Role of HAMLET and metabolism in treatment and pathogenesis of pneumococci

Antimicrobial resistance (AMR) is one of the principle public health problems in the 21st century, threatening the available treatment strategies for bacterial infections. Here, we present a human protein-lipid complex, HAMLET (human alpha-lactalbumin made lethal to tumor cells) purified from human milk as a potential therapeutic agent which has both tumoricidal and bactericidal activity. HAMLET’s anti-bacterial activity is selective, against respiratory pathogens with highest activity seen in Streptococcus pneumoniae (the pneumococcus). HAMLET-induced bacterial death was shown to require membrane depolarization and rupture by a sodium-dependent influx of calcium, interference with glycolysis and activation of kinases. In this thesis, to understand the role of HAMLET as a future therapeutic agent, we studied HAMLET-induced targets and pathways involved in pneumococcal death and host immunomodulatory effects, which can provide us with information about future potential bacterial targets and alternative treatment strategies. Additionally, to understand pneumococcal pathogenesis, we studied metabolism and biofilm formation in pneumococci with different niche-associated sugars (like galactose). In paper I, we observed that HAMLET results in inhibition of glycolysis and energy production in the cells. In paper II, we studied the interaction between HAMLET’s bacterial targets and observed that pneumococcal targets of HAMLET are either directly or indirectly related. In paper III, we observed that HAMLET induces immunomodulatory effects resulting in functional changes of monocyte-derived macrophages and dendritic cells. In paper IV, we observed that pneumococci grow slower and are less metabolically active in both planktonic and biofilm bacteria in the presence of galactose compared to glucose. Further, we show that galactose-grown bacteria disperse (spread) less in response to febrile temperature compared to glucose-grown bacteria. Overall, the results from this thesis suggest that HAMLET has dual anti-bacterial roles: first by directly killing bacteria and second by stimulating immune responses to eliminate bacteria. Additionally, in the presence of galactose pneumococcal growth and metabolism is slow, suggesting a role in bacterial pathogenesis (in vitro).