© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim wileyonlinelibrary.com 1 COMMUNICATION M13 Bacteriophage and Adeno-Associated Virus Hybrid for Novel Tissue Engineering Material with Gene Delivery Functions So Young Yoo,* Hyo-Eon Jin, Dong Shin Choi, Masae Kobayashi, Yohan Farouz, Sky Wang, and Seung-Wuk Lee* Dr. S. Y. Yoo, Dr. H.-E. Jin, D. S. Choi, M. Kobayashi, Y. Farouz, S. Wang, Prof. S.-W. Lee Department of Bioengineering University of California, Berkeley Physical Biosciences Division Lawrence Berkeley National Laboratory Berkeley Nanoscience and Nanoengineering Institute Berkeley, CA 94720, USA E-mail: yoosy@pusan.ac.kr; leesw@berkeley.edu Dr. S. Y. Yoo BIO-IT Foundry Technology Institute Pusan National University Busan 609-735, and Research Institute for Convergence of Biomedical Science and Technology Yangsan 626-770, Republic of Korea Y. Farouz Biology Department Ecole Polytechnique Route de Saclay 91128, Palaiseau, Cedex, France DOI: 10.1002/adhm.201500179 infect even by these virus-based approaches. [10] (2) Viruses often cause host immune responses that eliminate the infecting virus and its side effects on the surrounding cells. Gene delivery vec- tors must be able to escape the body's natural surveillance sys- tems, otherwise a resulting adverse immune reaction can cause serious illness or even death. [11] (3) Random insertion of viral genome into host genome can cause unexpected mutations that can give rise to abnormal cell formation such as cancer. [12] Therefore, selective, safe, stable, and predictable gene delivery is still challenging in terms of developing targeted gene delivery. Recent advances in the phage biotechnology provide remark- able pathways to develop novel biomaterials. Phages possess many desirable features that make them attractive as versatile gene delivery materials. Phage have little harmful effects and are easily removed from the body through lysosomal degrada- tion processes, causing few known side-effects. [13] Phage can be modified to display functional peptide motifs on their minor (pIII, pIX) and major (pVIII) coat proteins. [14–18] Large quanti- ties of identical phage building blocks can be easily prepared through bacterial amplification. Due to their long-filamentous shape (aspect ratio: 130), phage can self-assemble into nanofi- brous tissue-like matrix structures. Recently, we engineered various biochemical cues (i.e., RGD (Arg-Gly-Asp), IKVAV (Ile- Lys-Val-Ala-Val), and DGEA (Asp-Gly-Glu-Ala)) on major coat proteins (pVIII) of the phage and developed self-assembled tissue engineering materials for regulating cellular behaviors of desired cells. [15,16,19–22] Using this phage-based tissue matrix system, we demonstrated that engineered phages could regulate various cellular behaviors such as proliferation and differentia- tion. [14–16,19,20,23] Here, we developed novel tissue engineering materials with gene delivery functions through the hybridiza- tion of M13 bacteriophage (phage) and AAV. We engineered the M13 phage with RGD peptide on the major coat proteins. We then fused its gene with AAV gene with eukaryotic invading functional gene, ITR. The resulting M13–AAV hybrid phage exhibited enhanced internalization into target cells as compared to minor coat engineered phage with RGD or wild-type phage. The resulting hybrid phage formed nanofibrous matrices that could support the cellular growth and deliver desired gene information into the target tissues. Our novel M13–AAV hybrid phage matrices can provide selective, stable, and safe gene delivery ( Figure 1). We believe that it can be also developed as a topical therapeutic tissue patch in the future. In order to construct a novel phage tissue engineering mate- rial with gene delivery function ( Figure 2), we constructed the hybrid phage (M13 RGD8 –AAV GFP ) carrying both RGD major coat Reprogramming of cellular functions through gene delivery is critical in treating cellular/tissue abnormalities as well as in studying the mechanism of cellular behavior of many cells. [1] Many researchers have utilized nanosized natural [2] or syn- thetic materials such as surfactants, [3] peptide or protein nano- spheres, [4] and polymersomes for gene delivery. [5] Recently, virus-based gene delivery vehicles, such as retrovirus or adeno- virus, are considered as promising delivery vehicles for transfer of genetic information into target cells. [6] Among them, adeno- associated virus (AAV) has been largely adapted for therapeutic gene transfer due to its innocuousness and high resistance to extreme conditions. [7] The AAV is a nonpathogenic virus with a linear single-stranded DNA genome that contains eukaryotic gene integrating functional genes, named inverted terminal repeats (ITRs; 145 bp in length for AAV2), [8] at each end of the AAV viral genome terminus. The ITR possesses a T-shaped hairpin structure with self-complementary guanine-cyto- sine rich sequences; the gene enables integration of the viral genome into as well as rescue from a specific section of the host genome (19th chromosome in humans). [9] Despite the develop- ment of various viral gene delivery vehicles, their therapeutic application still remains challenging due to several reasons. (1) Viruses can hardly reach their desired targets. Viruses have lim- ited ranges of the type of host cells they can infect, although adenoviruses and AAV are able to infect a relatively broad range of cells efficiently. Still, some cell types are unmanageable to Adv. Healthcare Mater. 2015, DOI: 10.1002/adhm.201500179 www.advhealthmat.de www.MaterialsViews.com