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Introducing the ESAT-6 free IGRA, a companion diagnostic for TB vaccines based on ESAT-6

View Article: PubMed Central - PubMed

ABSTRACT

There is a need for an improved vaccine for tuberculosis. ESAT-6 is a cardinal vaccine antigen with unique properties and is included in several vaccine candidates in development. ESAT-6 is also the core antigen in the IFN-γ release assays (IGRA) used to diagnose latent infection, rendering IGRA tests unspecific after vaccination. This challenge has prompted the development of a companion diagnostic for ESAT-6 based vaccines, an ESAT-6 free IGRA. We screened a panel of seven potential new diagnostic antigens not recognized in BCG vaccinated individuals. Three highly recognized antigens EspC, EspF and Rv2348c were identified and combined with CFP10 in an ESAT-6 free antigen cocktail. The cocktail was prepared in a field-friendly format, lyophilized with heparin in ready-to-use vacutainer tubes. The diagnostic performance of the ESAT-6 free IGRA was determined in a cross-validation study. Compared IGRA, the ESAT-6 free IGRA induced a comparable magnitude of IFN-γ release, and the diagnostic performance was on par with Quantiferon (sensitivity 84% vs 79%; specificity 99% vs 97%). The comparable performance of the ESAT-6 free IGRA to IGRA suggests potential as companion diagnostic for ESAT-6 containing vaccines and as adjunct test for latent infection.

No MeSH data available.


IFN-γ release to QFT antigens, 10 screened Mtb specific antigens and selected regions within antigens.Diluted whole blood from 48 patients with TB (A) 34 Egyptian and 14 from Greenland), 18 Mtb infected controls from Greenland (B) and 56 uninfected controls from Egypt (C) was stimulated 24 hours with overlapping peptides (as described in Table 1) in 200 ul volume. IFN-γ release was determined using in-house ELISA and presented following subtraction of IFN-γ release in an unstimulated control well. Median values are indicated in red.
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f1: IFN-γ release to QFT antigens, 10 screened Mtb specific antigens and selected regions within antigens.Diluted whole blood from 48 patients with TB (A) 34 Egyptian and 14 from Greenland), 18 Mtb infected controls from Greenland (B) and 56 uninfected controls from Egypt (C) was stimulated 24 hours with overlapping peptides (as described in Table 1) in 200 ul volume. IFN-γ release was determined using in-house ELISA and presented following subtraction of IFN-γ release in an unstimulated control well. Median values are indicated in red.

Mentions: IFN-γ release from antigen stimulated whole blood culture in 48 patients with microbiologically or microscopy confirmed pulmonary TB (34 from Egypt and 14 from Greenland), 18 Mtb infected controls (Greenland), and 56 controls with negative IGRA (Egypt) were compared in terms of magnitude of IFN-γ release, and responder frequency (Fig. 1 and Table 2). ESAT-6, CFP10 and EspC were the most frequently recognized (>40% responders of TB and LTBI), Rv2348-B and EspF were less frequently recognized (15–40%) whereas TB7.7, EspJ, EccD1, PE35 and the N-terminal of Rv2348c (Rv2348-A) were infrequently detected. The group of LTBI donors had a wider antigen repertoire compared to the TB patients, whom almost exclusively focused on ESAT-6, CFP10 and EspC. There were no significant differences in the antigen recognition repertoire among TB patients from Greenland and Egypt (data not shown).


Introducing the ESAT-6 free IGRA, a companion diagnostic for TB vaccines based on ESAT-6
IFN-γ release to QFT antigens, 10 screened Mtb specific antigens and selected regions within antigens.Diluted whole blood from 48 patients with TB (A) 34 Egyptian and 14 from Greenland), 18 Mtb infected controls from Greenland (B) and 56 uninfected controls from Egypt (C) was stimulated 24 hours with overlapping peptides (as described in Table 1) in 200 ul volume. IFN-γ release was determined using in-house ELISA and presented following subtraction of IFN-γ release in an unstimulated control well. Median values are indicated in red.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5384086&req=5

f1: IFN-γ release to QFT antigens, 10 screened Mtb specific antigens and selected regions within antigens.Diluted whole blood from 48 patients with TB (A) 34 Egyptian and 14 from Greenland), 18 Mtb infected controls from Greenland (B) and 56 uninfected controls from Egypt (C) was stimulated 24 hours with overlapping peptides (as described in Table 1) in 200 ul volume. IFN-γ release was determined using in-house ELISA and presented following subtraction of IFN-γ release in an unstimulated control well. Median values are indicated in red.
Mentions: IFN-γ release from antigen stimulated whole blood culture in 48 patients with microbiologically or microscopy confirmed pulmonary TB (34 from Egypt and 14 from Greenland), 18 Mtb infected controls (Greenland), and 56 controls with negative IGRA (Egypt) were compared in terms of magnitude of IFN-γ release, and responder frequency (Fig. 1 and Table 2). ESAT-6, CFP10 and EspC were the most frequently recognized (>40% responders of TB and LTBI), Rv2348-B and EspF were less frequently recognized (15–40%) whereas TB7.7, EspJ, EccD1, PE35 and the N-terminal of Rv2348c (Rv2348-A) were infrequently detected. The group of LTBI donors had a wider antigen repertoire compared to the TB patients, whom almost exclusively focused on ESAT-6, CFP10 and EspC. There were no significant differences in the antigen recognition repertoire among TB patients from Greenland and Egypt (data not shown).

View Article: PubMed Central - PubMed

ABSTRACT

There is a need for an improved vaccine for tuberculosis. ESAT-6 is a cardinal vaccine antigen with unique properties and is included in several vaccine candidates in development. ESAT-6 is also the core antigen in the IFN-γ release assays (IGRA) used to diagnose latent infection, rendering IGRA tests unspecific after vaccination. This challenge has prompted the development of a companion diagnostic for ESAT-6 based vaccines, an ESAT-6 free IGRA. We screened a panel of seven potential new diagnostic antigens not recognized in BCG vaccinated individuals. Three highly recognized antigens EspC, EspF and Rv2348c were identified and combined with CFP10 in an ESAT-6 free antigen cocktail. The cocktail was prepared in a field-friendly format, lyophilized with heparin in ready-to-use vacutainer tubes. The diagnostic performance of the ESAT-6 free IGRA was determined in a cross-validation study. Compared IGRA, the ESAT-6 free IGRA induced a comparable magnitude of IFN-γ release, and the diagnostic performance was on par with Quantiferon (sensitivity 84% vs 79%; specificity 99% vs 97%). The comparable performance of the ESAT-6 free IGRA to IGRA suggests potential as companion diagnostic for ESAT-6 containing vaccines and as adjunct test for latent infection.

No MeSH data available.