HER3-targeted protein chimera forms endosomolytic capsomeres and self-assembles into stealth nucleocapsids for systemic tumor homing of RNA interference in vivo

Abstract

RNA interference represents a potent intervention for cancer treatment but requires a robust delivery agent for transporting gene-modulating molecules, such as small interfering RNAs (siRNAs). Although numerous molecular approaches for siRNA delivery are adequate in vitro, delivery to therapeutic targets in vivo is limited by payload integrity, cell targeting, efficient cell uptake, and membrane penetration. We constructed nonviral biomaterials to transport small nucleic acids to cell targets, including tumor cells, on the basis of the self-assembling and cell-penetrating activities of the adenovirus capsid penton base. Our recombinant penton base chimera contains polypeptide domains designed for noncovalent assembly with anionic molecules and tumor homing. Here, structural modeling, molecular dynamics simulations, and functional assays suggest that it forms pentameric units resembling viral capsomeres that assemble into larger capsid-like structures when combined with siRNA cargo. Pentamerization forms a barrel lined with charged residues mediating pH-responsive dissociation and exposing masked domains, providing insight on the endosomolytic mechanism. The therapeutic impact was examined on tumors expressing high levels of HER3/ErbB3 that are resistant to clinical inhibitors. Our findings suggest that our construct may utilize ligand mimicry to avoid host attack and target the siRNA to HER3+ tumors by forming multivalent capsid-like structures.

Systemic ligand-mimicking bioparticles cross the blood-brain barrier and reduce growth of intracranial triple-negative breast cancer using the human epidermal growth factor receptor 3 (HER3) to mediate both routes

“…Through ligand mimicry, HPK bioparticles evade immuno-recognition by anti-adenovirus antiserum and exhibit no detectable stimulation of neutralizing antibodies in contrast to whole adenovirus in immune-competent mice. 44 In the present study we demonstrate that HPK can form nucleic acid-containing bioparticles that not only invade HER3+ tumors but also distribute to the brain in contrast to cargo lacking neuregulin-directed delivery. We subsequently found that HER3 is prominently expressed on both mouse and human brain endothelium and associates with systemic bioparticle transfer into the brain parenchyma.…”

“…The mixtures were subjected to ultrafiltration using 100K molecular-weight-cutoff membranes to isolate assembled particles from unassembled components. The dsODN concentration in particles was quantified by spectrophotometric absorbance at 260 nm after heparin-mediated release of nucleic acids as previously described 44 . Doxorubin concentration was determined using absorbance at 480 nm and Beer's Law using Extinction Coefficient of Dox = 8030 M-1 cm-1.…”

“…We have bioengineered a tumor-invading protein, HPK (previously known as HerPBK10 41 ), that can home in on breast tumors resisting trastuzumab (Tz) and other ErbB receptor family inhibitors through its high avidity targeting of HER3. 42 Unlike antibodies, HPK uses the receptor engagement of the natural HER3 ligand, heregulin (or neuregulin) 43 to exploit a ligand-induced route of cell entry. HPK also takes advantage of the capsid-forming and membrane penetration functions of the adenovirus-derived penton base protein to encapsulate and deposit macromolecular cargo into HER3-expressing cells.…”

“…HPK also takes advantage of the capsid-forming and membrane penetration functions of the adenovirus-derived penton base protein to encapsulate and deposit macromolecular cargo into HER3-expressing cells. 44 Cargo encapsulation forms biologically-based particles or bioparticles that use neuregulin mimicry to breach HER3+ tumor cells while reducing HER3 activation by wild-type neuregulins 44 . Through ligand mimicry, HPK bioparticles evade immuno-recognition by anti-adenovirus antiserum and exhibit no detectable stimulation of neutralizing antibodies in contrast to whole adenovirus in immune-competent mice.…”

The Adenovirus Dodecahedron: Beyond the Platonic Story

“…In a similar work using the non-dodecamerizing HAdV5 penton base fused to a decalysine (called 3PO), it has been clearly shown that addition of protamine protected DNA from serum nucleases [54]. Such system has been further explored by adding a heregulin ligand to bind with high affinity Her2/3 or Her2/4 overexpressed on certain aggressive breast cancers [55] and a recent work reported the delivery of RNAi in HER3+ tumors in vivo [56].…”