ABSTRACT
Purpose of Study: Previous Osteopathic manipulative treatment (OMT) research has shown evidence of increased lymphatic movement resulting in increased leukocyte and cytokine flow. One study even showed increased antibody titer response in patients when used in conjunction with the Hepatitis B vaccine. Given previous studies, our group conducted a year-long longitudinal randomized controlled clinical trial to evaluate the ability of OMT to improve the COVID-19 vaccine immune response. Methods Used: Subjects were randomized into either the control arm or OMT intervention arm. OMT consisted of myofascial release of the thoracic inlet, pectoral traction, diaphragm release with MFR, splenic pump, and thoracic pump applied the day of and the day following each vaccination session. All subjects in each group received the Pfizer mRNA COVID-19 vaccine. All subjects had blood drawn on day 0 (1st vaccine), day 7, day 21 (2nd vaccine), days 28, 90, 210, and 365. Anti-spike IgG immunoglobulin titers (AS IgG) were measured at all time points for all subjects. Side effects, adverse events, and medication usage in response to the vaccines or OMT was documented. Breakthrough cases with symptomology and medication usage was documented for both groups. The study was approved by the WesternU IRB committee, protocol #FB21/IRB026. Summary of Results: Data for 91 subjects were analyzed with 41 male (45.1%) and 50 female (54.9%). Age distribution was comparable between the two groups. Side effects and medication usage reported by the subjects was similar between groups (p>0.1). AS IgG measured at baseline distinguished between previously infected individuals and those naive to COVID-19, regardless of OMT treatment. For all time points measured, the average AS IgG in subjects trended higher in OMT group than control group. Two-way ANOVA analysis showed statistical significance at 1 week after 2nd injection (p<0.001) in the COVID-19 naive population. 13 symptomatic breakthrough infections were reported in the control group and 12 in the OMT group. Length of symptoms were reported as 8.36 +/- 4.60 days (control) and 4.62 +/- 2.60 days (OMT) (p<0.05). Length of medication usage was 3.64 +/- 3.58 days (control) and 1.23 +/- 1.24 days (OMT) (p<0.1). Conclusion(s): Both groups had comparable side effects after COVID-19 vaccination with no adverse events linked to OMT, indicating that OMT is a safe adjunct that can be used with COVID-19 vaccination. The data showed an enhanced immune response by OMT, as evidenced by increased levels of AS IgG in previously naive subjects. Although both control and OMT groups had similar rates of symptomatic breakthrough infection, the OMT data shows reduced length of symptoms and medication duration in this population when compared to control breakthrough infections. This study is underpowered for statistical significance at each time point and future vaccination studies should recruit more patients to confirm the trends seen here.
ABSTRACT
Purpose of Study: Antibiotic resistance remains one of the largest healthcare and public health challenges. Several studies have documented that the spread of antibiotic resistant bacteria in nosocomial settings has been exacerbated worldwide due to increased rates of hospitalization and intubation in the wake of the COVID-19 pandemic. One way to address antibiotic resistance is to identify novel compounds that inhibit essential microbial processes. Two-component regulatory systems are important mediators of signal transduction that allow bacteria to communicate with and respond to changes in their environment. The WalRK system is a two-component system that is conserved and essential for viability in many Gram-positive human pathogens. We hypothesize that a ligand that specifically binds with the DNA-interaction surface of the WalR protein can lead to cell death and can serve as a lead compound for future drug development efforts. Methods Used: We describe the development process of an assay to identify WalR binding compounds. In silico molecular dynamics docking approaches were utilized to identify potential WalR binding compounds from virtual compound libraries. To assess their WalR-binding capacity in vitro, overexpression strains for several WalR recombinant constructs were engineered and protein constructs were purified to homogenicity. Isothermal titration calorimetry (ITC) is a technique that measures heat release or absorption when two molecules interact. A MicroCal PEAQ ITC instrument was utilized to develop a WalR-binding assay. Summary of Results: WalR is a two-domain protein featuring a regulatory and a DNA-binding domain. Two constructs, a truncated DNA-binding domain and a full-length protein construct proved soluble, and pure quantities necessary to conduct ITC measurements could be successfully obtained (12 mg full-length protein and 23 mg truncated protein). These proteins were amenable to ITC experiments. We found that experiments were best run with at least a two-fold increase of ligand concentration to protein concentration supplied in identical buffer conditions over nineteen injections. We are currently assessing the binding affinities of our in silico hit compounds. Conclusion(s): Our results show that ITC enables the detailed, rapid, and reproducible characterization of the binding relationship between the DNA-binding domain of the WalR protein and any potential ligands. The protocol discussed herein will enable further drug discovery studies on the WalR response regulator protein to identify and characterize inhibitors, providing insight towards the development of novel antimicrobial compound.