research; research and development; residence; Resistance; Rights; Safety; safety testing; Schedule; Services; Soil; Staphylococcus aureus; Streptococcus pneumoniae; success; Symptoms; Technology; Temperature; Testing; Therapeutic; Time; United States Food and Drug Administration; Universities; Validation; Visual Acuity; Wisconsin; Work; Wound Healing