HAMLET: Functional Properties and Therapeutic Potential

James Ho CS; Anna Rydström; Maria Trulsson; Johannes Bålfors; Petter Storm; Manoj Puthia; Aftab Nadeem; Catharina Svanborg

Disclosures

Future Oncol. 2012;8(10):1301-1313. 

In This Article

HAMLET-like Complexes

Following the discovery of HAMLET, several HAMLET-like complexes have been produced. The properties of HAMLET-like complexes using α-lactalbumin derived from different species suggested that the conversion of α-lactalbumin to its tumoricidal form tolerates a certain degree of sequence variation. Bovine, equine, porcine and caprine α-lactalbumin differed greatly in the efficiency of complex formation, although the complexes with oleic acid showed similar tumoricidal activities.[66] Lysozyme, which is the closest structural homolog of α-lactalbumin, also forms cytotoxic complexes with oleic acid called ELOA.[67] In contrast with conventional noncalcium-binding c-type lysozyme, equine lysozyme binds calcium and exhibits enzymatic activity. Similar to α-lactalbumin, equine lysozyme forms a range of partially folded states under destabilizing conditions and also populates various kinetic intermediates, while retaining its native-like core conformation.

A few significant differences between ELOA and HAMLET should be discussed. ELOA exists as an oligomer, binding as many as 48 oleic acid molecules in a single complex, compared with four to eight oleic acids bound to the HAMLET monomers. This 'lipid overload' is likely to change the interaction with tumor cell membranes from a programmed cell death response to cell lysis, as high levels of oleic acid are cytotolytic.[68–71]

Other HAMLET-like complexes have been produced by alternative methods, including mixing procedures under acidic[72] or alkaline[42] pH conditions or by heat denaturation.[73] By using proteolytic fragments of bovine α-lactalbumin mixed with oleic acid, Tolin et al. suggested that the protein component mainly acts as a binding partner for the fatty acid.[40] However, the peptide preparations contained two- to three-fold more lipid compared with HAMLET, and target analysis in tumor cells has not yet been performed. The α-isoform of pike parvalbumin or bovine β-lactoglobulin[43] were both shown to form complexes with oleic acid, suggesting that proteins other than α-lactalbumin may act as binding partners for oleic acid. This was confirmed for β-lactoglobulin.[74] These complexes are cytotoxic for different cells lines, including human lung carcinoma cells A549,[72] laryngeal carcinoma (HEp-2) cells[70] and human monocytic cells (U937).[43] Mechanistic characterizations of the implicated cell death pathways are needed to define if the activity of the complexes is associated with the cytotoxicity of oleic acid.

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