Interferon resistance promotes oncolysis by influenza virus NS1‐deletion mutants

Abstract

NS1 protein of influenza virus is a virulence factor that counteracts Type I interferon (IFN)- Key words: conditionally-replicating-virus; STAT1; SCIDKnowledge of the pathogenesis of viral diseases and the ability to manipulate specific regions of viral genomes permit the construction of conditionally replicating viruses that are attenuated in normal cells but retain their ability to lyse tumor cells. [1][2][3] We show that the NS1 protein of influenza A virus is a virulence factor that counteracts the interferon (IFN)-mediated antiviral cellular response. 4 As a consequence, an influenza virus that lacks a functional NS1 protein due to an almost total deletion of the NS1 open reading frame (delNS1 virus) fails to replicate in normal cells and is apathogenic for wild-type mice. 5 The delNS1 virus, however, grows to titers similar to wild-type virus in dsRNA-activated kinase (PKR) knockout mice. 6 The conditionally replicating phenotype of the delNS1 virus in PKR-defective systems could be exploited for virally-induced oncolysis in tumors expressing oncogenic ras. 7 This observation is based on the fact that oncogenic ras inhibits PKR activation. 8 For this reason, melanoma cells became permissive for productive delNS1 virus replication upon transfection and expression of oncogenic N-ras. Moreover, delNS1 virus treatment of subcutaneous N-ras-expressing melanomas in SCID mice showed that this virus has tumor-ablative potentials in vivo.The delNS1 virus was also shown to replicate effectively in STAT1 knockout mice. 9 STAT1 exists in 2 isoforms, a 91-kD protein (STAT1␣) and a 84-kD splice variant (STAT1␤). 10 Upon activation by Type I IFN signaling STAT1 forms a heterodimer with STAT2 and becomes an essential part of the IFN-receptor induced transcription complex ISGF3. A reduced expression of STAT1 is associated with IFN resistance. 11 Alterations in the IFN-dependent signal cascades, including changes in STAT1 and Type I IFN-receptor molecules, have been described to occur frequently in malignantly transformed cells. For example, several melanoma and lymphoma cell lines contain no or reduced levels of STAT1. 12,13 In addition, leukemia cell lines were shown to be defective in IFN genes. 14 We hypothesized, therefore, that IFN resistance is a common tumor characteristic, which may allow delNS1 virus-mediated oncolysis.We analyzed the growth of delNS1 virus in IFN-resistant tumor cell lines of various histological origins. We compared the oncolytic effect of the delNS1 virus with a second virus, in which the NS1 was only deleted to the N-terminal 99 amino acids (NS1-99). In mice, the latter virus has intermediate attenuation properties ranging between delNS1 and wild-type viruses. 15 The NS1-99 virus gave us the possibility to investigate whether the attenuation level of NS1-deletion viruses influences the efficiency of oncolysis in vivo. The in vitro growth of the NS1-deletion mutants inversely correlated with the IFN resistance of the assayed cell lines. Both delNS1 and NS1-99 viruses induced a tumor-ablative ef...

An engineered avian-origin influenza A virus for pancreatic ductal adenocarcinoma virotherapy

“…To limit viral replication in IFN-competent healthy cells but not in IFN-impaired tumour cells, the H7N3 NS1 gene was mutated to produce a protein of only 77 aa (NS1-77), resulting in the complete loss of the ED (Hale et al, 2008). A second NS1-77 truncated IAV was also generated using the genetic backbone of the well-studied laboratory human strain H1N1 A/Puerto Rico/8/34 (PR8), previously investigated for use as an OV against other types of cancers (Bergmann et al, 2001;Muster et al, 2004;Sturlan et al, 2010).…”

“…Indeed, no recombinant IAV of any origin has been appraised so far for PDA treatment, and the relative small number of previous investigations concerning the use of IAV for virotherapy were focused on the human strain H1N1 A/Puerto Rico/8/34 (Bergmann et al, 2001;Muster et al, 2004;Sturlan et al, 2010;Wolschek et al, 2011). Importantly, the choice of a particular viral isolate is likely to affect its efficacy as an OV because IAVs differ in their ability to counteract host IFN response (Geiss et al, 2002;Hayman et al, 2006;Kochs et al, 2007), induce apoptosis (Kasloff et al, 2014) and in their sensitivity to host ISGs (Dittmann et al, 2008).…”

“…However, in cancer cells with deficient IFN responses, such mutants have been shown to replicate and cause cell death (Bergmann et al, 2001;Muster et al, 2004). A recombinant influenza virus with NS1 truncation rather than complete deletion was less attenuated and shown to be more effective as an oncolytic treatment in xenograft mouse model of human melanoma (Muster et al, 2004).…”

Anticancer Gene Transfer for Cancer Gene Therapy

“…Essentially specific pathways are required for the activation of replication cycle and due to extensive de-regulation of such pathways in cancer cells. These attempts to alter the viral tropism using pathway dependencies for viral replication have been utilized in the first generation of pre-clinical or clinically approved oncolytic viruses like Adenovirus (Ad): ONYX-015, dl922-947, Herpes Simplex Virus (HSV): G207, R3616, R1716, bM24-TE, Newcastle Disease Virus (NDV), Influenza Virus (IFA), Vesicular Stomatitis Virus (VSV) (Bischoff et al, 1996;O'Shea et al, 2004;Heise et al, 2000;Fueyo et al, 2000;Mineta et al, 1995;Cinatl et al, 2004;Reichard et al, 1992;Kuroda et al, 2006;Stojdl et al, 2003;Muster et al, 2004). The second approach for manipulating viral tropism and selective replication is to engineer viruses with genes responsible for replication controlled by tumor or tissue specific promoters.…”

“…The latter is a host defense pathway, which halts protein synthesis of virally-infected cells and induces their death. Since activated Ras leads to inactivation of PKR, oncolytics like HSV-1 or Influenza virus A (IFA) are able to conditionally replicate only in cancer cells using this common defect of protein synthesis inhibition in tumors (Smith et al, 2006;Veerapong et al, 2007;Bergmann et al, 2001;Muster et al, 2004). Ras pathway has a central role in tumor initiation, progression and sensitivity to treatment.…”