Trimethyl Chitosan Nanoparticles Encapsulated Protective Antigen Protects the Mice Against Anthrax

Abstract

Anthrax is an era old deadly disease against which there are only two currently available licensed vaccines named anthrax vaccine adsorbed and precipitated (AVP). Though they can provide a protective immunity, their multiple side-effects owing to their ill-defined composition and presence of toxic proteins (LF and EF) of Bacillus anthracis, the causative organism of anthrax, in the vaccine formulation makes their widespread use objectionable. Hence, an anthrax vaccine that contains well-defined and controlled components would be highly desirable. In this context, we have evaluated the potential of various vaccine formulations comprising of protective antigen (PA) encapsulated trimethyl-chitosan nanoparticles (TMC-PA) in conjunction with either CpG-C ODN 2395 (CpG) or Poly I:C. Each formulation was administered via three different routes, viz., subcutaneous (SC), intramuscular (IM), and intraperitoneal in female BALB/c mice. Irrespective of the route of immunization, CpG or Poly I:C adjuvanted TMC-PA nanoparticles induced a significantly higher humoral response (total serum IgG and its isotypes viz., IgG1, IgG2a, and IgG2b), compared to their CpG or Poly I:C PA counterparts. This clearly demonstrates the synergistic behavior of CpG and Poly I:C with TMC nanoparticles. The adjuvant potential of TMC nanoparticles could be observed in all the three routes as the TMC-PA nanoparticles by themselves induced IgG titers (1–1.5 × 105) significantly higher than both CpG PA and Poly I:C PA groups (2–8 × 104). The effect of formulations on T-helper (Th) cell development was assessed by quantifying the Th1-dependant (TNF-α, IFN-γ, and IL-2), Th2-dependant (IL-4, IL-6, and IL-10), and Th17-type (IL-17A) cytokines. Adjuvanation with CpG and Poly I:C, the TMC-PA nanoparticles triggered a Th1 skewed immune response, as suggested by an increase in the levels of total IgG2a along with IFN-γ cytokine production. Interestingly, the TMC-PA group showed a Th2-biased immune response. Upon challenge with the B. anthracis Ames strain, CpG and Poly I:C adjuvanted TMC-PA nanoparticles immunized via the SC and IM routes showed the highest protective efficacy of ~83%. Altogether, the results suggest that CpG or Poly I:C adjuvanted, PA-loaded TMC nanoparticles could be used as an effective, non-toxic, second generation subunit-vaccine candidate against anthrax.

<p>Chitosan And N, N, N-Trimethyl Chitosan Nanoparticle Encapsulation Of <em>Ocimum Gratissimum</em> Essential Oil: Optimised Synthesis, In Vitro Release And Bioactivity</p>

“…40 Malik et al also reported the smooth and irregular shape of antigenloaded TMCNPs. 22 The sizes of the OGEO-TMCNPs conformed to those reported by Sotelo-Boyás et al 10 .…”

“…At pH 3.0, cumulative EO release for OGEO-TMCNPs and OGEO-CSNPs were 77.35% and 70.39%, respectively. Malik et al reported a 30% protein release at pH 7.4 after 6 h with an increase to 55% within the first 24 h and reaching a steady state release after 48 h. 22 OGEO-TMCNPs exhibited higher EO release at all pH conditions compared to the OGEO-CSNPs. This is the result of the increased EO loading capacity of TMC nanoparticles with the influence of the methyl group.…”

“…[15][16][17][18][19] TMC has shown a potential use as an adjuvant and in gene delivery due to its ability to form polyplexes with DNA. 17,22 The aim of this study is to propose a novel approach for the optimised synthesis of Ocimum gratissimum essential oil (OGEO)-loaded nanoparticles with distinct physiochemical properties. Also, to the best of our knowledge, the bioactivities and in vitro release kinetic pattern of OGEO-encapsulated CS and TMC nanoparticles at varying physiological pH conditions (pH 3 and pH 7.4) have not been elucidated in the literature to date.…”

<p>Crystalline and Amorphous Preparation of Aluminum Hydroxide Nanoparticles Enhances Protective Antigen Domain 4 Specific Immunogenicity and Provides Protection Against Anthrax</p>

“…dosing schedule to elicit correct protective antibodies against the disease. The development of improved vaccine candidates against anthrax has been pursued for a long time with studies like polylactic-co-glycolic acid (PLGA) encapsulated D4 nanoparticles, 6 and PLGA-dendron nanoparticle-based PA-DNA vaccine, 16 dual-function viral nanoparticle, 17 CpG-Ficoll nanoparticle adjuvant encapsulating PA, 18 and Tri-methyl chitosan nanoparticles containing PA, 19 where the authors have shown induction of a better immune response and disease protection as compared to the traditional anthrax vaccines. Moreover, alum, being a Th2-type adjuvant, fails to clear the intracellular pathogens, thus rendering the vaccines ineffective in certain diseases like tuberculosis.…”

Development of Enzyme Linked Immunosorbent Assay for humoral immuneresponse and infection monitoring of anthrax

“…Although the existing vaccines such as AVA are safe and effective, there has been a surge in the need for anthrax vaccines due to the increased terroristic attacks in the recent decades. Furthermore, the prevalence of such bacteria among humans has highlighted the necessity of research and developing methods of producing subunit vaccines (Malik et al, 2018). PA is one of the most important antigens in the anthrax which is used in the design of vaccines and several commercial diagnostic kits.…”