**1. Background**

Virus-like particles (VLPs) are empty viral shells derived from the expression, in a suitable host cell, of viral structural proteins such as capsid or coat proteins. Over-expression of these proteins allows them to spontaneously self-assemble into VLPs (Figure 1). VLPs can also be derived from envelope proteins following over-expression of envelope proteins together with other structural proteins such as the pre-membrane proteins, capsid proteins, or over-expression of the envelope protein with Gag protein [1]. VLPs derived from the latter are known as enveloped VLPs. Thus, VLPs are empty shells which consist of one or more types of multimeric coat or envelope proteins arranged geometrically into dense, repetitive (multivalent) arrays [2–4]. They are morphologically and structurally similar to viruses from which the coat proteins are derived, except for the fact that they lack the viral genome. VLPs are highly immunogenic, even at small doses of antigens (as low as 500 ng) [5]. The following features have been credited for making VLPs highly immunogenic:

(i) Coat proteins that form the capsid are multivalently displayed to the immune system. Multivalent display enhances cross-linking of B-cell receptors on naïve B cells, leading to a stronger B-cell activation, proliferation, and differentiation, secretion of high-affinity antibodies, and the generation of long-lived memory B cells [6–9].


**Figure 1.** The assembly of bacteriophage MS2 coat proteins into VLPs (virus-like particles). The genome of MS2 bacteriophage (top image) with the coat protein (light blue). Cloning of the coat protein into a bacterial expression vector and expression of the protein in a suitable bacterial host gives rise to coat proteins (capsomers). The capsomeres form pentamers and hexamers; twelve pentamers and 20 hexamers spontaneously self-assemble to form a VLP (composed of 180 capsomeres). RdRP stands for RNA dependent RNA polymerase.

The aforementioned features, in addition to the fact that VLPs are noninfectious and are safe, have made VLPs attractive biological agents for vaccine design and development. A number of VLP-based prophylactic vaccines have been approved by the Food and Drug Administration to protect against human papillomaviruses (HPVs) and hepatitis B virus (HBV) infections. Two VLP-based HBV vaccines are credited for reducing incidences of HBV-related hepatocellular carcinomas and mortalities, worldwide, with protection lasting for up to 30 years in some individuals [2,15]. On the other hand, three VLP-based HPV vaccines (Gadarsil-9, Cervarix, and Gardasil-4 (discontinued in the US)) have been approved within the last decade to protect against HPV infections.
