The Development of a COVID19 Oral Vaccine Consisting of Bacillus Subtilis Spores

• Sponsor: DreamTec Research Limited (Industry)
• Overall Status: Completed
• CT.gov ID: NCT05057923
• Collaborator: (none)
• Enrollment: 9
• Location: 1
• Arms: 2
• Actual Duration (Months): 3.7
• Patients Per Site Per Month: 2.4

Study Details

Study Description

Brief Summary

This is a study trial to assess the effectiveness of the immune response stimulated by the genetically engineered Bacillus subtilis which express and display Spike protein of the SARS-COV2 on the spore coat.

Condition or Disease	Intervention/Treatment	Phase
COVID-19 Pneumonia	Biological: Bacillus subtilis	N/A

Detailed Description

Bacillus subtilis is regarded as safe organism by The Food and Drug Administration and it is presented in most food sources. Preliminary experiments have shown that the genetically engineered Bacillus subtilis can express and display receptor binding domain of spike protein of the SARS-COV2 on its spore coat, thus successfully induce the secretion of cytokines of human cells in vitro. Previous experiments also successfully demonstrated that a increased detection of neutralizing IgG and igM levels in mice after oral administrated with the Bacillus subtilis. This suggests that the transgenic spores of Bacillus subtilis have successfully activated the immune system, producing high-affinity neutralizing antibodies and memory B cells. Furthermore, no adverse effects were shown in all the mices. The engineered Bacillus subtilis will be further studied in a human trials through oral administration to test its safety and the immune effect resulted in human bodys.

Study Design

• Study Type: Interventional
• Actual Enrollment: 9 participants
• Allocation: Non-Randomized
• Intervention Model: Parallel Assignment
• Intervention Model Description: two parallel experimental groups were designed. 1. the vaccinated volunteers and 2. unvaccinated groups.two parallel experimental groups were designed.the vaccinated volunteers and 2. unvaccinated groups.
• Masking: None (Open Label)
• Primary Purpose: Basic Science
• Official Title: The Development of a COVID19 Oral Vaccine Consisting of Bacillus Subtilis Spores Expressing and Displaying the Receptor Binding Domain of Spike Protein of SARS-COV2
• Actual Study Start Date: Jul 10, 2021
• Actual Primary Completion Date: Oct 20, 2021
• Actual Study Completion Date: Nov 1, 2021

Arms and Interventions

Arm	Intervention/Treatment
Experimental: generation of neutralizing antibody for unvaccinated participants participants received vaccine 1 capsule of 1×10^10 CFU of B. subtilis spore at day 0, 14, and 28 respectively.	Biological: Bacillus subtilis; Bacillus subtilis, a harmless intestinal commensal, has earned in recent years, great reputation as a vaccine production host and delivery vector with advantages such as low cost, safe for human consumption and straightforward administration. The technology team has succeeded engineering Bacillus subtilis with spore coat proteins resembling the proteins of the nucleus and spikes of coronal virus. This product could have a vaccine like activity within the intestinal environment.
Experimental: neutralizing antibody booster for vaccinated participants participants after 4-month vaccinated with Sinovac received 1 capsule of 1×10^11 CFU of B. subtilis spore	Biological: Bacillus subtilis; Bacillus subtilis, a harmless intestinal commensal, has earned in recent years, great reputation as a vaccine production host and delivery vector with advantages such as low cost, safe for human consumption and straightforward administration. The technology team has succeeded engineering Bacillus subtilis with spore coat proteins resembling the proteins of the nucleus and spikes of coronal virus. This product could have a vaccine like activity within the intestinal environment.

Outcome Measures

Primary Outcome Measures

1. Serum Neutralizing Receptor Binding Domain IgG Antibody Concentration [Day 0, 27, 42 post oral administration]

   Concentration of neutralizing IgG antibody against receptor binding domain of spike protein in SARS-COV2

Secondary Outcome Measures

1. Lentirival Pseudovirus Neutralization Assay (Wild Type of SARS-CoV2) [Day 0, 27, 42 post oral administration]

   The ability of neutralization against SARS-CoV-2 was tested by an in vitro pseudo-virus neutralization assay. The lentivirus carrying a GFP gene was pseudotyped with the spike protein from a wild type of SARS-CoV-2. The pseudoviruses were then pre-incubated with serially diluted serum samples from orally vaccinated volunteers before being added to A549 lung carcinoma cells expressing human ACE2 and human TMPRSS2. The percentage of infection rate was measured with a fluorescent plate reader by counting GFP-positive cells. The results were fitted with a non-linear regression model. The dilution of the serum sample resulted in a 50% reduction of infection rate is designated as EC50. The results were presented as "NA" when the serum samples failed to neutralize pseudovirus infection.

2. Lentirival Pseudovirus Neutralization Assay (D614G SARS-COV2 Variant) [Day 0, 27, 42 post oral administration]

   The ability of neutralization against SARS-CoV-2 was tested by an in vitro pseudo-virus neutralization assay. The lentivirus carrying a GFP gene was pseudotyped with the spike protein from a D614G variant of SARS-CoV-2. The pseudoviruses were then pre-incubated with serially diluted serum samples from orally vaccinated volunteers before being added to A549 lung carcinoma cells expressing human ACE2 and human TMPRSS2. The percentage of infection rate was measured with a fluorescent plate reader by counting GFP-positive cells. The results were fitted with a non-linear regression model. The dilution of the serum sample resulted in a 50% reduction of infection rate is designated as EC50. The results were presented as "NA" when the serum samples failed to neutralize pseudovirus infection.

Eligibility Criteria

Criteria

• Ages Eligible for Study: 25 Years and Older
• Sexes Eligible for Study: All
• Accepts Healthy Volunteers: Yes

Inclusion Criteria:

• healthy

• age over 25 years

• the outcome of the following examinations should be clinically insignificant: medical and surgical history (hypo-, hypertension, allergy, other diseases, major surgery, micturition, defecation, sleep, illness within the last 4 weeks prior to the start of the trial);

• participant vaccinated with Sinovac over 4 months

• anti-SARS CoV 2 neutralizing antibody is negative in serum.

Exclusion Criteria:

• pregnant women

• history of COVID-19 infection or showing COVID-19 infection symptoms

• having had contact to people with known COVID-19 infection in the last 14 days

• having fever (> 37.4oC in the last 24 hours), dry cough or feeling tired and having aches and pains, nasal congestion, runny nose, sore throat and diarrhea.

• positive real time RT-PCR COVID-19 test.

• persons with autoimmune diseases

• allergic diathesis or any clinically significant allergic disease (i.e. asthma)

• any condition that might impair the immune response

• recent or current immunosuppressive medication

• any other vaccine application 30 days before the first dose

Contacts and Locations

Locations

	Site	City	State	Country	Postal Code
1	Zentrogene Bioscience Laboratory Ltd	Hong Kong		Hong Kong

Sponsors and Collaborators

• DreamTec Research Limited

Investigators

• Principal Investigator: WAI YEUNG KWONG, PhD, DreamTec Research Limited
• Study Director: Chun Chau Sung, PhD, DreamTec Research Limited

Study Documents (Full-Text)

• Study Protocol, Statistical Analysis Plan, and Informed Consent Form - Mar 15, 2021

More Information

Publications

None provided.

• Responsible Party: DreamTec Research Limited
• ClinicalTrials.gov Identifier: NCT05057923
• Other Study ID Numbers: PRP/008/21FX
• First Posted: Sep 27, 2021
• Last Update Posted: Dec 23, 2021
• Last Verified: Dec 1, 2021

• Studies a U.S. FDA-regulated Drug Product: No
• Studies a U.S. FDA-regulated Device Product: No
• Keywords provided by DreamTec Research Limited: Covid-19; Vaccine Reaction; Bacillus subtilis
• Additional relevant MeSH terms: COVID-19

Study Results

Participant Flow

Recruitment Details	Participants voluntary enrolled or recruitted by Genfortune Pharmaceuticals Limited
Pre-assignment Detail	6 unvaccinated participants were recruited and their level of serum neutralizing antibody against spike protein of SARS-CoV2 were lower than 0.003 ug/ml.

Arm/Group Title	Generation of Neutralizing Antibody for Unvaccinated Participants
Arm/Group Description	participants received vaccine 1 capsule of 1×10^10 CFU of B. subtilis spore at day 0, 14, and 28 respectively. Bacillus subtilis: Bacillus subtilis, a harmless intestinal commensal, has earned in recent years, great reputation as a vaccine production host and delivery vector with advantages such as low cost, safe for human consumption and straightforward administration. The technology team has succeeded engineering Bacillus subtilis with spore coat proteins resembling the proteins of the nucleus and spikes of coronal virus. This product could have a vaccine like activity within the intestinal environment.
Period Title: Overall Study
STARTED	6
COMPLETED	6
NOT COMPLETED	0

Baseline Characteristics

Arm/Group Title	Generation of Neutralizing Antibody for Unvaccinated Participants
Arm/Group Description	participants received vaccine 1 capsule of 1×10^10 CFU of B. subtilis spore at day 0, 14, and 28 respectively. Bacillus subtilis: Bacillus subtilis, a harmless intestinal commensal, has earned in recent years, great reputation as a vaccine production host and delivery vector with advantages such as low cost, safe for human consumption and straightforward administration. The technology team has succeeded engineering Bacillus subtilis with spore coat proteins resembling the proteins of the nucleus and spikes of coronal virus. This product could have a vaccine like activity within the intestinal environment.
Overall Participants	6
Age (Count of Participants)
<=18 years	0 0%
Between 18 and 65 years	4 66.7%
>=65 years	2 33.3%
Sex: Female, Male (Count of Participants)
Female	3 50%
Male	3 50%
Race (NIH/OMB) (Count of Participants)
American Indian or Alaska Native	0 0%
Asian	6 100%
Native Hawaiian or Other Pacific Islander	0 0%
Black or African American	0 0%
White	0 0%
More than one race	0 0%
Unknown or Not Reported	0 0%
Region of Enrollment (participants) [Number]
East Asia	6 100%

Outcome Measures

1. Primary Outcome

Title	Serum Neutralizing Receptor Binding Domain IgG Antibody Concentration
Description	Concentration of neutralizing IgG antibody against receptor binding domain of spike protein in SARS-COV2
Time Frame	Day 0, 27, 42 post oral administration

Outcome Measure Data

Analysis Population Description

Participants were orally administered 5x10^7 spores/kg person of recombinant B. subtilis spores mixed with sodium alginate in enteric coated capsules at day 1, 14, and 28. Furthermore, 5 mL of blood samples were drawn at day 0, 27, 42, and the serum samples were analyzed. The titer of neutralizing antibodies was determined with a CLIA-based assay.

Arm/Group Title	Generation of Neutralizing Antibody for Unvaccinated Participants
Arm/Group Description	participants received vaccine 1 capsule of 1×10^10 CFU of B. subtilis spore at day 0, 14, and 28 respectively. Bacillus subtilis: Bacillus subtilis, a harmless intestinal commensal, has earned in recent years, great reputation as a vaccine production host and delivery vector with advantages such as low cost, safe for human consumption and straightforward administration. The technology team has succeeded engineering Bacillus subtilis with spore coat proteins resembling the proteins of the nucleus and spikes of coronal virus. This product could have a vaccine like activity within the intestinal environment.
Measure Participants	6
Day 0	0.2 (0.1)
Day 27	9.3 (5.4)
Day 42	35.8 (8.8)

2. Secondary Outcome

Title	Lentirival Pseudovirus Neutralization Assay (Wild Type of SARS-CoV2)
Description	The ability of neutralization against SARS-CoV-2 was tested by an in vitro pseudo-virus neutralization assay. The lentivirus carrying a GFP gene was pseudotyped with the spike protein from a wild type of SARS-CoV-2. The pseudoviruses were then pre-incubated with serially diluted serum samples from orally vaccinated volunteers before being added to A549 lung carcinoma cells expressing human ACE2 and human TMPRSS2. The percentage of infection rate was measured with a fluorescent plate reader by counting GFP-positive cells. The results were fitted with a non-linear regression model. The dilution of the serum sample resulted in a 50% reduction of infection rate is designated as EC50. The results were presented as "NA" when the serum samples failed to neutralize pseudovirus infection.
Time Frame	Day 0, 27, 42 post oral administration

Outcome Measure Data

Analysis Population Description

[Not Specified]

Arm/Group Title	Generation of Neutralizing Antibody for Unvaccinated Participants
Arm/Group Description	participants received vaccine 1 capsule of 1×10^10 CFU of B. subtilis spore at day 0, 14, and 28 respectively. Bacillus subtilis: Bacillus subtilis, a harmless intestinal commensal, has earned in recent years, great reputation as a vaccine production host and delivery vector with advantages such as low cost, safe for human consumption and straightforward administration. The technology team has succeeded engineering Bacillus subtilis with spore coat proteins resembling the proteins of the nucleus and spikes of coronal virus. This product could have a vaccine like activity within the intestinal environment.
Measure Participants	6
EC50 of Day 27 Serum diltuion in Pseudovirus inhibition assay	NA (NA)
EC50 of Day 0 Serum diltuion in Pseudovirus inhibition assay	NA (NA)
EC50 of Day 42 Serum diltuion in Pseudovirus inhibition assay	76.33 (38.7)

3. Secondary Outcome

Title	Lentirival Pseudovirus Neutralization Assay (D614G SARS-COV2 Variant)
Description	The ability of neutralization against SARS-CoV-2 was tested by an in vitro pseudo-virus neutralization assay. The lentivirus carrying a GFP gene was pseudotyped with the spike protein from a D614G variant of SARS-CoV-2. The pseudoviruses were then pre-incubated with serially diluted serum samples from orally vaccinated volunteers before being added to A549 lung carcinoma cells expressing human ACE2 and human TMPRSS2. The percentage of infection rate was measured with a fluorescent plate reader by counting GFP-positive cells. The results were fitted with a non-linear regression model. The dilution of the serum sample resulted in a 50% reduction of infection rate is designated as EC50. The results were presented as "NA" when the serum samples failed to neutralize pseudovirus infection.
Time Frame	Day 0, 27, 42 post oral administration

Outcome Measure Data

Analysis Population Description

[Not Specified]

Arm/Group Title	Generation of Neutralizing Antibody for Unvaccinated Participants
Arm/Group Description	participants received vaccine 1 capsule of 1×10^10 CFU of B. subtilis spore at day 0, 14, and 28 respectively. Bacillus subtilis: Bacillus subtilis, a harmless intestinal commensal, has earned in recent years, great reputation as a vaccine production host and delivery vector with advantages such as low cost, safe for human consumption and straightforward administration. The technology team has succeeded engineering Bacillus subtilis with spore coat proteins resembling the proteins of the nucleus and spikes of coronal virus. This product could have a vaccine like activity within the intestinal environment.
Measure Participants	6
EC50 of Day 27 Serum diltuion in Pseudovirus inhibition assay	NA (NA)
EC50 of Day 0 Serum diltuion in Pseudovirus inhibition assay	NA (NA)
EC50 of Day 42 Serum diltuion in Pseudovirus inhibition assay	82.4 (29.2)

Adverse Events

Time Frame	Adverse event (AE) data were collected from Day 0 (post-vaccination) up to Day 30 post-final vaccination.
Adverse Event Reporting Description	All AEs were presented.
Arm/Group Title	Generation of Neutralizing Antibody for Unvaccinated Participants
Arm/Group Description	participants received vaccine 1 capsule of 1×10^10 CFU of B. subtilis spore at day 0, 14, and 28 respectively. Bacillus subtilis: Bacillus subtilis, a harmless intestinal commensal, has earned in recent years, great reputation as a vaccine production host and delivery vector with advantages such as low cost, safe for human consumption and straightforward administration. The technology team has succeeded engineering Bacillus subtilis with spore coat proteins resembling the proteins of the nucleus and spikes of coronal virus. This product could have a vaccine like activity within the intestinal environment.
All Cause Mortality
	Generation of Neutralizing Antibody for Unvaccinated Participants
	Affected / at Risk (%)	# Events
Total	0/6 (0%)
Serious Adverse Events
	Generation of Neutralizing Antibody for Unvaccinated Participants
	Affected / at Risk (%)	# Events
Total	0/6 (0%)
Other (Not Including Serious) Adverse Events
	Generation of Neutralizing Antibody for Unvaccinated Participants
	Affected / at Risk (%)	# Events
Total	0/6 (0%)

Limitations/Caveats

[Not Specified]

More Information

Certain Agreements

All Principal Investigators ARE employed by the organization sponsoring the study.

There is NOT an agreement between Principal Investigators and the Sponsor (or its agents) that restricts the PI's rights to discuss or publish trial results after the trial is completed.