Abstract
Replicating viruses have broad applications in biomedicine, notably in cancer virotherapy and in the design of attenuated vaccines; however, uncontrolled virus replication in vulnerable tissues can give pathology and often restricts the use of potent strains. Increased knowledge of tissue-selective microRNA expression now affords the possibility of engineering replicating viruses that are attenuated at the RNA level in sites of potential pathology, but retain wild-type replication activity at sites not expressing the relevant microRNA. To assess the usefulness of this approach for the DNA virus adenovirus, we have engineered a hepatocyte-safe wild-type adenovirus 5 (Ad5), which normally mediates significant toxicity and is potentially lethal in mice. To do this, we have included binding sites for hepatocyte-selective microRNA mir-122 within the 3′ UTR of the E1A transcription cassette. Imaging versions of these viruses, produced by fusing E1A with luciferase, showed that inclusion of mir-122 binding sites caused up to 80-fold decreased hepatic expression of E1A following intravenous delivery to mice. Animals administered a ten-times lethal dose of wild-type Ad5 (5×1010 viral particles/mouse) showed substantial hepatic genome replication and extensive liver pathology, while inclusion of 4 microRNA binding sites decreased replication 50-fold and virtually abrogated liver toxicity. This modified wild-type virus retained full activity within cancer cells and provided a potent, liver-safe oncolytic virus. In addition to providing many potent new viruses for cancer virotherapy, microRNA control of virus replication should provide a new strategy for designing safe attenuated vaccines applied across a broad range of viral diseases.
Highlights
Viruses have a highly successful history as prophylactic vaccines and are being developed for their intrinsic anticancer activities [1]
Attenuated viruses have found important applications in medicine, including their use as vaccines and their experimental development as selective cancer-killing agents, so-called ‘‘virotherapy.’’ Wild-type versions are often most effective in both of these settings; attenuated viruses have usually been developed to decrease the risk of significant viral pathology
We have engineered wild-type adenovirus for recognition by a microRNA expressed in hepatocytes, producing a virus that retains wild-type infection and replication at sites of therapeutic activity but is severely attenuated in hepatocytes, both in vitro and in vivo
Summary
Viruses have a highly successful history as prophylactic vaccines and are being developed for their intrinsic anticancer activities [1]. Several ‘conditionally-replicating’ viruses have been engineered for activation by tumour-associated changes, showing greater potency in cancer cells than in normal cells. These agents are generally attenuated compared to the equivalent wild-type virus even in cancer tissues, and have so far shown little therapeutic activity in clinical trials [4,5]. For both vaccination and cancer virotherapy it would be attractive to produce viruses that show wild-type replication activity at therapeutic sites For both vaccination and cancer virotherapy it would be attractive to produce viruses that show wild-type replication activity at therapeutic sites (eg. within tumours or at sites of antigen presentation) but are attenuated at sites of potential pathology
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