Development of a broad spectrum glycoconjugate vaccine to prevent wound and disseminated infections with Klebsiella pneumoniae and Pseudomonas aeruginosa

Abstract

Klebsiella pneumoniae (KP) and Pseudomonas aeruginosa (PA) are important human pathogens that are associated with a range of infection types, including wound and disseminated infections. Treatment has been complicated by rising rates of antimicrobial resistance. Immunoprophylactic strategies are not constrained by antimicrobial resistance mechanisms. Vaccines against these organisms would be important public health tools, yet they are not available. KP surface O polysaccharides (OPS) are protective antigens in animal models of infection. Similarly, PA flagellin (Fla), the major subunit of the flagellar filament, is required for virulence and is a target of protective antibodies in animal models. We report herein the development of a combined KP and PA glycoconjugate vaccine comprised of the four most common KP OPS types associated with human infections (O1, O2, O3, O5), chemically linked to the two Fla types of PA (FlaA, FlaB). Conjugation of KP OPS to PA Fla enhanced anti-polysaccharide immune responses and produced a formulation that generated antibody titers to the four KP OPS types and both PA Fla antigens in rabbits. Passive transfer of vaccine-induced rabbit antisera reduced the bacterial burden and protected mice against fatal intravenous KP infection. Mice passively transferred with conjugate-induced antisera were also protected against PA infection after thermal injury with a FlaB-expressing isolate, but not a FlaA isolate. Taken together, these promising preclinical results provide important proof-of-concept for a broad spectrum human vaccine to prevent KP and PA infections.

Neuraminidase 1–mediated desialylation of the mucin 1 ectodomain releases a decoy receptor that protects against Pseudomonas aeruginosa lung infection

“…Pa-derived flagellin was purified and adsorbed with polymyxin B-agarose to remove bacterial LPS, after which less than 0.1 LPS units/g of protein was detected by the Limulus amebocyte lysate assay as described (20). Flagellin was purified from STm strain CVD1925, a derivative of the WT invasive Malian strain I77, as described (79,80). Both Pa and STm flagellins exhibited a single Coomassie Blue-stained protein band following SDS-PAGE.…”

“…Both rFlaA and rFlaB proteins were purified as described (79) and exhibited a single Coomassie Blue-stained protein band following SDS-PAGE. For preparation of mouse polyclonal antisera to rFlaA and rFlaB, 6-to 7-week-old female Crl:CD-1 mice (Charles River Laboratories) were injected in the right gastrocnemius muscle on 0, 14, and 28 days with 2.5 g of rFlaA or rFlaB flagellins, as described (80). Blood was collected by retro-orbital bleeding at 14 days following the final immunization and centrifuged, and the serum stored at Ϫ20°C.…”

A murine model demonstrates capsule-independent adaptive immune protection in survivors of Klebsiella pneumoniae respiratory tract infection

“…ATCC 43816) that are highly and rapidly lethal in mice (Fodah et al, 2014;Hsieh et al, 2013;Lau et al, 2007;Lavender et al, 2004;Lawlor et al, 2005;Lin et al, 2014;Wu et al, 2009), precluding their utility in illuminating natural adaptive immune responses to live K. pneumoniae. Instead, experiments have assessed adaptive immunity elicited by heat-killed organisms and a variety of specific K. pneumoniae immunogenic factors, including outer membrane vesicles, O-antigens, type 3 fimbriae and purified capsule (Chen et al, 2011;Cryz et al, 1985;Lee et al, 2015;Amezcua Vesely et al, 2019;Lavender et al, 2005;Trautmann et al, 2004;Hegerle et al, 2018). Early work in rodents and subsequently humans indicated that immunization with capsule elicits serotype-specific antibody responses (Cryz et al, 1985;Alcantar-Curiel et al, 1993;Cryz et al, 1988Cryz et al, , 1984Cryz et al, , 1986a; however, capsule is not the sole driver of protective immunity during infection (Lee et al, 2015;Lundberg et al, 2013).…”

Impact of vaccines on antimicrobial resistance

“…Both P. aeruginosa and K. pneumoniae account for many nosocomial infections that are increasingly complicated to treat due to antibiotic resistance. A glycoconjugate vaccine against K. pneumoniae and P. aeruginosa infections is also in preclinical development; results so far are promising for development of this broad-spectrum human vaccine (Hegerle et al, 2018). The vaccine formulation includes the four predominant surface O polysaccharides associated with K. pneumoniae infection, conjugated to a protein carrier to enhance immunity.…”

“…Flagellin proteins from P. aeruginosa were selected as carrier proteins in order to produce a vaccine targeting both K. pneumoniae and P. aeruginosa infections. This approach to extend vaccine coverage to other pathogens has only been used in one other vaccine to date, a pneumococcal conjugate vaccine that uses protein D from non-typeable H. influenzae (NTHi) as a carrier (Hegerle et al, 2018).…”