skip to main content


Title: MHC‐restricted Ag85B‐specific CD8 + T cells are enhanced by recombinant BCG prime and DNA boost immunization in mice
Abstract

Despite efforts to develop effective treatments and vaccines,Mycobacterium tuberculosis(Mtb), particularly pulmonaryMtb, continues to provide major health challenges worldwide. To improve immunization against the persistent health challenge ofMtbinfection, we have studied the CD8+T cell response to Bacillus Calmette‐Guérin (BCG) and recombinant BCG (rBCG) in mice. Here, we generated CD8+T cells with an rBCG‐based vaccine encoding the Ag85B protein ofM. kansasii, termed rBCG‐Mkan85B, followed by boosting with plasmid DNA expressing the Ag85B gene (DNA‐Mkan85B). We identified two MHC‐I (H2‐Kd)‐restricted epitopes that induce cross‐reactive responses toMtband other related mycobacteria in both BALB/c (H2d) and CB6F1 (H2b/d) mice. The H2‐Kd‐restricted peptide epitopes elicited polyfunctional CD8+T cell responses that were also highly cross‐reactive with those of other proteins of the Ag85 complex. Tetramer staining indicated that the two H2‐Kd‐restricted epitopes elicit distinct CD8+T cell populations, a result explained by the X‐ray structure of the two peptide/H2‐Kdcomplexes. These results suggest that rBCG‐Mkan85B vector‐based immunization and DNA‐Mkan85B boost may enhance CD8+T cell response toMtb, and might help to overcome the limited effectiveness of the current BCG in eliciting tuberculosis immunity.

 
more » « less
PAR ID:
10457409
Author(s) / Creator(s):
 ;  ;  ;  ;  ;  ;  ;  
Publisher / Repository:
Wiley Blackwell (John Wiley & Sons)
Date Published:
Journal Name:
European Journal of Immunology
Volume:
49
Issue:
9
ISSN:
0014-2980
Page Range / eLocation ID:
p. 1399-1414
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. Abstract

    Peptide nanofibers are useful for many biological applications, including immunotherapy, tissue engineering, and drug delivery. The robust lengthwise assembly of these peptides into nanofibers is typically difficult to control, resulting in polydisperse fiber lengths and an incomplete understanding of how nanofiber length affects biological responses. Here, rationally designed capping peptides control the length of helical peptide nanofibers with unique precision. These designed peptides bind the tips of elongated nanofibers to shorten and narrow their length distributions. Demonstrating their use as immunotherapies, capped nanofibers are preferentially cross‐presented by dendritic cells compared to uncapped nanofibers. Due to increased cross‐presentation, these capped nanofibers trigger stronger CD8+T‐cell responses in mice than uncapped nanofibers. This strategy illustrates a means for controlling the length of supramolecular peptide nanofibers to modulate their immunogenicity in the context of immunotherapies.

     
    more » « less
  2. Tuberculosis (TB) is the world’s deadliest infectious disease, with over 1.5 million deaths and 10 million new cases reported anually. The causative organismMycobacterium tuberculosis(Mtb) can take nearly 40 d to culture, a required step to determine the pathogen’s antibiotic susceptibility. Both rapid identification and rapid antibiotic susceptibility testing of Mtb are essential for effective patient treatment and combating antimicrobial resistance. Here, we demonstrate a rapid, culture-free, and antibiotic incubation-free drug susceptibility test for TB using Raman spectroscopy and machine learning. We collect few-to-single-cell Raman spectra from over 25,000 cells of the Mtb complex strain Bacillus Calmette-Guérin (BCG) resistant to one of the four mainstay anti-TB drugs, isoniazid, rifampicin, moxifloxacin, and amikacin, as well as a pan-susceptible wildtype strain. By training a neural network on this data, we classify the antibiotic resistance profile of each strain, both on dried samples and on patient sputum samples. On dried samples, we achieve >98% resistant versus susceptible classification accuracy across all five BCG strains. In patient sputum samples, we achieve ~79% average classification accuracy. We develop a feature recognition algorithm in order to verify that our machine learning model is using biologically relevant spectral features to assess the resistance profiles of our mycobacterial strains. Finally, we demonstrate how this approach can be deployed in resource-limited settings by developing a low-cost, portable Raman microscope that costs <$5,000. We show how this instrument and our machine learning model enable combined microscopy and spectroscopy for accurate few-to-single-cell drug susceptibility testing of BCG.

     
    more » « less
  3. Introduction

    Immunotherapies have shown great promise, but are not effective for all tumors types and are effective in less than 3% of patients with pancreatic ductal adenocarcinomas (PDAC). To make an immune treatment that is effective for more cancer patients and those with PDAC specifically, we genetically engineered Salmonella to deliver exogenous antigens directly into the cytoplasm of tumor cells. We hypothesized that intracellular delivery of an exogenous immunization antigen would activate antigen-specific CD8 T cells and reduce tumors in immunized mice.

    Methods

    To test this hypothesis, we administered intracellular delivering (ID) Salmonella that deliver ovalbumin as a model antigen into tumor-bearing, ovalbumin-vaccinated mice. ID Salmonella delivers antigens by autonomously lysing in cells after the induction of cell invasion.

    Results

    We showed that the delivered ovalbumin disperses throughout the cytoplasm of cells in culture and in tumors. This delivery into the cytoplasm is essential for antigen cross-presentation. We showed that co-culture of ovalbumin-recipient cancer cells with ovalbumin-specific CD8 T cells triggered a cytotoxic T cell response. After the adoptive transfer of OT-I CD8 T cells, intracellular delivery of ovalbumin reduced tumor growth and eliminated tumors. This effect was dependent on the presence of the ovalbumin-specific T cells. Following vaccination with the exogenous antigen in mice, intracellular delivery of the antigen cleared 43% of established KPC pancreatic tumors, increased survival, and prevented tumor re-implantation.

    Discussion

    This response in the immunosuppressive KPC model demonstrates the potential to treat tumors that do not respond to checkpoint inhibitors, and the response to re-challenge indicates that new immunity was established against intrinsic tumor antigens. In the clinic, ID Salmonella could be used to deliver a protein antigen from a childhood immunization to refocus pre-existing T cell immunity against tumors. As an off-the-shelf immunotherapy, this bacterial system has the potential to be effective in a broad range of cancer patients.

     
    more » « less
  4. Abstract

    Traditional bolus vaccines often fail to sustain robust adaptive immune responses, typically requiring multiple booster shots for optimal efficacy. Additionally, these provide few opportunities to control the resulting subclasses of antibodies produced, which can mediate effector functions relevant to distinct disease settings. Here, it is found that three scaffold‐based vaccines, fabricated from poly(lactide‐co‐glycolide) (PLG), mesoporous silica rods, and alginate cryogels, induce robust, long‐term antibody responses to a model peptide antigen gonadotropin‐releasing hormone with single‐shot immunization. Compared to a bolus vaccine, PLG vaccines prolong germinal center formation and T follicular helper cell responses. Altering the presentation and release of the adjuvant (cytosine‐guanosine oligodeoxynucleotide, CpG) tunes the resulting IgG subclasses. Further, PLG vaccines elicit strong humoral responses against disease‐associated antigens HER2 peptide and pathogenicE. coli, protecting mice againstE. colichallenge more effectively than a bolus vaccine. Scaffold‐based vaccines may thus enable potent, durable and versatile humoral immune responses against disease.

     
    more » « less
  5. Abstract

    Widespread vaccination is essential to global health. Significant barriers exist to improving vaccine coverage in lower‐ and middle‐income countries, including the costly requirements for cold‐chain distribution and trained medical personnel to administer the vaccines. A heat‐stable and highly porous tablet vaccine that can be administered sublingually via simple dissolution under the tongue is described. SIMPL tablet vaccines (Supramolecular IMmunization with Peptides subLingually) are produced by freeze‐drying a mixture of self‐assembling peptide‐polymer nanofibers, sugars, and adjuvant. Sublingual immunization with SIMPL tablets raises antibody responses against both a model epitope from ovalbumin and a clinically relevant epitope fromMycobacterium tuberculosis. Further, sublingual antibody responses are not diminished after heating the tablets for 1 week at 45 °C, in contrast to a more conventional carrier vaccine (KLH). This approach directly addresses the need for a heat‐stable and easily deliverable vaccine to improve equity in global vaccine coverage.

     
    more » « less