Studies on the Interaction of Tumor-Derived HD5 Alpha Defensins with Adenoviruses and Implications for Oncolytic Adenovirus Therapy

Abstract

Defensins are small antimicrobial peptides capable of neutralizing human adenovirus (HAdV) in vitro by binding capsid proteins and blocking endosomal escape of virus. In humans, the alpha defensin HD5 is produced by specialized epithelial cells of the gastrointestinal and genito-urinary tracts. Here, we demonstrate, using patient biopsy specimens, that HD5 is also expressed as an active, secreted peptide by epithelial ovarian and lung cancer cells in situ. This finding prompted us to study the role of HD5 in infection and spread of replication-competent, oncolytic HAdV type 3 (HAdV3). HAdV3 produces large amounts of penton-dodecahedra (PtDd), virus-like particles, during replication. We have previously shown that PtDd are involved in opening epithelial junctions, thus facilitating lateral spread of de novo-produced virions. Here, we describe a second function of PtDd, namely, the blocking of HD5. A central tool to prove that viral PtDd neutralize HD5 and support spread of progeny virus was an HAdV3 mutant virus in which formation of PtDd was disabled (mut-Ad3GFP, where GFP is green fluorescent protein). We demonstrated that viral spread of mut-Ad3GFP was blocked by synthetic HD5 whereas that of the wild-type (wt) form (wt-Ad3GFP) was only minimally impacted. In human colon cancer Caco-2 cells, induction of cellular HD5 expression by fibroblast growth factor 9 (FGF9) significantly inhibited viral spread and progeny virus production of mut-Ad3GFP but not of wt-Ad3GFP. Finally, the ectopic expression of HD5 in tumor cells diminished the in vivo oncolytic activity of mut-Ad3GFP but not of wt-Ad3GFP. These data suggest a new mechanism of HAdV3 to overcome innate antiviral host responses. Our study has implications for oncolytic adenovirus therapy. IMPORTANCE Previously, it has been reported that human defensin HD5 inactivates specific human adenoviruses by binding to capsid proteins and blocking endosomal escape of virus. The central new findings described in our manuscript are the following: (i) the discovery of a new mechanism used by human adenovirus serotype 3 to overcome innate antiviral host responses that is based on the capacity of HAdV3 to produce subviral penton-dodecahedral particles that act as decoys for HD5, thus preventing the inactivation of virus progeny produced upon replication; (ii) the demonstration that ectopic HD5 expression in cancer cells decreases the oncolytic efficacy of a serotype 5-based adenovirus vector; and (iii) the demonstration that epithelial ovarian and lung cancers express HD5. The study improves our understanding of how adenoviruses establish infection in epithelial tissues and has implications for cancer therapy with oncolytic adenoviruses.KEYWORDS adenovirus, cancer, defensins, innate immunity

Upregulated Expression of Intestinal Antimicrobial Peptide HD5 Associated with Renal Function in IgA Nephropathy

“…Elevated serum HD6 levels were found to be associated with gender in IgAN patients. The current findings suggested that HD5 rather than HD6 There were some evidences suggesting that HD5 or HD6 persists in serum with very low concentration [36,38,39]. We also detected serum HD5 by ELISA and found a relatively low level of HD5 in peripheral blood both from IgAN patients and normal controls.…”

“…Measurement of Serum and Urine Levels of HD5 and HD6. Serum and urine levels of HD5 and HD6 were measured by the human HD5 and HD6 ELISA kit (USCN Life Science, Wuhan, China) according to the manufacturer's instructions [36]. Serum was diluted 1 : 10 in the appropriate buffer and transferred to microtitre plates for incubation procedures.…”

Oncolytic Viruses—Interaction of Virus and Tumor Cells in the Battle to Eliminate Cancer

“…This inhibition presents a barrier to effective viral spread and successful oncolytic therapy. However, type 3 Ad has evolved to manufacture a decoy capsid which sequesters HD5 and renders the virus able to escape endosomes ( 62 ). This finding suggests that the mechanism used by this strain of Ad can be mimicked in order to improve efficacy of existing or potential oncolytic Ad therapies.…”

Synergistic antitumor effect of the combination of a dual cancer‑specific oncolytic adenovirus and cisplatin on lung cancer cells

“…However, an effective cancer treatment strategy also needs a safe delivery vehicle that can express apoptin protein persistently and deliver it effectively. The human adenovirus serotype 5 (Ad5) is one of such vector that has been used widely to treat a variety of tumors ( 13 – 16 ). In addition, satisfactory curative effects in the treatment of tumors have been demonstrated by a combination of a recombinant Ad5 and cisplatin ( 17 , 18 ).…”