title: Preclinical study and phase I clinical safety evaluation of recombinant Mycobacterium tuberculosis ESAT6 protein.
abstract: BACKGROUND
  To investigate the ability of rESAT6 to identify different mycobacteria-sensitized guinea pigs and its safety in preclinical and phase I clinical study.
  
  
  METHODS
  Guinea pigs were sensitized with different Mycobacteria. After sensitization, all animals were intradermally injected with rESAT6 and either PPD or PPD-B. At 24 h after the injection, the erythema of the injection sites were measured using a double-blind method. For the preclinical safety study, different doses of rESAT6 and BSA were given 3 times intramuscularly to guinea pigs. On day 14 after the final immunization, the guinea pigs were intravenously injected with the same reagents in the hind legs and the allergic reactions were observed. A single-center, randomized, open phase I clinical trial was employed. The skin test was conducted in 32 healthy volunteers aged 19-65 years with 0.1 µg, 0.5 µg, and 1 µg rESAT6. Physical examination and laboratory tests were performed before and after the skin test and adverse reactions were monitored. The volunteers' local and systemic adverse reactions and adverse events were recorded for 7 days.
  
  
  RESULTS
  Positive PPD or PPD-B skin tests were observed in all Mycobacteria-sensitized guinea pigs; the diameters of erythema were all >10 mm. The rESAT6 protein induced a positive skin test result in the guinea pigs sensitized with MTB, M. bovis, M. africanum and M. kansasii; the diameters of erythema were 14.7±2.0, 9.3±3.8, 18.7±2.4, and 14.8±4.2 mm, respectively. A negative skin test result was detected in BCG-vaccinated and other NTM-sensitized guinea pigs. The rESAT6 caused no allergic symptoms, but many allergic reactions, such as cough, dyspnea, and even death, were observed in the guinea pigs who were administered BSA. During the phase I clinical trial, no adverse reactions were found in the 0.1 µg rESAT6 group, but in the 0.5 µg rESAT6 group 2 volunteers reported pain and 1 reported itching, and in the 1 µg rESAT6 group there was 1 case of pain, 1 case of itching, and 1 case of blister. No other local or systemic adverse reactions or events were reported.
  
  
  CONCLUSIONS
  The rESAT6 can differentiate effectively among MTB infection, BCG vaccination, and NTM infection and is safe in healthy volunteers.
authors.affiliation: National Institutes for Food and Drug Control, Beijing, PR China.
authors.forename: Wei-Xin
authors.initials: WX
authors.lastname: Du
authors.forename: Bao-Wen
authors.initials: BW
authors.lastname: Chen
authors.forename: Jin-Biao
authors.initials: JB
authors.lastname: Lu
authors.forename: Meng-Qiu
authors.initials: MQ
authors.lastname: Gao
authors.forename: Xiao-Bing
authors.initials: XB
authors.lastname: Shen
authors.forename: Lei
authors.initials: L
authors.lastname: Yang
authors.forename: Cheng
authors.initials: C
authors.lastname: Su
authors.forename: Guo-Zhi
authors.initials: GZ
authors.lastname: Wang
authors.forename: Qing-feng
authors.initials: QF
authors.lastname: Sun
authors.forename: Miao
authors.initials: M
authors.lastname: Xu
mesh_terms: D000328:Adult; D000368:Aged; D000818:Animals; D000942:Antigens, Bacterial / Q000008:administration & dosage / Q000009:adverse effects* / Q000276:immunology*; D001426:Bacterial Proteins / Q000008:administration & dosage / Q000009:adverse effects* / Q000276:immunology*; D004306:Dose-Response Relationship, Immunologic; D004353:Drug Evaluation, Preclinical; D005260:Female; D006168:Guinea Pigs; D006801:Humans; D007114:Immunization; D008297:Male; D008875:Middle Aged; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000009:adverse effects* / Q000276:immunology*; D014373:Tuberculin / Q000276:immunology; D014374:Tuberculin Test; D055815:Young Adult
chemical_list: D000942:Antigens, Bacterial; D001426:Bacterial Proteins; C093276:ESAT-6 protein, Mycobacterium tuberculosis; D011994:Recombinant Proteins; D014373:Tuberculin