Oncolytic virus

Abstract

The present invention relates to genetic constructs, as well as expression cassettes and oncolytic viruses comprising the genetic constructs, and to their use in methods for treating, preventing or ameliorating cancer. The genetic constructs, expression cassettes and oncolytic viruses are particularly, although not exclusively, useful for treating bladder cancer, such as non-muscle invasive bladder cancer.

Description

[0001]Oncolytic Virus The present invention relates to genetic constructs, as well as expression cassettes and oncolytic viruses comprising the genetic constructs, and to their use in methods for treating, preventing or ameliorating cancer. The genetic constructs, expression cassettes and oncolytic viruses are particularly, although not exclusively, useful for treating bladder cancer, such as non-muscle invasive bladder cancer. Non-muscle invasive bladder cancer (NMIBC) is a highly prevalent and significant health problem, with lifelong risk of recurrence. In the UK, there are around 10,000 new cases of NMIBC per year. Surgery is a standard treatment but is accompanied by high recurrence rate and tumour progression. Immunotherapy with live tuberculosis vaccine, bacillus Calmette-Guerin (BCG), into the bladder has been a standard treatment for 40 years, and is instilled after surgery in high-risk disease (intermediate and high-grade cancer). BCG works as an immunotherapy in part through its activation of receptors which are key to the initiation of host immune defence mechanisms, so called Toll-like receptor ligands (TLRs). Whilst it reduces the risk of recurrence, it does so at the high cost of local and systemic toxicity, which severely limits its use. As such, around a third of patients are unresponsive to BCG treatment, whilst another third experience severe side effects and must cease treatment. There is, therefore, an urgent unmet need for alternative treatments for a significant proportion of bladder cancer patients (up to 60%), who do not respond to, or cannot tolerate, BCG. In particular, there is the need for a new localised therapy capable of exploiting the immunomodulatory effects of BCG with improved efficacy, but without the toxic side effects. Accordingly, in a first aspect of the invention, there is provided a genetic construct comprising a first coding sequence, which encodes a TLR agonist, and a second coding sequence, which encodes a cytokine. The inventors have created a novel cancer-killing virus that is surprisingly effective for treating BCG non-responder models (and ultimately patients). Advantageously, as illustrated in Figures 2 and 3, the virus retains potent tumour cytotoxicity with enhanced immunogenicity by the inclusion of a TLR agonist (e.g. a novel bacterial protein) together with a cytokine (e.g. IL-15), which promotes the proliferation of NK cells and cytotoxic functions of CD8 T cells. Preferably, the genetic construct comprises an expression cassette, one embodiment of which is shown in Figure 1. Accordingly, in a second aspect of the invention, there is provided an expression cassette comprising the genetic construct of the first aspect. As can be seen in Figure 1, the construct comprises the first nucleotide sequence encoding the TLR agonist (e.g. a novel bacterial protein) and the second nucleotide sequence encoding the cytokine (e.g. IL-15). It will be appreciated, however, that other TLR agonists and cytokines may be used, as discussed herein. In one embodiment, the TLR agonist may be a bacterial TLR agonist. Alternatively, the TLR agonist may be a synthetic TLR agonist. Preferably, the TLR agonist is a TLR2 agonist, a TLR4 agonist, or a TLR5 agonist. More preferably, the TLR agonist is a TLR2 agonist or a TLR5 agonist. The TLR agonist may be a lipoprotein or a lipopeptide. Examples of TLR agonists may be found in Kaczanowska S, Joseph AM, Davila E. TLR agonists: our best frenemy in cancer immunotherapy. J Leukoc Biol.2013 Jun;93(6):847-63. doi: 10.1189 / jlb.1012501. Epub 2013 Mar 8. PMID: 23475577; PMCID: PMC3656332. In one preferred embodiment, the TLR2 agonist may be selected from a group consisting of: LpqH, LprA, LprG, LpqT, PhoS1, MPB83, MPT83, LAM, AraLAM, LM, PIM2 / 6, TDM, HSP70, MymA, PE_PGRS33, EsxL, PPE18, PPE26, PPE32, PPE57, Lrp, GPLs, PILAM, PPE60, OmpA and OmpC. Preferably, the TLR2 agonist is PPE60 from Mycobacterium tuberculosis. In another preferred embodiment, the TLR2 agonist is an outer membrane protein, OmpA and / or OmpC, from Shigella dysenteriae. In one preferred embodiment, the TLR4 agonist is FimH. The TLR4 agonist, FimH, may be from E. coli and / or Salmonella enterica serovar Typhimurium. In one preferred embodiment, the TLR5 agonist may be selected from a group consisting of: FliC, CagL, CagY, FllA and FlaB. CagL and CagY may be from Helicobacter pylori. FllA may be from E. coli. FlaB may be from Vibrio vulnificus. In one preferred embodiment, the TLR5 agonist is FliC. The TLR5 agonist, FliC, may be from E. coli and / or Burkholderia pseudomallei (BP). Alternatively, the TLR5 agonist, FliC, may be from other Burkholderia species or a Salmonella strain, including but not limited to: Salmonella typhimurium, Salmonella enteritidis, Salmonella choleraesuis, Salmonella gallinarum, Salmonella pullorum, and Salmonella enterica and subspecies and serovars thereof. As described in the Examples, when screening TLR ligands, the inventors identified that the bacterial flagellin protein, FliC_BP, is a particularly effective candidate. In particular, the inventors identified that FliC_BP can trigger TLR5- and TLR2-mediated signalling pathways, suggesting either the unique property of FliC_BP to bind to the TLR2 receptor or the occurrence of crosstalk between TLR5 (the primary receptor for flagellin) and TLR2. As such, this TLR5 / TLR2 ligand offers the opportunity to enhance antitumour immune responses seen with oncolytic viruses with an innate bacterial response. Most preferably, therefore, the TLR agonist is a TLR5 agonist. Most preferably, the TLR5 agonist is FliC_BP. FliC_BP is a bacterial flagellin protein from Burkholderia pseudomallei. FliC_BP (GenBank: ABA48561.1) may comprise an amino acid sequence of SEQ ID No: 1, which is provided herein, as follows: MVYRFSARAIRVHARSRRLAGRRTARRSHARFVRIQRWLTERSTQKIRAESRPAPSSFFGVRRNPAQGGR QRTSASARANSGIDGLIQLNLEIFMLGINSNINSLVAQQNLNGSQGALSQAITRLSSGKRINSAADDAAG LAIATRMQTQINGLNQGVSNANDGVSILQTASSGLTSLTNSLQRIRQLAVQASNGPLSASDASALQQEVA QQISEVNRIASQTNYNGKNILDGSAGTLSFQVGANVGQTVSVDLTQSMSAAKIGGGMVQTGQTLGTIKVA IDSSGAAWSSGSTGQETTQINVVSDGKGGFTFTDQNNQALSSTAVTAVFGSSTAGTGTAASPSFQTLALS TSATSALSATDQANATAMVAQINAVNKPQTVSNLDISTQTGAYQAMVSIDNALATVNNLQATLGAAQNRF TAIATTQQAGSNNLAQAQSQIQSADFAQETANLSRAQVLQQAGISVLAQANSLPQQVLKLLQ [SEQ ID No: 1] Accordingly, in a preferred embodiment, FliC_BP comprises or consists of an amino acid sequence substantially as set out in SEQ ID No: 1, or a fragment or variant thereof. In one embodiment, FliC_BP may be encoded by the nucleic acid sequence of SEQ ID No: 2, which is provided herein as follows: ATGGTTTACCGCTTTTCCGCGCGGGCGATCCGTGTCCACGCCCGTTCGCGCCGCCTTGCGGGCCGCCGGA CGGCGCGCCGTTCGCATGCGCGATTCGTGCGCATCCAACGTTGGCTCACCGAACGATCGACACAAAAAAT TCGGGCCGAATCTCGCCCTGCCCCCTCAAGTTTTTTTGGCGTACGCCGAAATCCTGCACAAGGCGGCCGG CAAAGAACGAGTGCTTCCGCCCGCGCCAATAGTGGCATTGATGGGCTTATACAGCTCAATTTGGAGATTT TCATGCTCGGAATCAACAGCAACATTAACTCGTTGGTCGCTCAACAGAACCTCAACGGCTCGCAAGGCGC CCTGTCCCAAGCGATCACCCGCCTGTCGTCGGGCAAGCGCATCAACAGCGCGGCGGACGATGCGGCCGGC CTCGCGATCGCCACCCGGATGCAAACGCAGATCAACGGCCTGAACCAGGGCGTGTCGAACGCGAACGACG GCGTGTCGATCCTGCAAACGGCATCGAGCGGCCTGACCTCGCTCACCAACAGCCTGCAGCGTATCCGCCA GCTCGCCGTGCAGGCCTCGAACGGCCCGCTGAGCGCGAGCGACGCGTCGGCGCTGCAACAGGAAGTCGCG CAGCAGATCTCGGAAGTGAACCGTATCGCTTCGCAGACGAACTACAACGGCAAGAACATCCTCGACGGCT CGGCAGGCACGCTGAGCTTCCAGGTCGGCGCGAACGTCGGCCAGACGGTCTCCGTCGACCTCACGCAAAG CATGTCGGCGGCGAAGATCGGCGGCGGCATGGTTCAGACGGGCCAGACGCTCGGCACGATCAAGGTGGCG ATCGACTCGAGCGGCGCGGCCTGGTCGTCGGGCAGCACCGGCCAGGAGACGACGCAGATCAACGTCGTGT CGGACGGCAAGGGCGGCTTCACGTTCACCGATCAGAACAACCAGGCGCTGTCGTCGACGGCCGTGACCGC CGTGTTCGGCTCGTCGACCGCCGGCACGGGCACGGCGGCCTCGCCGTCGTTCCAGACGCTGGCGCTGTCG ACTTCGGCAACCAGCGCGCTGTCCGCGACGGACCAGGCGAACGCCACGGCGATGGTTGCGCAGATCAACG CGGTCAACAAGCCGCAAACGGTCTCGAACCTCGACATCAGCACGCAGACGGGCGCGTACCAGGCGATGGT ATCGATCGACAACGCACTCGCGACGGTCAACAATCTGCAGGCAACGCTCGGCGCGGCGCAAAACCGCTTC ACCGCGATCGCGACGACGCAGCAAGCCGGCTCGAACAACCTCGCGCAGGCGCAATCGCAAATCCAGAGCG CGGACTTTGCTCAGGAAACCGCGAACCTGTCGCGCGCGCAAGTGCTCCAGCAAGCCGGCATCTCGGTGCT CGCGCAAGCGAACTCGCTGCCGCAGCAAGTGCTGAAGCTCCTGCAATAA [SEQ ID No: 2] Accordingly, in a preferred embodiment, FliC_BP is encoded by the nucleotide sequence substantially as set out in SEQ ID No: 2, or a fragment or variant thereof. When produced by Burkholderia pseudomallei, the flagella are normally non- glycosylated. Glycosylation of the Burkholderia pseudomallei flagella sequences may inhibit the TLR5 ligand function in a mammalian viral system. To check for N- glycosylation sites in the Burkholderia pseudomallei flagella, the inventors ran the sequences through Net N Glyc server website software.3x N-glycosylation sites were found (AA No 18, 270, 358). To avoid N-glycosylation sites, Asparagine's (N) on the glycosylation site were changed to glutamines (Q). Accordingly, in a preferred embodiment, FliC_BP may comprise an optimised amino acid sequence of SEQ ID No: 3, which is provided herein, as follows: LGINSNINSLVAQQNLQGSQGALSQAITRLSSGKRINSAADDAAGLAIATRMQTQINGLNQGVSNANDGV SILQTASSGLTSLTNSLQRIRQLAVQASNGPLSASDASALQQEVAQQISEVNRIASQTNYNGKNILDGSA GTLSFQVGANVGQTVSVDLTQSMSAAKIGGGMVQTGQTLGTIKVAIDSSGAAWSSGSTGQETTQINVVSD GKGGFTFTDQNNQALSSTAVTAVFGSSTAGTGTAASPSFQTLALSTSATSALSATDQAQATAMVAQINAV NKPQTVSNLDISTQTGAYQAMVSIDNALATVNNLQATLGAAQNRFTAIATTQQAGSNNLAQAQSQIQSAD FAQETAQLSRAQVLQQAGISVLAQANSLPQQVLKLLQ [SEQ ID No: 3] Accordingly, in a preferred embodiment, FliC_BP comprises or consists of an optimised amino acid sequence substantially as set out in SEQ ID No: 3, or a fragment or variant thereof. In one embodiment, FliC_BP may be encoded by the optimised nucleic acid sequence of SEQ ID No: 4, which is provided herein as follows: gccctgggaatcaatagtaacatcaactctctggtcgctcagcagaatctgcagggcagccagggcgccc tgtctcaggccatcaccagactgtcctctggcaagcggatcaattccgccgccgacgatgccgccggcct ggccatcgccaccagaatgcagacccagatcaatggcctgaaccagggcgtgtctaacgccaatgatggc gtgagcatcctgcagaccgcctccagcggcctgacatctctgaccaacagcctgcagaggatcagacagc tggccgtgcaggccagcaacggccctctgtctgcctccgatgccagcgccctgcagcaggaggtggccca gcagatctctgaggtgaacagaatcgccagccagaccaactacaatggcaagaacatcctggatggctct gccggcaccctgtcctttcaggtgggcgccaacgtgggccagaccgtgtctgtggatctgacccagtcta tgagcgccgccaagatcggcggcggcatggtgcagacaggccagaccctgggcacaatcaaggtggccat cgatagctccggcgccgcctggtctagcggctccaccggccaggagacaacccagatcaacgtggtgtcc gacggcaagggcggcttcacctttacagaccagaacaatcaggccctgtccagcacagccgtgaccgccg tgtttggctcctctacagccggcaccggcacagccgcctcccctagctttcagaccctggccctgtccac atctgccacatctgccctgtccgccacagatcaggcccaggccacggccatggtggcccagatcaatgcc gtgaacaagccccagacagtgtccaacctggatatcagcacccagaccggcgcctatcaggccatggtgt ccatcgacaacgccctggccaccgtgaacaatctgcaggccacactgggcgccgcccagaaccggtttac cgccatcgccacaacccagcaggccggctccaacaatctggcccaggcccagagccagatccagagcgcc gattttgcccaggagacagcccagctgagcagagcccaggtgctgcagcaggccgggatctccgtcctgg cacaggctaatagtctgccacagcaggtcctgaaactgctgcag [SEQ ID No: 4] Accordingly, in a preferred embodiment, FliC_BP is encoded by the optimised nucleotide sequence substantially as set out in SEQ ID No: 4, or a fragment or variant thereof. In order to secrete the TLR agonist (e.g. the FliC_BP flagella protein) out of the cells, the inventors further added a CD33 secretion signal. Accordingly, in a preferred embodiment, the first coding sequence of the genetic construct according to the first aspect encodes a CD33 secretion signal. Preferably, the CD33 secretion signal is 5’ of the sequence encoding the TLR agonist (e.g. FliC_BP). The CD33 secretion signal may comprise an amino acid sequence of SEQ ID No: 5, which is provided herein, as follows: MPLLLLLPLLWAGALA [SEQ ID No: 5] Accordingly, in a preferred embodiment, the CD33 secretion signal comprises or consists of an amino acid sequence substantially as set out in SEQ ID No: 5, or a fragment or variant thereof. In one embodiment, the CD33 secretion signal may be encoded by the nucleic acid sequence of SEQ ID No: 6, which is provided herein as follows: atg cct ctg ctg ctg ctg ctg cct ctg ctg tgg gct ggg gct ctg gcc [SEQ ID No: 6] Accordingly, in a preferred embodiment, the CD33 secretion signal is encoded by the nucleotide sequence substantially as set out in SEQ ID No: 6, or a fragment or variant thereof. Accordingly, it will be appreciated that FliC_BP and the CD33 secretion signal may together comprise an amino acid sequence of SEQ ID No: 7, which is provided herein, as follows: MPLLLLLPLLWAGALALGINSNINSLVAQQNLQGSQGALSQAITRLSSGKRINSAADDAAGLAIATRMQT QINGLNQGVSNANDGVSILQTASSGLTSLTNSLQRIRQLAVQASNGPLSASDASALQQEVAQQISEVNRI ASQTNYNGKNILDGSAGTLSFQVGANVGQTVSVDLTQSMSAAKIGGGMVQTGQTLGTIKVAIDSSGAAWS SGSTGQETTQINVVSDGKGGFTFTDQNNQALSSTAVTAVFGSSTAGTGTAASPSFQTLALSTSATSALSA TDQAQATAMVAQINAVNKPQTVSNLDISTQTGAYQAMVSIDNALATVNNLQATLGAAQNRFTAIATTQQA GSNNLAQAQSQIQSADFAQETAQLSRAQVLQQAGISVLAQANSLPQQVLKLLQ [SEQ ID No: 7] Accordingly, in a preferred embodiment, FliC_BP and the CD33 secretion signal comprises or consists of an amino acid sequence substantially as set out in SEQ ID No: 7, or a fragment or variant thereof. In one embodiment, FliC_BP and the CD33 secretion signal may be encoded by the nucleic acid sequence of SEQ ID No: 8, which is provided herein as follows: GGATCCGCCACCATGCCTCTGCTGCTGCTGCTGCCTCTGCTGTGGGCTGGGGCTCTGGCCCTGGGAATCA ATAGTAACATCAACTCTCTGGTCGCTCAGCAGAATCTGCAGGGCAGCCAGGGCGCCCTGTCTCAGGCCAT CACCAGACTGTCCTCTGGCAAGCGGATCAATTCCGCCGCCGACGATGCCGCCGGCCTGGCCATCGCCACC AGAATGCAGACCCAGATCAATGGCCTGAACCAGGGCGTGTCTAACGCCAATGATGGCGTGAGCATCCTGC AGACCGCCTCCAGCGGCCTGACATCTCTGACCAACAGCCTGCAGAGGATCAGACAGCTGGCCGTGCAGGC CAGCAACGGCCCTCTGTCTGCCTCCGATGCCAGCGCCCTGCAGCAGGAGGTGGCCCAGCAGATCTCTGAG GTGAACAGAATCGCCAGCCAGACCAACTACAATGGCAAGAACATCCTGGATGGCTCTGCCGGCACCCTGT CCTTTCAGGTGGGCGCCAACGTGGGCCAGACCGTGTCTGTGGATCTGACCCAGTCTATGAGCGCCGCCAA GATCGGCGGCGGCATGGTGCAGACAGGCCAGACCCTGGGCACAATCAAGGTGGCCATCGATAGCTCCGGC GCCGCCTGGTCTAGCGGCTCCACCGGCCAGGAGACAACCCAGATCAACGTGGTGTCCGACGGCAAGGGCG GCTTCACCTTTACAGACCAGAACAATCAGGCCCTGTCCAGCACAGCCGTGACCGCCGTGTTTGGCTCCTC TACAGCCGGCACCGGCACAGCCGCCTCCCCTAGCTTTCAGACCCTGGCCCTGTCCACATCTGCCACATCT GCCCTGTCCGCCACAGATCAGGCCCAGGCCACGGCCATGGTGGCCCAGATCAATGCCGTGAACAAGCCCC AGACAGTGTCCAACCTGGATATCAGCACCCAGACCGGCGCCTATCAGGCCATGGTGTCCATCGACAACGC CCTGGCCACCGTGAACAATCTGCAGGCCACACTGGGCGCCGCCCAGAACCGGTTTACCGCCATCGCCACA ACCCAGCAGGCCGGCTCCAACAATCTGGCCCAGGCCCAGAGCCAGATCCAGAGCGCCGATTTTGCCCAGG AGACAGCCCAGCTGAGCAGAGCCCAGGTGCTGCAGCAGGCCGGGATCTCCGTCCTGGCACAGGCTAATAG TCTGCCACAGCAGGTCCTGAAACTGCTGCAGTAACAATTGGAATTC [SEQ ID No: 8] Accordingly, in a preferred embodiment, FliC_BP and the CD33 secretion signal are encoded by the nucleotide sequence substantially as set out in SEQ ID No: 8, or a fragment or variant thereof. The cytokine may be a chemokine. The chemokine may be selected from a group consisting of: CXCL10, CXCL9, CXCL11, CXCL8, CXCL12, CCL2, CCL3 and CCL5. In a preferred embodiment, the cytokine is selected from a group consisting of: IL-2, IL-6, IL-10, IL-12, non-secreting IL-12, IL-15, IL-17, IL-21, IL-33, and type I and type II interferons (IFN- ^, IFN ^ ^, IFN- ^). Most preferably, however, the cytokine is IL-15. IL-15 may comprise an amino acid sequence of SEQ ID No: 9, which is provided herein, as follows: MDWTWILFLVAAATRVHSNWVNVISDLKKIEDLIQSMHIDATLYTESDVHPSCKVTAMKCFLLELQVISL ESGDASIHDTVENLIILANNSLSSNGNVTESGCKECEELEKKNIKEFLQSFVHIVQMFINTS [SEQ ID No: 9] Accordingly, in a preferred embodiment, IL-15 comprises or consists of an amino acid sequence substantially as set out in SEQ ID No: 9, or a fragment or variant thereof. In one embodiment, IL-15 may be encoded by the nucleic acid sequence of SEQ ID No: 10, which is provided herein as follows: ATGGACTGGACCTGGATTCTGTTCCTGGTGGCAGCAGCAACAAGGGTGCACAGCAACTGGGTGAATGTGA TCTCCGACCTGAAGAAGATCGAGGATCTGATCCAGAGCATGCACATCGACGCCACCCTGTACACAGAGTC CGATGTGCACCCCTCTTGCAAGGTGACCGCCATGAAGTGTTTTCTGCTGGAGCTGCAGGTCATCTCTCTG GAGAGCGGCGACGCCTCCATCCACGATACCGTGGAGAACCTGATCATCCTGGCCAACAATTCTCTGAGCT CCAACGGCAATGTGACAGAGAGCGGCTGCAAGGAGTGTGAGGAGCTGGAGAAGAAGAACATCAAGGAGTT CCTGCAGTCCTTTGTGCACATCGTGCAGATGTTCATCAATACATCT [SEQ ID No: 10] Accordingly, in a preferred embodiment, IL-15 is encoded by the nucleotide sequence substantially as set out in SEQ ID No: 10, or a fragment or variant thereof. Preferably, the first coding sequence encoding the TLR agonist and the second coding sequence encoding the cytokine are under the control of one or more promoters. Most preferably, the first coding sequence encoding the TLR agonist is under the control of a first promoter, and the second coding sequence encoding the cytokine is under the control of a second promoter. Alternatively, in another embodiment, the gene encoding the TLR agonist and the gene encoding the cytokine are under the control of a single promoter. Accordingly, in this embodiment, the genetic construct according to the first aspect comprises a first promoter, operably linked to the first coding sequence encoding the TLR agonist and the second coding sequence encoding the cytokine. Preferably, when under the control of a single promoter, the first and second coding sequence are expressed as a fusion protein. The first and second coding sequence may be separated by a nucleotide sequence encoding a fusion protein linker. In a preferred embodiment, the genetic construct according to the first aspect comprises a first promoter, operably linked to the first coding sequence encoding the TLR agonist, and / or a second promoter operably linked to the second coding sequence encoding the cytokine. The or each promoter may be any nucleotide sequence that is capable of inducing RNA polymerase to bind to, and transcribe, the coding sequence. The or each promoter may be a constitutive promoter or a regulated promoter. Examples of constitutive promoters include, but are not limited to, the cytomegalovirus (CMV) promoter, the RSV promoter, and the T7 polymerase promoter. Accordingly, in a preferred embodiment, the or each promoter may be selected from a group consisting of: CMV, RSV, T7 polymerase, EF1a, SV40, PGK1, Ac5, UBI, MP-84 and MP-135. Advantageously, the micro-promoters, MP-84 and MP-135, of adeno- associated virus are small in size and so they allow more space for the recombinant genes in smaller viruses. In a preferred embodiment, the first promoter operably linked to the first coding sequence encoding the TLR agonist (e.g. FliC_BP), is the cytomegalovirus (CMV) promoter. One embodiment of the nucleotide sequence encoding the cytomegalovirus (CMV) promoter is referred to herein as SEQ ID No: 12, as follows: agctctgcttatatagacctcccaccgtacacgcctaccgcccatttgcgtcaatggggcggagttgtta cgacattttggaaagtcccgttgattttggtgccaaaacaaactcccattgacgtcaatggggtggagac ttggaaatccccgtgagtcaaaccgctatccacgcccattgatgtactgccaaaaccgcatcaccatggt aatagcgatgactaatacgtagatgtactgccaagtaggaaagtcccataaggtcatgtactgggcataa tgccaggcgggccatttaccgtcattgacgtcaatagggggcgtacttggcatatgatacacttgatgta ctgccaagtgggcagtttaccgtaaatagtccacccattgacgtcaatggaaagtccctattggcgttac tatgggaacatacgtcattattgacgtcaatgggcgggggtcgttgggcggtcagccaggcgggccattt accgtaagttatgtaacgcggaactccatatatgggctatgaactaatgaccccgtaattgattactatt aataactagtcaataatcaatgtc [SEQ ID No: 12] Therefore, preferably the first promoter comprises or consists of a nucleic acid sequence substantially as set out in SEQ ID No: 12, or a fragment or variant thereof. Preferably, the second promoter operably linked to the second coding sequence encoding the cytokine, is the RSV promoter. One embodiment of the nucleotide sequence encoding the RSV promoter is referred to herein as SEQ ID No: 13, as follows: aatgtagtcttatgcaatacacttgtagtcttgcaacatggtaacgatgagttagcaacatgccttacaa ggagagaaaaagcaccgtgcatgccgattggtggaagtaaggtggtacgatcgtgccttattaggaaggc aacagacaggtctgacatggattggacgaaccactgaattccgcattgcagagataattgtatttaagtg cctagctcgatacaataaacgccatttgaccattcaccacattggtgtgcacc [SEQ ID No: 13] Therefore, preferably the second promoter comprises or consists of a nucleic acid sequence substantially as set out in SEQ ID No: 13, or a fragment or variant thereof. As can be seen in Figure 1, the genetic construct (or expression cassette) comprises two nucleotide sequences encoding two polyA tails. Preferably, the genetic construct or expression cassette comprises a nucleotide sequence encoding a poly-A tail. More preferably, the genetic construct or expression cassette comprises a first and second nucleotide sequence encoding a first and second poly-A tail, respectively. Preferably, the first and second nucleotide sequences encoding the poly-A tails flank the cassette. Preferably, the first poly-A tail coding sequence is disposed 3’ of the first coding sequence encoding the TLR agonist (e.g. FliC_BP). Preferably, the second poly-A tail coding sequence is disposed 3’ of the second coding sequence encoding the cytokine (e.g. IL-15). One embodiment of the nucleotide sequence encoding the first and / or second poly-A tail (BGH poly-A tail for CMV) is referred to herein as SEQ ID No: 14, as follows: ccatagagcccaccgcatccccagcatgcctgctattgtcttcccaatcctcccccttgctgtcctgccc caccccaccccccagaatagaatgacacctactcagacaatgcgatgcaatttcctcattttattaggaa aggacagtgggagtggcaccttccagggtcaaggaaggcacgggggaggggcaaacaacagatggctggc aactagaaggcacag [SEQ ID No: 14] Preferably, therefore, the first and / or second poly-A tail comprises a nucleic acid sequence substantially as set out in SEQ ID No: 14, or a fragment or variant thereof. One embodiment of the nucleotide sequence encoding the first and / or second poly-A tail (SV40 poly-A tail for RSV) is referred to herein as SEQ ID No: 15, as follows: aacttgtttattgcagcttataatggttacaaataaagcaatagcatcacaaatttcacaaataaagcat ttttttcactgc [SEQ ID No: 15] Preferably, therefore, the first and / or second poly-A tail comprises a nucleic acid sequence substantially as set out in SEQ ID No: 15, or a fragment or variant thereof. Preferably, the genetic construct of the first aspect or the expression cassette of the second aspect is harboured on an oncolytic virus. Accordingly, in a third aspect of the invention, there is provided an oncolytic virus comprising the genetic construct of the first aspect or the expression cassette of the second aspect. As can be seen in Figure 1, the oncolytic virus comprises the first nucleotide sequence encoding the TLR agonist (e.g. a novel bacterial protein, preferably FliC_BP) under the control of the first promoter (e.g. CMV), and the second nucleotide sequence encoding the cytokine (e.g. IL-15) under the control of the second promoter (e.g. RSV). The virus also comprises two flanking polyA tails either side of the expression cassette. The inventors found that FliC_BP is a particularly effective candidate for enhancing antitumour immune responses seen with oncolytic viruses with an innate bacterial response. In particular, the inventors discovered that FLiC BP can trigger TLR5- and TLR2-mediated signalling pathways, suggesting either the unique property of FliC_BP to bind to the TLR2 receptor or the occurrence of crosstalk between TLR5 (the primary receptor for flagellin) and TLR2. This crosstalk between TLR2 and TLR5 likely amplifies the production of proinflammatory cytokines, thereby contributing to a more robust and immediate immune response. The inventors believe that they are the first to have integrated an expression cassette encoding FliC_BP onto an oncolytic virus. Accordingly, in a fourth aspect of the invention, there is provided an oncolytic virus comprising a nucleotide sequence, which encodes Burkholderia pseudomallei flagellin protein (FliC_BP). The nucleotide sequence encoding FliC_BP may be as described above in relation to the first or second aspect. For example, the nucleotide sequence encoding FliC_BP may comprise or consist of the nucleotide sequence substantially as set out in SEQ ID No: 2 or 4, or a fragment or variant thereof. Preferably, the oncolytic virus according to the fourth aspect comprises a first promoter, operably linked to the nucleotide sequence, which encodes FliC_BP. The promoter may be as described above in relation to the first or second aspect. For example, the promoter may be selected from cytomegalovirus (CMV) promoter, the RSV promoter, and the T7 polymerase promoter. In a preferred embodiment, the promoter operably linked to the nucleotide sequence encoding FliC_BP, is the cytomegalovirus (CMV) promoter, preferably comprising or consisting of a nucleic acid sequence substantially as set out in SEQ ID No: 12, or a fragment or variant thereof. As used herein, the term oncolytic virus in relation to the third or fourth aspect will be understood to be a virus that infects and lyses cancer cells, but not normal or healthy cells. The oncolytic virus may be a native virus that is naturally oncolytic or may be engineered by modifying one or more viral genes so as to increase tumour selectivity. Preferably, the oncolytic virus is selected from a group consisting of: herpes simplex virus (HSV), Coxsackie virus, Maraba virus, measles virus (MV), Newcastle disease virus (NDV), poliovirus, reovirus, retroviruses, Seneca Valley virus (SVV), Alphavirus such as Semliki Forest virus (SFV) and Sindbis virus (SINV), Vesicular stomatitis virus (VSV), Sindbis virus (SBV), Adenovirus, Poxvirus, Parvovirus, Flavivirus such as Zika virus, Paramyxo virus, Picornavirus, and Rhabdovirus. Most preferably, the oncolytic virus is herpes simplex virus (HSV). The HSV virus may be HSV-1 or HSV-2. Preferably, the HSV virus is HSV-1. Most preferably, the HSV virus is the HSV-1 strain 17+. The inventors have discovered that the oncolytic virus (e.g. HSV) contributes to the therapeutic effect when it is in a live replicating state, resulting in maximal oncolysis of bladder tumour cells. Advantageously, the live replicating oncolytic virus facilitates the direct destruction of cancer cells and promotes the release of therapeutic proteins, such as IL-15 and FliC_BP along with tumour antigens. This release is essential for stimulating both innate and adaptive immune responses against tumour antigens and viral proteins leading to sustained anti-tumour immunity. Accordingly, in some embodiments, the oncolytic virus is a live, replicating virus. In some embodiments, the oncolytic virus is live, replicating HSV. In some embodiments, the oncolytic virus is live, replicating HSV-1 or HSV-2. The HSV-1 neurovirulence protein, infected cell protein 34.5 (ICP34.5), is necessary for HSV-1 infection of neurons and other healthy cells as it binds to, and blocks, PKR, allowing for viral replication. ICP34.5 blocks PKR pathway activation by binding to, and activating, the PP1α phosphatase, which dephosphorylates eIF-2α preventing the shutdown of protein translation by eIF-2. As such, for the infection of cancer cells, it is preferable that the ICP34.5 gene is not present. The HSV-1 genome is a single, linear molecule of double-stranded DNA approximately 152,000 bp in length. It is divided into two unique segments called long (U L ) and short (U S ). Short regions of repeated sequence occur at the genome ends and between the L and S segments. ICP34.5 is encoded within the two short regions of repeated sequence of UL. Therefore, there is two copies of the gene per genome. The first ICP34.5 gene is encoded between 513-1539bp and the second is encoded between 124832-125858bp in the genome. Accordingly, in a preferred embodiment, the oncolytic virus comprises a functionally deleted ICP34.5 gene. The ICP34.5 gene may be functionally deleted if it is rendered absent, disrupted or non-functional. More preferably, the oncolytic herpes simplex virus comprises a functional deletion of at least two copies of the ICP34.5 gene. Advantageously, this functional deletion has implications for eliminating the pathogenesis of HSV-1 and also enhances viral replication in cancer cells. ICP34.5- deficient HSV-1 infection of healthy cells activates PKR, resulting in abortive infection. In contrast, this HSV can replicate in cancer cells because PKR activity is not activated. Functional deletion of the ICP34.5 gene may be achieved by introducing one or more mutation into the viral DNA, such that the expressed protein is non-functional or has no low activity. Alternatively, functional deletion of the ICP34.5 gene may be achieved by contacting the virus with an inhibitor of the encoded protein, for example RNA, e.g. siRNA, specific to ICP34.5. However, preferably, functional deletion of the ICP34.5 gene is achieved by integrating the genetic construct of the first aspect or the expression cassette according to the second aspect into the ICP34.5 gene. Preferably, the oncolytic virus comprises a functionally deleted ICP47 gene. The ICP47 gene may be functionally deleted if it is rendered absent, disrupted or non-functional. This ICP47 functional deletion has multiple effects that improve the lytic activity of HSV-1 ICP34.5 / − mutants. US11 encodes for UL11, which binds PKR, preventing eIF- 2α phosphorylation, thus permitting HSV-1 replication. Furthermore, US11 is normally expressed later during infection well after ICP34.5 would inhibit PKR. However, the deletion in such HSV virus results in the regulation of US11 by the α47 promoter, which causes UL11 to be expressed as an immediate early gene, and this blocks PKR activity before PKR is able to terminate protein synthesis. In many cancer cells, the type I IFN signalling pathways are disrupted, and in the absence of ICP34.5, viral replication occurs in cancer cells lacking normal IFN signalling, whereas normal cells will be less permissive because IFN signalling is intact. This feature of HSV-1 ICP34.5 / − ICP47 - mutants adds an important safety component because infection of normal cells will be aborted in the absence of PKR and type I IFN signalling defects. Accordingly, in a preferred embodiment, the oncolytic virus according to the third aspect may comprise, in this specified order, a 5’ coding sequence encoding a cytokine (e.g. IL-15), and a 3’ coding sequence encoding a TLR agonist (e.g. FliC_BP). Alternatively, the oncolytic virus according to the third aspect may comprise, in this specified order, a 5’ coding sequence encoding a TLR agonist (e.g. FliC_BP), and a 3’ coding sequence encoding a cytokine (e.g. IL-15). The use of 5’ and 3’ indicates that the features are either upstream or downstream, and is not intended to indicate that the features are necessarily terminal features. In a particular embodiment, the oncolytic virus according to the third aspect may comprise in this specified order, a 5’ coding sequence encoding a cytokine (e.g. IL-15), a first promoter (e.g. RSV), a second promoter (e.g. CMV) and a 3’ coding sequence encoding a TLR agonist (e.g. FliC_BP). Alternatively, the oncolytic virus according to the third aspect may comprise, in this specified order, a 5’ coding sequence encoding a TLR agonist (e.g. FliC_BP), a first promoter (e.g. CMV), a second promoter (e.g. RSV) and a 3’ coding sequence encoding a cytokine (e.g. IL-15). Alternatively, in another embodiment, the oncolytic virus according to the third aspect may comprise in this specified order, a 5’ first promoter (e.g. RSV), a coding sequence encoding a cytokine (e.g. IL-15), a second promoter (e.g. CMV) and a 3’ coding sequence encoding a TLR agonist (e.g. FliC_BP). Alternatively, the oncolytic virus according to the third aspect may comprise, in this specified order, a 5’ first promoter (e.g. CMV), a coding sequence encoding a TLR agonist (e.g. FliC_BP), a second promoter (e.g. RSV) and a 3’ coding sequence encoding a cytokine (e.g. IL-15). In a particular embodiment, the oncolytic virus according to the third aspect may comprise in this specified order, a 5’ poly-A tail, a coding sequence encoding a cytokine (e.g. IL-15), a first promoter (e.g. RSV), a second promoter (e.g. CMV), a coding sequence encoding a TLR agonist (e.g. FliC_BP), and a 3’ poly-A tail. Alternatively, the oncolytic virus according to the third aspect may comprise, in this specified order, a 5’ poly-A tail, a coding sequence encoding a TLR agonist (e.g. FliC_BP), a first promoter (e.g. CMV), a second promoter (e.g. RSV) and a coding sequence encoding a cytokine (e.g. IL-15), and a 3’ poly-A tail. Alternatively, in another particular embodiment, the oncolytic virus according to the third aspect may comprise in this specified order, a 5’ first promoter (e.g. RSV), a coding sequence encoding a cytokine (e.g. IL-15), a poly-A tail, a second promoter (e.g. CMV), a coding sequence encoding a TLR agonist (e.g. FliC_BP), and a 3’ poly-A tail. Alternatively, the oncolytic virus according to the third aspect may comprise, in this specified order, a 5’ first promoter (e.g. CMV), a coding sequence encoding a TLR agonist (e.g. FliC_BP), a poly-A tail, a second promoter (e.g. RSV), a coding sequence encoding a cytokine (e.g. IL-15), and a 3’ poly-A tail. In a preferred embodiment, the oncolytic virus according to the fourth aspect may comprise, in this specified order, a 5’ promoter (e.g. CMV), and a 3’ coding sequence encoding a TLR agonist (e.g. FliC_BP). In a particular embodiment, the oncolytic virus according to the fourth aspect may comprise in this specified order, a 5’ promoter (e.g. CMV), a coding sequence encoding a TLR agonist (e.g. FliC_BP), and a 3’ poly-A tail. The inventors have demonstrated that the combination of a TLR agonist and cytokine in the context of an oncolytic virus (e.g. HSV), creates enhanced acquired immunity to the tumour whilst potentiating the oncolytic activity of the virus. The inventors surprisingly discovered, therefore, that an antigen is not required to produce a therapy for cancer. Accordingly, in some embodiments, the genetic construct, expression cassette and oncolytic virus according to the invention, do not comprise an antigen (e.g. a tumour or cancer antigen). From the foregoing, the skilled person will appreciate the nucleotide and amino acid sequence of an embodiment of the oncolytic virus of the third and fourth aspect. In some embodiments, a shuttle vector may be used to produce the oncolytic virus according to the invention. Thus, for the avoidance of doubt, the nucleotide sequence encoding the shuttle vector (known as p-34.5 DC RSV CMV) used to produce the oncolytic virus according to the invention, is provided herein as SEQ ID No: 11, as follows: gctgcggccgccagcgcggcggggcccggccaaccagcgtccgccgagtcttcggggcccggcccattgg gcgggagttaccgcccaatgggccgggccgcccacttcccggtatggtaattaaaaacttgcaagaggcc ttgttccgcttcccggtatggtaattagaaactcattaatgggcggccccggccgcccttcccgcttccg gcaattcccgcggcccttaatgggcaaccccggtattccccgcctcccgcgccgcgcgtaaccactcccc tggggttccgggttatgctaattgcttttttggcggaacacacggcccctcgcgcattggcccgcgggtc gctcaatgaacccgcattggtcccctggggttccgggtatggtaatgagtttcttcgggaaggcgggaag ccccggggcaccgacgcaggccaagcccctgttgcgtcggcgggaggggcatgctaatggggttctttgg gggacaccgggttgggcccccaaatcgggggccgggccgtgcatgctaatgatattctttgggggcgccg ggttggtccccggggacggggccgccccgcggtgggcctgcctcccctgggacgcgcggccattggggga atcgtcactgccgcccctttggggaggggaaaggcgtggggtataagttagccctggcccgacagtctgg tcgcatttgcacctcggcactcggagcgagacgcagcagccaggcagactcgggccgccccctctccgca tcaccacagaagccccgcctacgttgcgacccccagggaccctccgtccgcgaccctccagccgcatacg acccccatggagccccgccccggagcgagtacccgccggcctgagggccgcccccagcgcgaggtgaggg gccgggcgccatgtctggggcgccatattggggggcgccatattggggggcgccatgttgggggaccccc gacccttacactggaaccggccgccatgttgggggacccccactcatacacgggagccgggcgccatgtt ggggcgccatgttagggggcgtggaaccccgtgacactatatatacagggaccgggggcgccatgttagg gggtgcggaaccccctgaccctatatatacagggaccggggtcgccctgttgggggtcgccatgtgaccc cctgactttatatatacagacccccaacacatacacatggcccctttgactcagacgcagggcccggggt cgccgtgggaccccctgactcatacacagagacacgcccccacaacaaacacacaaggaccggggtcgcc gtgttgggggcgtggtccccactgactcatacgcaggccccccttactcacacgcatctaggggggtggg gaggagccgcccgccatatttgggggacgccgtgggacccccgactccggtgcgtctggagggcgggaga agagggaagaagaggggtcgggatctgccggtctccctatagtgagtcgtattaatttcgataagccagg ttaacctgcattaatgaatcggccaacgcgcggggagaggcggtttgcgtattgggcgctcttccgcttc ctcgctcactgactcgctgcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggta atacggttatccacagaatcaggggataacgcaggaaagaacatgtgagcaaaaggccagcaaaaggcca ggaaccgtaaaaaggccgcgttgctggcgtttttccataggctccgcccccctgacgagcatcacaaaaa tcgacgctcaagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctggaagc tccctcgtgcgctctcctgttccgaccctgccgcttaccggatacctgtccgcctttctcccttcgggaa gcgtggcgctttctcaatgctcacgctgtaggtatctcagttcggtgtaggtcgttcgctccaagctggg ctgtgtgcacgaaccccccgttcagcccgaccgctgcgccttatccggtaactatcgtcttgagtccaac ccggtaagacacgacttatcgccactggcagcagccactggtaacaggattagcagagcgaggtatgtag gcggtgctacagagttcttgaagtggtggcctaactacggctacactagaaggacagtatttggtatctg cgctctgctgaagccagttaccttcggaaaaagagttggtagctcttgatccggcaaacaaaccaccgct ggtagcggtggtttttttgtttgcaagcagcagattacgcgcagaaaaaaaggatctcaagaagatcctt tgatcttttctacggggtctgacgctcagtggaacgaaaactcacgttaagggattttggtcatgagatt atcaaaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaatcaatctaaagtatatat gagtaaacttggtctgacagttaccaatgcttaatcagtgaggcacctatctcagcgatctgtctatttc gttcatccatagttgcctgactccccgtcgtgtagataactacgatacgggagggcttaccatctggccc cagtgctgcaatgataccgcgagacccacgctcaccggctccagatttatcagcaataaaccagccagcc ggaagggccgagcgcagaagtggtcctgcaactttatccgcctccatccagtctattaattgttgccggg aagctagagtaagtagttcgccagttaatagtttgcgcaacgttgttgccattgctacaggcatcgtggt gtcacgctcgtcgtttggtatggcttcattcagctccggttcccaacgatcaaggcgagttacatgatcc cccatgttgtgcaaaaaagcggttagctccttcggtcctccgatcgttgtcagaagtaagttggccgcag tgttatcactcatggttatggcagcactgcataattctcttactgtcatgccatccgtaagatgcttttc tgtgactggtgagtactcaaccaagtcattctgagaatagtgtatgcggcgaccgagttgctcttgcccg gcgtcaatacgggataataccgcgccacatagcagaactttaaaagtgctcatcattggaaaacgttctt cggggcgaaaactctcaaggatcttaccgctgttgagatccagttcgatgtaacccactcgtgcacccaa ctgatcttcagcatcttttactttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgca aaaaagggaataagggcgacacggaaatgttgaatactcatactcttcctttttcaatattattgaagca tttatcagggttattgtctcatgagcggatacatatttgaatgtatttagaaaaataaacaaataggggt tccgcgcacatttccccgaaaagtgccacctgacgtctaagaaaccattattatcatgacattaacctat aaaaataggcgtatcacgaggccctttcgtctcgcgcgtttcggtgatgacggtgaaaacctctgacaca tgcagctcccggagacggtcacagcttgtctgtaagcggatgccgggagcagacaagcccgtcagggcgc gtcagcgggtgttggcgggtgtcggggctggcttaactatgcggcatcagagcagattgtactgagagtg caccatatggacatattgtcgttagaacgcggctacaattaatacataaccttatgtatcatacacatac gatttaggtgacactatagaactcgagcagctgaagcttgcatgcctgcaggtcgactctagaggatctc tgacctgagattggcggcactgaggtagagatgcccgaacccccccgagggagcgcgggacgcgccgggg agggctggggccggggagggctggggccggggagggctggggccggggagggctggggccggggagggct ggggccggggagggctggggccggggagggctggggctggggagggctggggctggggagggggcggtgg tgtgtagcaggagcggtgtgttgcgccggggtacgtctggaggagcgggaggtgcgcggtgacgtgtgga tgaggaacaggagttgttgcgcggtgagttgtcgctgtgagttgtgttgttgggcaggtgtggtggatga cgtgacgtgtgacgtgcggagtgcgccgtgctctgttggtttcacctgtggcagcccgggccccccgcgg gcgcgcgcgcgcgcaaaaaaggcgggcggcggtccgggcggcgtgcgcgcgcgcggcgggcgtggggggc ggggccgcgggagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcggggggag gagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcgggggg aggagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcgggg ggaggagcggggggaggagcggccagacgccgaaaacgggccccccccaaaacacaccccccgggggtcg cgcgcggccctttaaagcggtggcggcgggcagcccgggccccccgcggccgagactagcgagttagaca ggcaagcactactcgcctctgcacgcacatgcttgcctgtcaaactctaccaccccggcacgctctctgt ctccatggcccgccgccgccgccatcgcggcccccgccgcccccggccgcccgggcccacgggcgccgtc ccaaccgcacagtcccaggtaacctccacgcccaactcggaacccgcggtcaggagcgcgcccgcggccg cagcaacctggttctttccgcctcagaagccatagagcccaccgcatccccagcatgcctgctattgtct tcccaatcctcccccttgctgtcctgccccaccccaccccccagaatagaatgacacctactcagacaat gcgatgcaatttcctcattttattaggaaaggacagtgggagtggcaccttccagggtcaaggaaggcac gggggaggggcaaacaacagatggctggcaactagaaggcacagtcgaggctgatcagcgagctctagca tttaggtgacactatagaatagggccctctagctcgagcaattgcacgtgatggatccagcttgggtctc cctatagtgagtcgtattaatttcgataagccagtaagcagtgggttctctagttagccagagagctctg cttatatagacctcccaccgtacacgcctaccgcccatttgcgtcaatggggcggagttgttacgacatt ttggaaagtcccgttgattttggtgccaaaacaaactcccattgacgtcaatggggtggagacttggaaa tccccgtgagtcaaaccgctatccacgcccattgatgtactgccaaaaccgcatcaccatggtaatagcg atgactaatacgtagatgtactgccaagtaggaaagtcccataaggtcatgtactgggcataatgccagg cgggccatttaccgtcattgacgtcaatagggggcgtacttggcatatgatacacttgatgtactgccaa gtgggcagtttaccgtaaatagtccacccattgacgtcaatggaaagtccctattggcgttactatggga acatacgtcattattgacgtcaatgggcgggggtcgttgggcggtcagccaggcgggccatttaccgtaa gttatgtaacgcggaactccatatatgggctatgaactaatgaccccgtaattgattactattaataact agtcaataatcaatgtcaacgcgtatatctggcccgtacatcgcgatgtacgggccagatatacgcgtat ctgaggggactagggtgtgtttaggcgaaaagcggggcttcggttgtacgcggttaggagtcccctcagg atatagtagtttcgcttttgcatagggagggggaaatgtagtcttatgcaatacacttgtagtcttgcaa catggtaacgatgagttagcaacatgccttacaaggagagaaaaagcaccgtgcatgccgattggtggaa gtaaggtggtacgatcgtgccttattaggaaggcaacagacaggtctgacatggattggacgaaccactg aattccgcattgcagagataattgtatttaagtgcctagctcgatacaataaacgccatttgaccattca ccacattggtgtgcacctccaagcttgcggccgcgtttaaaccctgcaggtctagataataacgggcagg ccatgtctgcccgtatttcgcgtaaggaaatccattatgtactatttaaaaaacacaaacttttggatgt tcggtttattctttttcttttacttttttatcatgggagcctacttcccgtttttcccgatttggctaca tgacatcaaccatatcagcaaaagtgatacgggtattatttttgccgctatttctctgttctcgctatta ttccaaccgctgtttggtctgctttctgacaaactcggaacttgtttattgcagcttataatggttacaa ataaagcaatagcatcacaaatttcacaaataaagcatttttttcactgcattctagttgtggtttgtgt cgacttagatc [SEQ ID No: 11] Alternatively, in another embodiment, the nucleotide sequence encoding the shuttle vector (known as p-34.5 DC CMV S-FLiC-M3 RSV) used to produce the oncolytic virus according to the invention, is provided herein as SEQ ID No: 16, as follows: gctgcggccgccagcgcggcggggcccggccaaccagcgtccgccgagtcttcggggcccggcccattgg gcgggagttaccgcccaatgggccgggccgcccacttcccggtatggtaattaaaaacttgcaagaggcc ttgttccgcttcccggtatggtaattagaaactcattaatgggcggccccggccgcccttcccgcttccg gcaattcccgcggcccttaatgggcaaccccggtattccccgcctcccgcgccgcgcgtaaccactcccc tggggttccgggttatgctaattgcttttttggcggaacacacggcccctcgcgcattggcccgcgggtc gctcaatgaacccgcattggtcccctggggttccgggtatggtaatgagtttcttcgggaaggcgggaag ccccggggcaccgacgcaggccaagcccctgttgcgtcggcgggaggggcatgctaatggggttctttgg gggacaccgggttgggcccccaaatcgggggccgggccgtgcatgctaatgatattctttgggggcgccg ggttggtccccggggacggggccgccccgcggtgggcctgcctcccctgggacgcgcggccattggggga atcgtcactgccgcccctttggggaggggaaaggcgtggggtataagttagccctggcccgacagtctgg tcgcatttgcacctcggcactcggagcgagacgcagcagccaggcagactcgggccgccccctctccgca tcaccacagaagccccgcctacgttgcgacccccagggaccctccgtccgcgaccctccagccgcatacg acccccatggagccccgccccggagcgagtacccgccggcctgagggccgcccccagcgcgaggtgaggg gccgggcgccatgtctggggcgccatattggggggcgccatattggggggcgccatgttgggggaccccc gacccttacactggaaccggccgccatgttgggggacccccactcatacacgggagccgggcgccatgtt ggggcgccatgttagggggcgtggaaccccgtgacactatatatacagggaccgggggcgccatgttagg gggtgcggaaccccctgaccctatatatacagggaccggggtcgccctgttgggggtcgccatgtgaccc cctgactttatatatacagacccccaacacatacacatggcccctttgactcagacgcagggcccggggt cgccgtgggaccccctgactcatacacagagacacgcccccacaacaaacacacaaggaccggggtcgcc gtgttgggggcgtggtccccactgactcatacgcaggccccccttactcacacgcatctaggggggtggg gaggagccgcccgccatatttgggggacgccgtgggacccccgactccggtgcgtctggagggcgggaga agagggaagaagaggggtcgggatctgccggtctccctatagtgagtcgtattaatttcgataagccagg ttaacctgcattaatgaatcggccaacgcgcggggagaggcggtttgcgtattgggcgctcttccgcttc ctcgctcactgactcgctgcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggta atacggttatccacagaatcaggggataacgcaggaaagaacatgtgagcaaaaggccagcaaaaggcca ggaaccgtaaaaaggccgcgttgctggcgtttttccataggctccgcccccctgacgagcatcacaaaaa tcgacgctcaagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctggaagc tccctcgtgcgctctcctgttccgaccctgccgcttaccggatacctgtccgcctttctcccttcgggaa gcgtggcgctttctcaatgctcacgctgtaggtatctcagttcggtgtaggtcgttcgctccaagctggg ctgtgtgcacgaaccccccgttcagcccgaccgctgcgccttatccggtaactatcgtcttgagtccaac ccggtaagacacgacttatcgccactggcagcagccactggtaacaggattagcagagcgaggtatgtag gcggtgctacagagttcttgaagtggtggcctaactacggctacactagaaggacagtatttggtatctg cgctctgctgaagccagttaccttcggaaaaagagttggtagctcttgatccggcaaacaaaccaccgct ggtagcggtggtttttttgtttgcaagcagcagattacgcgcagaaaaaaaggatctcaagaagatcctt tgatcttttctacggggtctgacgctcagtggaacgaaaactcacgttaagggattttggtcatgagatt atcaaaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaatcaatctaaagtatatat gagtaaacttggtctgacagttaccaatgcttaatcagtgaggcacctatctcagcgatctgtctatttc gttcatccatagttgcctgactccccgtcgtgtagataactacgatacgggagggcttaccatctggccc cagtgctgcaatgataccgcgagacccacgctcaccggctccagatttatcagcaataaaccagccagcc ggaagggccgagcgcagaagtggtcctgcaactttatccgcctccatccagtctattaattgttgccggg aagctagagtaagtagttcgccagttaatagtttgcgcaacgttgttgccattgctacaggcatcgtggt gtcacgctcgtcgtttggtatggcttcattcagctccggttcccaacgatcaaggcgagttacatgatcc cccatgttgtgcaaaaaagcggttagctccttcggtcctccgatcgttgtcagaagtaagttggccgcag tgttatcactcatggttatggcagcactgcataattctcttactgtcatgccatccgtaagatgcttttc tgtgactggtgagtactcaaccaagtcattctgagaatagtgtatgcggcgaccgagttgctcttgcccg gcgtcaatacgggataataccgcgccacatagcagaactttaaaagtgctcatcattggaaaacgttctt cggggcgaaaactctcaaggatcttaccgctgttgagatccagttcgatgtaacccactcgtgcacccaa ctgatcttcagcatcttttactttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgca aaaaagggaataagggcgacacggaaatgttgaatactcatactcttcctttttcaatattattgaagca tttatcagggttattgtctcatgagcggatacatatttgaatgtatttagaaaaataaacaaataggggt tccgcgcacatttccccgaaaagtgccacctgacgtctaagaaaccattattatcatgacattaacctat aaaaataggcgtatcacgaggccctttcgtctcgcgcgtttcggtgatgacggtgaaaacctctgacaca tgcagctcccggagacggtcacagcttgtctgtaagcggatgccgggagcagacaagcccgtcagggcgc gtcagcgggtgttggcgggtgtcggggctggcttaactatgcggcatcagagcagattgtactgagagtg caccatatggacatattgtcgttagaacgcggctacaattaatacataaccttatgtatcatacacatac gatttaggtgacactatagaactcgagcagctgaagcttgcatgcctgcaggtcgactctagaggatctc tgacctgagattggcggcactgaggtagagatgcccgaacccccccgagggagcgcgggacgcgccgggg agggctggggccggggagggctggggccggggagggctggggccggggagggctggggccggggagggct ggggccggggagggctggggccggggagggctggggctggggagggctggggctggggagggggcggtgg tgtgtagcaggagcggtgtgttgcgccggggtacgtctggaggagcgggaggtgcgcggtgacgtgtgga tgaggaacaggagttgttgcgcggtgagttgtcgctgtgagttgtgttgttgggcaggtgtggtggatga cgtgacgtgtgacgtgcggagtgcgccgtgctctgttggtttcacctgtggcagcccgggccccccgcgg gcgcgcgcgcgcgcaaaaaaggcgggcggcggtccgggcggcgtgcgcgcgcgcggcgggcgtggggggc ggggccgcgggagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcggggggag gagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcgggggg aggagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcgggg ggaggagcggggggaggagcggccagacgccgaaaacgggccccccccaaaacacaccccccgggggtcg cgcgcggccctttaaagcggtggcggcgggcagcccgggccccccgcggccgagactagcgagttagaca ggcaagcactactcgcctctgcacgcacatgcttgcctgtcaaactctaccaccccggcacgctctctgt ctccatggcccgccgccgccgccatcgcggcccccgccgcccccggccgcccgggcccacgggcgccgtc ccaaccgcacagtcccaggtaacctccacgcccaactcggaacccgcggtcaggagcgcgcccgcggccg cagcaacctggttctttccgcctcagaagccatagagcccaccgcatccccagcatgcctgctattgtct tcccaatcctcccccttgctgtcctgccccaccccaccccccagaatagaatgacacctactcagacaat gcgatgcaatttcctcattttattaggaaaggacagtgggagtggcaccttccagggtcaaggaaggcac gggggaggggcaaacaacagatggctggcaactagaaggcacagtcgaggctgatcagcgagctctagca tttaggtgacactatagaatagggccctctagctcgagcAATTGTTACTGCAGCAGTTTCAGGACCTGCT GTGGCAGACTATTAGCCTGTGCCAGGACGGAGATCCCGGCCTGCTGCAGCACCTGGGCTCTGCTCAGCTG GGCTGTCTCCTGGGCAAAATCGGCGCTCTGGATCTGGCTCTGGGCCTGGGCCAGATTGTTGGAGCCGGCC TGCTGGGTTGTGGCGATGGCGGTAAACCGGTTCTGGGCGGCGCCCAGTGTGGCCTGCAGATTGTTCACGG TGGCCAGGGCGTTGTCGATGGACACCATGGCCTGATAGGCGCCGGTCTGGGTGCTGATATCCAGGTTGGA CACTGTCTGGGGCTTGTTCACGGCATTGATCTGGGCCACCATGGCCGTGGCCTGGGCCTGATCTGTGGCG GACAGGGCAGATGTGGCAGATGTGGACAGGGCCAGGGTCTGAAAGCTAGGGGAGGCGGCTGTGCCGGTGC CGGCTGTAGAGGAGCCAAACACGGCGGTCACGGCTGTGCTGGACAGGGCCTGATTGTTCTGGTCTGTAAA GGTGAAGCCGCCCTTGCCGTCGGACACCACGTTGATCTGGGTTGTCTCCTGGCCGGTGGAGCCGCTAGAC CAGGCGGCGCCGGAGCTATCGATGGCCACCTTGATTGTGCCCAGGGTCTGGCCTGTCTGCACCATGCCGC CGCCGATCTTGGCGGCGCTCATAGACTGGGTCAGATCCACAGACACGGTCTGGCCCACGTTGGCGCCCAC CTGAAAGGACAGGGTGCCGGCAGAGCCATCCAGGATGTTCTTGCCATTGTAGTTGGTCTGGCTGGCGATT CTGTTCACCTCAGAGATCTGCTGGGCCACCTCCTGCTGCAGGGCGCTGGCATCGGAGGCAGACAGAGGGC CGTTGCTGGCCTGCACGGCCAGCTGTCTGATCCTCTGCAGGCTGTTGGTCAGAGATGTCAGGCCGCTGGA GGCGGTCTGCAGGATGCTCACGCCATCATTGGCGTTAGACACGCCCTGGTTCAGGCCATTGATCTGGGTC TGCATTCTGGTGGCGATGGCCAGGCCGGCGGCATCGTCGGCGGCGGAATTGATCCGCTTGCCAGAGGACA GTCTGGTGATGGCCTGAGACAGGGCGCCCTGGCTGCCCTGCAGATTCTGCTGAGCGACCAGAGAGTTGAT GTTACTATTGATTCCCAGGGCCAGAGCCCCAGCCCACAGCAGAGGCAGCAGCAGCAGCAGAGGCATGGTG GCGgatccagcttgggtctccctatagtgagtcgtattaatttcgataagccagtaagcagtgggttctc tagttagccagagagctctgcttatatagacctcccaccgtacacgcctaccgcccatttgcgtcaatgg ggcggagttgttacgacattttggaaagtcccgttgattttggtgccaaaacaaactcccattgacgtca atggggtggagacttggaaatccccgtgagtcaaaccgctatccacgcccattgatgtactgccaaaacc gcatcaccatggtaatagcgatgactaatacgtagatgtactgccaagtaggaaagtcccataaggtcat gtactgggcataatgccaggcgggccatttaccgtcattgacgtcaatagggggcgtacttggcatatga tacacttgatgtactgccaagtgggcagtttaccgtaaatagtccacccattgacgtcaatggaaagtcc ctattggcgttactatgggaacatacgtcattattgacgtcaatgggcgggggtcgttgggcggtcagcc aggcgggccatttaccgtaagttatgtaacgcggaactccatatatgggctatgaactaatgaccccgta attgattactattaataactagtcaataatcaatgtcaacgcgtatatctggcccgtacatcgcgatgta cgggccagatatacgcgtatctgaggggactagggtgtgtttaggcgaaaagcggggcttcggttgtacg cggttaggagtcccctcaggatatagtagtttcgcttttgcatagggagggggaaatgtagtcttatgca atacacttgtagtcttgcaacatggtaacgatgagttagcaacatgccttacaaggagagaaaaagcacc gtgcatgccgattggtggaagtaaggtggtacgatcgtgccttattaggaaggcaacagacaggtctgac atggattggacgaaccactgaattccgcattgcagagataattgtatttaagtgcctagctcgatacaat aaacgccatttgaccattcaccacattggtgtgcacctccaagcttgcggccgcgtttaaaccctgcagg tctagataataacgggcaggccatgtctgcccgtatttcgcgtaaggaaatccattatgtactatttaaa aaacacaaacttttggatgttcggtttattctttttcttttacttttttatcatgggagcctacttcccg tttttcccgatttggctacatgacatcaaccatatcagcaaaagtgatacgggtattatttttgccgcta tttctctgttctcgctattattccaaccgctgtttggtctgctttctgacaaactcggaacttgtttatt gcagcttataatggttacaaataaagcaatagcatcacaaatttcacaaataaagcatttttttcactgc attctagttgtggtttgtgtcgacttagatc [SEQ ID No: 16] Alternatively, in another embodiment, the nucleotide sequence encoding the shuttle vector (known as p-34.5 DC CMV RSV IL-15ED) used to produce the oncolytic virus according to the invention, is provided herein as SEQ ID No: 17, as follows: gctgcggccgccagcgcggcggggcccggccaaccagcgtccgccgagtcttcggggcccggcccattgg gcgggagttaccgcccaatgggccgggccgcccacttcccggtatggtaattaaaaacttgcaagaggcc ttgttccgcttcccggtatggtaattagaaactcattaatgggcggccccggccgcccttcccgcttccg gcaattcccgcggcccttaatgggcaaccccggtattccccgcctcccgcgccgcgcgtaaccactcccc tggggttccgggttatgctaattgcttttttggcggaacacacggcccctcgcgcattggcccgcgggtc gctcaatgaacccgcattggtcccctggggttccgggtatggtaatgagtttcttcgggaaggcgggaag ccccggggcaccgacgcaggccaagcccctgttgcgtcggcgggaggggcatgctaatggggttctttgg gggacaccgggttgggcccccaaatcgggggccgggccgtgcatgctaatgatattctttgggggcgccg ggttggtccccggggacggggccgccccgcggtgggcctgcctcccctgggacgcgcggccattggggga atcgtcactgccgcccctttggggaggggaaaggcgtggggtataagttagccctggcccgacagtctgg tcgcatttgcacctcggcactcggagcgagacgcagcagccaggcagactcgggccgccccctctccgca tcaccacagaagccccgcctacgttgcgacccccagggaccctccgtccgcgaccctccagccgcatacg acccccatggagccccgccccggagcgagtacccgccggcctgagggccgcccccagcgcgaggtgaggg gccgggcgccatgtctggggcgccatattggggggcgccatattggggggcgccatgttgggggaccccc gacccttacactggaaccggccgccatgttgggggacccccactcatacacgggagccgggcgccatgtt ggggcgccatgttagggggcgtggaaccccgtgacactatatatacagggaccgggggcgccatgttagg gggtgcggaaccccctgaccctatatatacagggaccggggtcgccctgttgggggtcgccatgtgaccc cctgactttatatatacagacccccaacacatacacatggcccctttgactcagacgcagggcccggggt cgccgtgggaccccctgactcatacacagagacacgcccccacaacaaacacacaaggaccggggtcgcc gtgttgggggcgtggtccccactgactcatacgcaggccccccttactcacacgcatctaggggggtggg gaggagccgcccgccatatttgggggacgccgtgggacccccgactccggtgcgtctggagggcgggaga agagggaagaagaggggtcgggatctgccggtctccctatagtgagtcgtattaatttcgataagccagg ttaacctgcattaatgaatcggccaacgcgcggggagaggcggtttgcgtattgggcgctcttccgcttc ctcgctcactgactcgctgcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggta atacggttatccacagaatcaggggataacgcaggaaagaacatgtgagcaaaaggccagcaaaaggcca ggaaccgtaaaaaggccgcgttgctggcgtttttccataggctccgcccccctgacgagcatcacaaaaa tcgacgctcaagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctggaagc tccctcgtgcgctctcctgttccgaccctgccgcttaccggatacctgtccgcctttctcccttcgggaa gcgtggcgctttctcaatgctcacgctgtaggtatctcagttcggtgtaggtcgttcgctccaagctggg ctgtgtgcacgaaccccccgttcagcccgaccgctgcgccttatccggtaactatcgtcttgagtccaac ccggtaagacacgacttatcgccactggcagcagccactggtaacaggattagcagagcgaggtatgtag gcggtgctacagagttcttgaagtggtggcctaactacggctacactagaaggacagtatttggtatctg cgctctgctgaagccagttaccttcggaaaaagagttggtagctcttgatccggcaaacaaaccaccgct ggtagcggtggtttttttgtttgcaagcagcagattacgcgcagaaaaaaaggatctcaagaagatcctt tgatcttttctacggggtctgacgctcagtggaacgaaaactcacgttaagggattttggtcatgagatt atcaaaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaatcaatctaaagtatatat gagtaaacttggtctgacagttaccaatgcttaatcagtgaggcacctatctcagcgatctgtctatttc gttcatccatagttgcctgactccccgtcgtgtagataactacgatacgggagggcttaccatctggccc cagtgctgcaatgataccgcgagacccacgctcaccggctccagatttatcagcaataaaccagccagcc ggaagggccgagcgcagaagtggtcctgcaactttatccgcctccatccagtctattaattgttgccggg aagctagagtaagtagttcgccagttaatagtttgcgcaacgttgttgccattgctacaggcatcgtggt gtcacgctcgtcgtttggtatggcttcattcagctccggttcccaacgatcaaggcgagttacatgatcc cccatgttgtgcaaaaaagcggttagctccttcggtcctccgatcgttgtcagaagtaagttggccgcag tgttatcactcatggttatggcagcactgcataattctcttactgtcatgccatccgtaagatgcttttc tgtgactggtgagtactcaaccaagtcattctgagaatagtgtatgcggcgaccgagttgctcttgcccg gcgtcaatacgggataataccgcgccacatagcagaactttaaaagtgctcatcattggaaaacgttctt cggggcgaaaactctcaaggatcttaccgctgttgagatccagttcgatgtaacccactcgtgcacccaa ctgatcttcagcatcttttactttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgca aaaaagggaataagggcgacacggaaatgttgaatactcatactcttcctttttcaatattattgaagca tttatcagggttattgtctcatgagcggatacatatttgaatgtatttagaaaaataaacaaataggggt tccgcgcacatttccccgaaaagtgccacctgacgtctaagaaaccattattatcatgacattaacctat aaaaataggcgtatcacgaggccctttcgtctcgcgcgtttcggtgatgacggtgaaaacctctgacaca tgcagctcccggagacggtcacagcttgtctgtaagcggatgccgggagcagacaagcccgtcagggcgc gtcagcgggtgttggcgggtgtcggggctggcttaactatgcggcatcagagcagattgtactgagagtg caccatatggacatattgtcgttagaacgcggctacaattaatacataaccttatgtatcatacacatac gatttaggtgacactatagaactcgagcagctgaagcttgcatgcctgcaggtcgactctagaggatctc tgacctgagattggcggcactgaggtagagatgcccgaacccccccgagggagcgcgggacgcgccgggg agggctggggccggggagggctggggccggggagggctggggccggggagggctggggccggggagggct ggggccggggagggctggggccggggagggctggggctggggagggctggggctggggagggggcggtgg tgtgtagcaggagcggtgtgttgcgccggggtacgtctggaggagcgggaggtgcgcggtgacgtgtgga tgaggaacaggagttgttgcgcggtgagttgtcgctgtgagttgtgttgttgggcaggtgtggtggatga cgtgacgtgtgacgtgcggagtgcgccgtgctctgttggtttcacctgtggcagcccgggccccccgcgg gcgcgcgcgcgcgcaaaaaaggcgggcggcggtccgggcggcgtgcgcgcgcgcggcgggcgtggggggc ggggccgcgggagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcggggggag gagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcgggggg aggagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcgggg ggaggagcggggggaggagcggccagacgccgaaaacgggccccccccaaaacacaccccccgggggtcg cgcgcggccctttaaagcggtggcggcgggcagcccgggccccccgcggccgagactagcgagttagaca ggcaagcactactcgcctctgcacgcacatgcttgcctgtcaaactctaccaccccggcacgctctctgt ctccatggcccgccgccgccgccatcgcggcccccgccgcccccggccgcccgggcccacgggcgccgtc ccaaccgcacagtcccaggtaacctccacgcccaactcggaacccgcggtcaggagcgcgcccgcggccg cagcaacctggttctttccgcctcagaagccatagagcccaccgcatccccagcatgcctgctattgtct tcccaatcctcccccttgctgtcctgccccaccccaccccccagaatagaatgacacctactcagacaat gcgatgcaatttcctcattttattaggaaaggacagtgggagtggcaccttccagggtcaaggaaggcac gggggaggggcaaacaacagatggctggcaactagaaggcacagtcgaggctgatcagcgagctctagca tttaggtgacactatagaatagggccctctagctcgagcaattgcacgtgatggatccagcttgggtctc cctatagtgagtcgtattaatttcgataagccagtaagcagtgggttctctagttagccagagagctctg cttatatagacctcccaccgtacacgcctaccgcccatttgcgtcaatggggcggagttgttacgacatt ttggaaagtcccgttgattttggtgccaaaacaaactcccattgacgtcaatggggtggagacttggaaa tccccgtgagtcaaaccgctatccacgcccattgatgtactgccaaaaccgcatcaccatggtaatagcg atgactaatacgtagatgtactgccaagtaggaaagtcccataaggtcatgtactgggcataatgccagg cgggccatttaccgtcattgacgtcaatagggggcgtacttggcatatgatacacttgatgtactgccaa gtgggcagtttaccgtaaatagtccacccattgacgtcaatggaaagtccctattggcgttactatggga acatacgtcattattgacgtcaatgggcgggggtcgttgggcggtcagccaggcgggccatttaccgtaa gttatgtaacgcggaactccatatatgggctatgaactaatgaccccgtaattgattactattaataact agtcaataatcaatgtcaacgcgtatatctggcccgtacatcgcgatgtacgggccagatatacgcgtat ctgaggggactagggtgtgtttaggcgaaaagcggggcttcggttgtacgcggttaggagtcccctcagg atatagtagtttcgcttttgcatagggagggggaaatgtagtcttatgcaatacacttgtagtcttgcaa catggtaacgatgagttagcaacatgccttacaaggagagaaaaagcaccgtgcatgccgattggtggaa gtaaggtggtacgatcgtgccttattaggaaggcaacagacaggtctgacatggattggacgaaccactg aattccgcattgcagagataattgtatttaagtgcctagctcgatacaataaacgccatttgaccattca ccacattggtgtgcacctccaagcttgcggccgcgtttAAACGCCACCATGGACTGGACCTGGATTCTGT TCCTGGTGGCAGCAGCAACAAGGGTGCACAGCAACTGGGTGAATGTGATCTCCGACCTGAAGAAGATCGA GGATCTGATCCAGAGCATGCACATCGACGCCACCCTGTACACAGAGTCCGATGTGCACCCCTCTTGCAAG GTGACCGCCATGAAGTGTTTTCTGCTGGAGCTGCAGGTCATCTCTCTGGAGAGCGGCGACGCCTCCATCC ACGATACCGTGGAGAACCTGATCATCCTGGCCAACAATTCTCTGAGCTCCAACGGCAATGTGACAGAGAG CGGCTGCAAGGAGTGTGAGGAGCTGGAGAAGAAGAACATCAAGGAGTTCCTGCAGTCCTTTGTGCACATC GTGCAGATGTTCATCAATACATCTTAACCAACCGCtgtttggtctgctttctgacaaactcggaacttgt ttattgcagcttataatggttacaaataaagcaatagcatcacaaatttcacaaataaagcatttttttc actgcattctagttgtggtttgtgtcgacttagatc [SEQ ID No: 17] Alternatively, in another embodiment, the nucleotide sequence encoding the shuttle vector (known as p-34.5 DC RSV CMV S-FLiC-M3) used to produce the oncolytic virus according to the invention, is provided herein as SEQ ID No: 18, as follows: gctgcggccgccagcgcggcggggcccggccaaccagcgtccgccgagtcttcggggcccggcccattgg gcgggagttaccgcccaatgggccgggccgcccacttcccggtatggtaattaaaaacttgcaagaggcc ttgttccgcttcccggtatggtaattagaaactcattaatgggcggccccggccgcccttcccgcttccg gcaattcccgcggcccttaatgggcaaccccggtattccccgcctcccgcgccgcgcgtaaccactcccc tggggttccgggttatgctaattgcttttttggcggaacacacggcccctcgcgcattggcccgcgggtc gctcaatgaacccgcattggtcccctggggttccgggtatggtaatgagtttcttcgggaaggcgggaag ccccggggcaccgacgcaggccaagcccctgttgcgtcggcgggaggggcatgctaatggggttctttgg gggacaccgggttgggcccccaaatcgggggccgggccgtgcatgctaatgatattctttgggggcgccg ggttggtccccggggacggggccgccccgcggtgggcctgcctcccctgggacgcgcggccattggggga atcgtcactgccgcccctttggggaggggaaaggcgtggggtataagttagccctggcccgacagtctgg tcgcatttgcacctcggcactcggagcgagacgcagcagccaggcagactcgggccgccccctctccgca tcaccacagaagccccgcctacgttgcgacccccagggaccctccgtccgcgaccctccagccgcatacg acccccatggagccccgccccggagcgagtacccgccggcctgagggccgcccccagcgcgaggtgaggg gccgggcgccatgtctggggcgccatattggggggcgccatattggggggcgccatgttgggggaccccc gacccttacactggaaccggccgccatgttgggggacccccactcatacacgggagccgggcgccatgtt ggggcgccatgttagggggcgtggaaccccgtgacactatatatacagggaccgggggcgccatgttagg gggtgcggaaccccctgaccctatatatacagggaccggggtcgccctgttgggggtcgccatgtgaccc cctgactttatatatacagacccccaacacatacacatggcccctttgactcagacgcagggcccggggt cgccgtgggaccccctgactcatacacagagacacgcccccacaacaaacacacaaggaccggggtcgcc gtgttgggggcgtggtccccactgactcatacgcaggccccccttactcacacgcatctaggggggtggg gaggagccgcccgccatatttgggggacgccgtgggacccccgactccggtgcgtctggagggcgggaga agagggaagaagaggggtcgggatctgccggtctccctatagtgagtcgtattaatttcgataagccagg ttaacctgcattaatgaatcggccaacgcgcggggagaggcggtttgcgtattgggcgctcttccgcttc ctcgctcactgactcgctgcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggta atacggttatccacagaatcaggggataacgcaggaaagaacatgtgagcaaaaggccagcaaaaggcca ggaaccgtaaaaaggccgcgttgctggcgtttttccataggctccgcccccctgacgagcatcacaaaaa tcgacgctcaagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctggaagc tccctcgtgcgctctcctgttccgaccctgccgcttaccggatacctgtccgcctttctcccttcgggaa gcgtggcgctttctcaatgctcacgctgtaggtatctcagttcggtgtaggtcgttcgctccaagctggg ctgtgtgcacgaaccccccgttcagcccgaccgctgcgccttatccggtaactatcgtcttgagtccaac ccggtaagacacgacttatcgccactggcagcagccactggtaacaggattagcagagcgaggtatgtag gcggtgctacagagttcttgaagtggtggcctaactacggctacactagaaggacagtatttggtatctg cgctctgctgaagccagttaccttcggaaaaagagttggtagctcttgatccggcaaacaaaccaccgct ggtagcggtggtttttttgtttgcaagcagcagattacgcgcagaaaaaaaggatctcaagaagatcctt tgatcttttctacggggtctgacgctcagtggaacgaaaactcacgttaagggattttggtcatgagatt atcaaaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaatcaatctaaagtatatat gagtaaacttggtctgacagttaccaatgcttaatcagtgaggcacctatctcagcgatctgtctatttc gttcatccatagttgcctgactccccgtcgtgtagataactacgatacgggagggcttaccatctggccc cagtgctgcaatgataccgcgagacccacgctcaccggctccagatttatcagcaataaaccagccagcc ggaagggccgagcgcagaagtggtcctgcaactttatccgcctccatccagtctattaattgttgccggg aagctagagtaagtagttcgccagttaatagtttgcgcaacgttgttgccattgctacaggcatcgtggt gtcacgctcgtcgtttggtatggcttcattcagctccggttcccaacgatcaaggcgagttacatgatcc cccatgttgtgcaaaaaagcggttagctccttcggtcctccgatcgttgtcagaagtaagttggccgcag tgttatcactcatggttatggcagcactgcataattctcttactgtcatgccatccgtaagatgcttttc tgtgactggtgagtactcaaccaagtcattctgagaatagtgtatgcggcgaccgagttgctcttgcccg gcgtcaatacgggataataccgcgccacatagcagaactttaaaagtgctcatcattggaaaacgttctt cggggcgaaaactctcaaggatcttaccgctgttgagatccagttcgatgtaacccactcgtgcacccaa ctgatcttcagcatcttttactttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgca aaaaagggaataagggcgacacggaaatgttgaatactcatactcttcctttttcaatattattgaagca tttatcagggttattgtctcatgagcggatacatatttgaatgtatttagaaaaataaacaaataggggt tccgcgcacatttccccgaaaagtgccacctgacgtctaagaaaccattattatcatgacattaacctat aaaaataggcgtatcacgaggccctttcgtctcgcgcgtttcggtgatgacggtgaaaacctctgacaca tgcagctcccggagacggtcacagcttgtctgtaagcggatgccgggagcagacaagcccgtcagggcgc gtcagcgggtgttggcgggtgtcggggctggcttaactatgcggcatcagagcagattgtactgagagtg caccatatggacatattgtcgttagaacgcggctacaattaatacataaccttatgtatcatacacatac gatttaggtgacactatagaactcgagcagctgaagcttgcatgcctgcaggtcgactctagaggatctc tgacctgagattggcggcactgaggtagagatgcccgaacccccccgagggagcgcgggacgcgccgggg agggctggggccggggagggctggggccggggagggctggggccggggagggctggggccggggagggct ggggccggggagggctggggccggggagggctggggctggggagggctggggctggggagggggcggtgg tgtgtagcaggagcggtgtgttgcgccggggtacgtctggaggagcgggaggtgcgcggtgacgtgtgga tgaggaacaggagttgttgcgcggtgagttgtcgctgtgagttgtgttgttgggcaggtgtggtggatga cgtgacgtgtgacgtgcggagtgcgccgtgctctgttggtttcacctgtggcagcccgggccccccgcgg gcgcgcgcgcgcgcaaaaaaggcgggcggcggtccgggcggcgtgcgcgcgcgcggcgggcgtggggggc ggggccgcgggagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcggggggag gagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcgggggg aggagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcggggggaggagcgggg ggaggagcggggggaggagcggccagacgccgaaaacgggccccccccaaaacacaccccccgggggtcg cgcgcggccctttaaagcggtggcggcgggcagcccgggccccccgcggccgagactagcgagttagaca ggcaagcactactcgcctctgcacgcacatgcttgcctgtcaaactctaccaccccggcacgctctctgt ctccatggcccgccgccgccgccatcgcggcccccgccgcccccggccgcccgggcccacgggcgccgtc ccaaccgcacagtcccaggtaacctccacgcccaactcggaacccgcggtcaggagcgcgcccgcggccg cagcaacctggttctttccgcctcagaagccatagagcccaccgcatccccagcatgcctgctattgtct tcccaatcctcccccttgctgtcctgccccaccccaccccccagaatagaatgacacctactcagacaat gcgatgcaatttcctcattttattaggaaaggacagtgggagtggcaccttccagggtcaaggaaggcac gggggaggggcaaacaacagatggctggcaactagaaggcacagtcgaggctgatcagcgagctctagca tttaggtgacactatagaatagggccctctagctcgagcAATTGTTACTGCAGCAGTTTCAGGACCTGCT GTGGCAGACTATTAGCCTGTGCCAGGACGGAGATCCCGGCCTGCTGCAGCACCTGGGCTCTGCTCAGCTG GGCTGTCTCCTGGGCAAAATCGGCGCTCTGGATCTGGCTCTGGGCCTGGGCCAGATTGTTGGAGCCGGCC TGCTGGGTTGTGGCGATGGCGGTAAACCGGTTCTGGGCGGCGCCCAGTGTGGCCTGCAGATTGTTCACGG TGGCCAGGGCGTTGTCGATGGACACCATGGCCTGATAGGCGCCGGTCTGGGTGCTGATATCCAGGTTGGA CACTGTCTGGGGCTTGTTCACGGCATTGATCTGGGCCACCATGGCCGTGGCCTGGGCCTGATCTGTGGCG GACAGGGCAGATGTGGCAGATGTGGACAGGGCCAGGGTCTGAAAGCTAGGGGAGGCGGCTGTGCCGGTGC CGGCTGTAGAGGAGCCAAACACGGCGGTCACGGCTGTGCTGGACAGGGCCTGATTGTTCTGGTCTGTAAA GGTGAAGCCGCCCTTGCCGTCGGACACCACGTTGATCTGGGTTGTCTCCTGGCCGGTGGAGCCGCTAGAC CAGGCGGCGCCGGAGCTATCGATGGCCACCTTGATTGTGCCCAGGGTCTGGCCTGTCTGCACCATGCCGC CGCCGATCTTGGCGGCGCTCATAGACTGGGTCAGATCCACAGACACGGTCTGGCCCACGTTGGCGCCCAC CTGAAAGGACAGGGTGCCGGCAGAGCCATCCAGGATGTTCTTGCCATTGTAGTTGGTCTGGCTGGCGATT CTGTTCACCTCAGAGATCTGCTGGGCCACCTCCTGCTGCAGGGCGCTGGCATCGGAGGCAGACAGAGGGC CGTTGCTGGCCTGCACGGCCAGCTGTCTGATCCTCTGCAGGCTGTTGGTCAGAGATGTCAGGCCGCTGGA GGCGGTCTGCAGGATGCTCACGCCATCATTGGCGTTAGACACGCCCTGGTTCAGGCCATTGATCTGGGTC TGCATTCTGGTGGCGATGGCCAGGCCGGCGGCATCGTCGGCGGCGGAATTGATCCGCTTGCCAGAGGACA GTCTGGTGATGGCCTGAGACAGGGCGCCCTGGCTGCCCTGCAGATTCTGCTGAGCGACCAGAGAGTTGAT GTTACTATTGATTCCCAGGGCCAGAGCCCCAGCCCACAGCAGAGGCAGCAGCAGCAGCAGAGGCATGGTG GCGgatccagcttgggtctccctatagtgagtcgtattaatttcgataagccagtaagcagtgggttctc tagttagccagagagctctgcttatatagacctcccaccgtacacgcctaccgcccatttgcgtcaatgg ggcggagttgttacgacattttggaaagtcccgttgattttggtgccaaaacaaactcccattgacgtca atggggtggagacttggaaatccccgtgagtcaaaccgctatccacgcccattgatgtactgccaaaacc gcatcaccatggtaatagcgatgactaatacgtagatgtactgccaagtaggaaagtcccataaggtcat gtactgggcataatgccaggcgggccatttaccgtcattgacgtcaatagggggcgtacttggcatatga tacacttgatgtactgccaagtgggcagtttaccgtaaatagtccacccattgacgtcaatggaaagtcc ctattggcgttactatgggaacatacgtcattattgacgtcaatgggcgggggtcgttgggcggtcagcc aggcgggccatttaccgtaagttatgtaacgcggaactccatatatgggctatgaactaatgaccccgta attgattactattaataactagtcaataatcaatgtcaacgcgtatatctggcccgtacatcgcgatgta cgggccagatatacgcgtatctgaggggactagggtgtgtttaggcgaaaagcggggcttcggttgtacg cggttaggagtcccctcaggatatagtagtttcgcttttgcatagggagggggaaatgtagtcttatgca atacacttgtagtcttgcaacatggtaacgatgagttagcaacatgccttacaaggagagaaaaagcacc gtgcatgccgattggtggaagtaaggtggtacgatcgtgccttattaggaaggcaacagacaggtctgac atggattggacgaaccactgaattccgcattgcagagataattgtatttaagtgcctagctcgatacaat aaacgccatttgaccattcaccacattggtgtgcacctccaagcttgcggccgcgtttaaaccctgcagg tctagataataacgggcaggccatgtctgcccgtatttcgcgtaaggaaatccattatgtactatttaaa aaacacaaacttttggatgttcggtttattctttttcttttacttttttatcatgggagcctacttcccg tttttcccgatttggctacatgacatcaaccatatcagcaaaagtgatacgggtattatttttgccgcta tttctctgttctcgctattattccaaccgctgtttggtctgctttctgacaaactcggaacttgtttatt gcagcttataatggttacaaataaagcaatagcatcacaaatttcacaaataaagcatttttttcactgc attctagttgtggtttgtgtcgacttagatc [SEQ ID No: 18] Hence, in a preferred embodiment, the shuttle vector used to produce the oncolytic virus according to the invention is encoded by a nucleotide sequence as set out in SEQ ID No: 11, 1617, or 18, or a fragment or variant thereof. The oncolytic viruses according to the third and fourth aspects of the invention, are particularly suitable for therapy against cancer, preferably bladder cancer. Hence, according to a fifth aspect of the invention, there is provided the genetic construct according to the first aspect, the expression cassette according to the second aspect, or the oncolytic virus according to the third or fourth aspect, for use in therapy. According to a sixth aspect, there is provided the genetic construct according to the first aspect, the expression cassette according to the second aspect, or the oncolytic virus according to the third or fourth aspect, for use in treating, preventing or ameliorating cancer. According to a seventh aspect, there is provided a method of treating, preventing or ameliorating a cancer in a subject, the method comprising administering, or having administered, to a subject in need of such treatment, a therapeutically effective amount of the genetic construct according to the first aspect, the expression cassette according to the second aspect, or the oncolytic virus according to the third or fourth aspect. The cancer may be bladder cancer. In a preferred embodiment, the cancer is bladder cancer. More preferably, the cancer is non-muscle invasive bladder cancer (NMIBC). Preferably, the bladder cancer is BCG- refractory disease bladder cancer. Advantageously, the genetic construct, expression cassette and oncolytic virus according to the invention, facilitate the direct destruction of cancer cells and promote the release of therapeutic proteins, such as IL-15 and FliC_BP along with tumour antigens. This release is essential for stimulating both innate and adaptive immune responses against tumour antigens and viral proteins leading to sustained anti-tumour immunity. Accordingly, in some embodiments, the genetic construct, expression cassette and oncolytic virus according to the invention, stimulate the innate and adaptive immune response. In some embodiments, the genetic construct, expression cassette and oncolytic virus according to the invention stimulate TLR2- and TLR-5 mediated signalling pathways. It will be appreciated that the oncolytic virus according to the invention, may be used in a medicament, which may be used as a monotherapy (i.e. use of the oncolytic virus alone), for treatment against cancer. Alternatively, the oncolytic virus according to the invention may be used as an adjunct to, or in combination with, known therapies for treating, ameliorating, or preventing cancer. The oncolytic virus of the invention may be combined in compositions having a number of different forms depending, in particular, on the manner in which the composition is to be used. Thus, for example, the composition may be in the form of a powder, tablet, capsule, liquid, ointment, cream, gel, hydrogel, aerosol, spray, micellar solution, transdermal patch, liposome suspension, polyplex, emulsion, lipid nanoparticles (e.g. with peptide, DNA or RNA on the surface or encapsulated) or any other suitable form that may be administered to a person or animal in need of vaccination. The lipid nanoparticle may comprise one or more components selected from a group consisting of: a cationic lipid (which is preferably ionisable); phosphatidylcholine; cholesterol; and polyethylene glycol (PEG)-lipid. It will be appreciated that the vehicle of medicaments according to the invention should be one which is well-tolerated by the subject to whom it is given. Medicaments comprising the oncolytic virus of the invention may be used in a number of ways. For instance, oral administration may be required, in which case the agents may be contained within a composition that may, for example, be ingested orally in the form of a tablet, capsule or liquid. Compositions comprising agents and medicaments of the invention may be administered by inhalation (e.g. intranasally). Compositions may also be formulated for topical use. For instance, creams or ointments may be applied to the skin. The oncolytic virus of the invention may also be incorporated within a slow- or delayed- release device. Such devices may, for example, be inserted on or under the skin, and the medicament may be released over weeks or even months. The device may be located at least adjacent the treatment site. Such devices may be particularly advantageous when long-term treatment with the oncolytic virus is required, and which would normally require frequent administration (e.g. at least daily injection). In a preferred embodiment, however, medicaments according to the invention may be administered to a subject by injection into the blood stream, muscle, skin or directly into a site requiring treatment. Injections may be intravenous (bolus or infusion) or subcutaneous (bolus or infusion), or intradermal (bolus or infusion), or intramuscular (bolus or infusion). In some embodiments, injections are intravenous. Typically, injections are intravesical or intratumoral. For intravesical injections, these may be applied into the bladder by catheter. It will be appreciated that the amount of oncolytic virus that is required is determined by its biological activity and bioavailability, which in turn depends on the mode of administration, the physiochemical properties of the oncolytic virus and whether it is being used as a monotherapy or in a combined therapy. The frequency of administration will also be influenced by the half-life of the active agent within the subject being treated. Optimal dosages to be administered may be determined by those skilled in the art, and will vary with the particular the oncolytic virus in use, the strength of the composition, the mode of administration, and the type and advancement of the cancer. Additional factors depending on the particular subject being treated will result in a need to adjust dosages, including subject age, weight, gender, diet, and time of administration. Generally, a daily dose of between 0.001µg / kg of body weight and 100mg / kg of body weight of the oncolytic virus of the invention may be used for the immunisation, depending upon the agent used. More preferably, the daily dose of agent is between 1 ^g / kg of body weight and 100mg / kg of body weight, more preferably between 10 ^g / kg and ^ ^mg / kg body weight, and most preferably between approximately 100 ^g / kg and 10mg / kg body weight. In some embodiments, a dose of between 10E4 plaque forming units (PFU) / mL and 10E9 PFU / mL of the oncolytic virus of the invention may be used for the immunisation. Daily doses may be given as a single administration (e.g. a single daily injection). Alternatively, the oncolytic virus may require administration twice or more times during a day. As an example, the oncolytic virus may be administered as an initial primer and a subsequent boost(s), or two boosts administered at between a week or monthly intervals. Preferably, the oncolytic virus may be administered as an initial primer and a subsequent boost, administered between two to six weeks apart. Known procedures, such as those conventionally employed by the pharmaceutical industry (e.g. in vivo experimentation, clinical trials, etc.), may be used to form specific formulations of the oncolytic virus according to the invention and precise therapeutic regimes (such as daily doses of the agents and the frequency of administration). The inventors believe that they are the first to suggest an oncolytic virus comprising a gene encoding a TLR agonist and a gene encoding a cytokine, or an oncolytic virus comprising a gene encoding FliC_BP. According to an eighth aspect of the invention, there is provided a pharmaceutical composition comprising the genetic construct according to the first aspect, the expression cassette according to the second aspect, or the oncolytic virus according to the third or fourth aspect, and a pharmaceutically acceptable vehicle. The invention also provides in a ninth aspect, a process for making the pharmaceutical composition according to the eighth aspect, the process comprising combining a therapeutically effective amount of the genetic construct according to the first aspect, the expression cassette according to the second aspect, or the oncolytic virus according to the third or fourth aspect, with a pharmaceutically acceptable vehicle. A “subject” may be a vertebrate, mammal, or domestic animal. Hence, compositions and medicaments according to the invention may be used to treat any mammal, for example livestock (e.g. a horse), pets, or may be used in other veterinary applications. Most preferably, however, the subject is a human being. A “therapeutically effective amount” of the oncolytic virus is any amount which, when administered to a subject, is the amount of the aforementioned that is needed to ameliorate, prevent or treat any given disease, preferably prophylactically. For example, the oncolytic virus of the invention may be used from about 0.001 µg to about 1 mg, and preferably from about 0.001 µg to about 500 µg. It is preferred that the amount of the oncolytic virus is an amount from about 0.01 µg to about 250 µg, and most preferably from about 0.1 µg to about 100 µg. Preferably, the oncolytic virus according to the invention is administered at a dose of 1-50 ^g. The oncolytic virus of the invention may further comprise a pharmaceutically acceptable vehicle. A “pharmaceutically acceptable vehicle” as referred to herein, is any known compound or combination of known compounds that are known to those skilled in the art to be useful in formulating pharmaceutical compositions. In one embodiment, the pharmaceutically acceptable vehicle may be a solid, and the composition may be in the form of a powder or tablet. A solid pharmaceutically acceptable vehicle may include one or more substances which may also act as flavouring agents, lubricants, solubilisers, suspending agents, dyes, fillers, glidants, compression aids, inert binders, sweeteners, preservatives, dyes, coatings, or tablet- disintegrating agents. The vehicle may also be an encapsulating material. In powders, the vehicle is a finely divided solid that is in admixture with the finely divided active agents according to the invention. In tablets, the active agent (e.g. oncolytic virus according to the invention) may be mixed with a vehicle having the necessary compression properties in suitable proportions and compacted in the shape and size desired. The powders and tablets preferably contain up to 99% of the active agents. Suitable solid vehicles include, for example calcium phosphate, magnesium stearate, talc, sugars, lactose, dextrin, starch, gelatin, cellulose, polyvinylpyrrolidine, low melting waxes and ion exchange resins. In another embodiment, the pharmaceutical vehicle may be a gel and the composition may be in the form of a cream or the like. In another embodiment, the oncolytic virus or pharmaceutical composition may be lyophilised. However, the pharmaceutical vehicle may be a liquid, and the pharmaceutical composition is in the form of a solution. Liquid vehicles are used in preparing solutions, suspensions, emulsions, syrups, elixirs and pressurized compositions. The oncolytic virus according to the invention may be dissolved or suspended in a pharmaceutically acceptable liquid vehicle such as water, an organic solvent, a mixture of both or pharmaceutically acceptable oils or fats. The liquid vehicle can contain other suitable pharmaceutical additives such as solubilisers, emulsifiers, buffers, preservatives, sweeteners, flavouring agents, suspending agents, thickening agents, colours, viscosity regulators, stabilizers or osmo-regulators. Suitable examples of liquid vehicles for oral and parenteral administration include water (partially containing additives as above, e.g. cellulose derivatives, preferably sodium carboxymethyl cellulose solution), alcohols (including monohydric alcohols and polyhydric alcohols, e.g. glycols) and their derivatives, and oils (e.g. fractionated coconut oil and arachis oil). For parenteral administration, the vehicle can also be an oily ester such as ethyl oleate and isopropyl myristate. Sterile liquid vehicles are useful in sterile liquid form compositions for parenteral administration. The liquid vehicle for pressurized compositions can be a halogenated hydrocarbon or other pharmaceutically acceptable propellant. Liquid pharmaceutical compositions, which are sterile solutions or suspensions, can be utilized by, for example, subcutaneous, intradermal, intrathecal, epidural, intraperitoneal, intravenous and particularly intramuscular injection, or urinary catheterization via urethral catheter. The nucleic acid sequence, or expression cassette of the invention may be prepared as a sterile solid composition that may be dissolved or suspended at the time of administration using sterile water, saline, or other appropriate sterile injectable medium. The oncolytic virus of the invention may be administered orally in the form of a sterile solution or suspension containing other solutes or suspending agents (for example, enough saline or glucose to make the solution isotonic), bile salts, acacia, gelatin, sorbitan monoleate, polysorbate 80 (oleate esters of sorbitol and its anhydrides copolymerized with ethylene oxide) and the like. The oncolytic virus according to the invention can also be administered orally either in liquid or solid composition form. Compositions suitable for oral administration include solid forms, such as pills, capsules, granules, tablets, and powders, and liquid forms, such as solutions, syrups, elixirs, and suspensions. Forms useful for parenteral administration include sterile solutions, emulsions, and suspensions. The pharmaceutical composition may be frozen, freeze dried, or lyophilised. It will be appreciated that the invention extends to any nucleic acid or peptide or variant, derivative or analogue thereof, which comprises substantially the amino acid or nucleic acid sequences of any of the sequences referred to herein, including variants or fragments thereof. The terms “substantially the amino acid / nucleotide / peptide sequence”, “variant” and “fragment”, can be a sequence that has at least 40% sequence identity with the amino acid / nucleotide / peptide sequences of any one of the sequences referred to herein, for example 40% identity with the sequence identified as SEQ ID Nos: 1-20 and so on. Amino acid / polynucleotide / polypeptide sequences with a sequence identity which is greater than 65%, more preferably greater than 70%, even more preferably greater than 75%, and still more preferably greater than 80% sequence identity to any of the sequences referred to are also envisaged. Preferably, the amino acid / polynucleotide / polypeptide sequence has at least 85% identity with any of the sequences referred to, more preferably at least 90% identity, even more preferably at least 92% identity, even more preferably at least 95% identity, even more preferably at least 97% identity, even more preferably at least 98% identity and, most preferably at least 99% identity with any of the sequences referred to herein. The skilled technician will appreciate how to calculate the percentage identity between two amino acid / polynucleotide / polypeptide sequences. In order to calculate the percentage identity between two amino acid / polynucleotide / polypeptide sequences, an alignment of the two sequences must first be prepared, followed by calculation of the sequence identity value. The percentage identity for two sequences may take different values depending on:- (i) the method used to align the sequences, for example, ClustalW, BLAST, FASTA, Smith-Waterman (implemented in different programs), or structural alignment from 3D comparison; and (ii) the parameters used by the alignment method, for example, local vs global alignment, the pair-score matrix used (e.g. BLOSUM62, PAM250, Gonnet etc.), and gap-penalty, e.g. functional form and constants. Having made the alignment, there are many different ways of calculating percentage identity between the two sequences. For example, one may divide the number of identities by: (i) the length of shortest sequence; (ii) the length of alignment; (iii) the mean length of sequence; (iv) the number of non-gap positions; or (v) the number of equivalenced positions excluding overhangs. Furthermore, it will be appreciated that percentage identity is also strongly length dependent. Therefore, the shorter a pair of sequences is, the higher the sequence identity one may expect to occur by chance. Hence, it will be appreciated that the accurate alignment of protein or DNA sequences is a complex process. The popular multiple alignment program ClustalW (Thompson et al., 1994, Nucleic Acids Research, 22, 4673-4680; Thompson et al., 1997, Nucleic Acids Research, 24, 4876-4882) is a preferred way for generating multiple alignments of proteins or DNA in accordance with the invention. Suitable parameters for ClustalW may be as follows: For DNA alignments: Gap Open Penalty = 15.0, Gap Extension Penalty = 6.66, and Matrix = Identity. For protein alignments: Gap Open Penalty = 10.0, Gap Extension Penalty = 0.2, and Matrix = Gonnet. For DNA and Protein alignments: ENDGAP = -1, and GAPDIST = 4. Those skilled in the art will be aware that it may be necessary to vary these and other parameters for optimal sequence alignment. Preferably, calculation of percentage identities between two amino acid / polynucleotide / polypeptide sequences may then be calculated from such an alignment as (N / T)*100, where N is the number of positions at which the sequences share an identical residue, and T is the total number of positions compared including gaps and either including or excluding overhangs. Preferably, overhangs are included in the calculation. Hence, a most preferred method for calculating percentage identity between two sequences comprises (i) preparing a sequence alignment using the ClustalW program using a suitable set of parameters, for example, as set out above; and (ii) inserting the values of N and T into the following formula:- Sequence Identity = (N / T)*100. Alternative methods for identifying similar sequences will be known to those skilled in the art. For example, a substantially similar nucleotide sequence will be encoded by a sequence which hybridizes to DNA sequences or their complements under stringent conditions. By stringent conditions, the inventors mean the nucleotide hybridises to filter-bound DNA or RNA in 3x sodium chloride / sodium citrate (SSC) at approximately 45ºC followed by at least one wash in 0.2x SSC / 0.1% SDS at approximately 20-65ºC. Alternatively, a substantially similar polypeptide may differ by at least 1, but less than 5, 10, 20, 50 or 100 amino acids from the sequences shown in, for example, in those of SEQ ID Nos: 1 to 20 that are amino acid sequences. Due to the degeneracy of the genetic code, it is clear that any nucleic acid sequence described herein could be varied or changed without substantially affecting the sequence of the protein encoded thereby, to provide a functional variant thereof. Suitable nucleotide variants are those having a sequence altered by the substitution of different codons that encode the same amino acid within the sequence, thus producing a silent (synonymous) change. Other suitable variants are those having homologous nucleotide sequences but comprising all, or portions of, sequence, which are altered by the substitution of different codons that encode an amino acid with a side chain of similar biophysical properties to the amino acid it substitutes, to produce a conservative change. For example, small non-polar, hydrophobic amino acids include glycine, alanine, leucine, isoleucine, valine, proline, and methionine. Large non-polar, hydrophobic amino acids include phenylalanine, tryptophan and tyrosine. The polar neutral amino acids include serine, threonine, cysteine, asparagine and glutamine. The positively charged (basic) amino acids include lysine, arginine and histidine. The negatively charged (acidic) amino acids include aspartic acid and glutamic acid. It will therefore be appreciated which amino acids may be replaced with an amino acid having similar biophysical properties, and the skilled technician will know the nucleotide sequences encoding these amino acids. All of the features described herein (including any accompanying claims, abstract and drawings), and / or all of the steps of any method or process so disclosed, may be combined with any of the above aspects in any combination, except combinations where at least some of such features and / or steps are mutually exclusive. For a better understanding of the invention, and to show how embodiments of the same may be carried into effect, reference will now be made, by way of example, to the accompanying Figures, in which:- Figure 1 shows a schematic representation of one embodiment of the oncolytic virus according to the invention. The oncolytic Herpes Simplex virus (oHSV-FliC_BP-IL-15) was derived from wild type (WT) HSV-1 strain 17+, and has two gene deletions, i.e. that of the genes encoding (i) ICP34.5 and (ii) ICP47. oHSV-FliC_BP-IL-15 expresses the Burkholderia pseudomallei flagellin protein (Genbank ABA48561.1, a TLR5 ligand) under the CMV promoter, and IL-15 (GenBank: AF031167.1) under the RSV promoter. Figure 2 illustrates the expression of TLR Ligands and IL-15 from plasmids and oHSV viruses. TLR ligand plasmids, or oHSV viruses were screened using TLR receptor / NF- kB reporter assays: a) TLR4 & 5 (Promega); b) TLR2 ligand plasmids (Invivogen); c) oHSV-FliC_BP (Promega). d) Concentration of IL-15 detected using IL-15 cell bioassay (Promega) in supernatant from 5637 cells infected for 48hrs with oHSV-IL-15 at MOI 1. e) A soluble ectodomain of human TLR5 - TLR5 inhibitor (α Hu TLR5-Fc, InvivoGen) to the TLR5 receptor, was used to partly block the TLR5-MyD88-NF-kB pathway activated by supernatant from 5637 cells infected for 24hrs with oHSV-FliC_BP at MOI 1. f) Immuno-staining for flagellin bacteria protein from BP (green) carried out on 5637 cells infected with oHSV-FliC_BP virus at MOI 1 for 24hrs. g) THP-1 cells (+PMA) were treated oHSV or oHSV FLIC BP which had been UV inactive. The THP-1 were incubated for 24hrs and qPCR for innate cytokines was carried out. Figure 3 shows a) Average tumour volume of subcutaneous MB49 tumours treated with 108pfu of oHSV or oHSV-FliC_BP or oHSV-IL-15, or oHSV-FliC_BP-IL-15 (n=10) 4x time over 8 days. b) Kaplan-Meier survival curves. c) Rechallenge experiment with the same bladder cancer cell line (MB49) on naive and cured mice. e-g) FACS analysis measuring TNF-α in spherocytes from cured and naive mice, co-culture, alone or with MB49 tumours cells or PMA / Ionomycin. e) CD4 f) CD8 g) NK. Figure 4 illustrates the knockout of ICP47 from the HSV-1 genome. A plasmid (pUC57 del ICP47) was designed and synthesized, containing sequences of two HSV-1 flank regions on either side of ICP47. A PCR generated CMV EGFP pA cassette was generated from pcDNA3.1+ EGFP. The PCR product was cut with Bbs-1 and then inserted between flanking regions of ICP47 to produce pUC57 del ICP47 CMV EGFP pA. This viral shuttle vector was linearized (ssp-1) and added to HSV-1 infected cellular BHK DNA and transfected with Calcium Phosphate into BHK cells. The resulting homologous recombinant was plaque purified resulting in clones of HSV-117+ 47- EGFP. The CMV EGFP pA cassette was removed from the virus (HSV-117+ 47- EGFP) by homologous recombination using pUC57 del ICP47, which resulted in clones of HSV-117+ 47-. Figure 5 shows a plasmid map of p34.5 DC EGFP containing sequences of two HSV-1 flanking regions either side of ICP34.5. p34.5 DC EGFP contains a double expression cassette (RSV-pA, CMV pA). Under the CMV promoter is the synthetic green fluorescent gene (EGFP). Figure 6 shows a plasmid map of p34.5 CMV RSV IL-15ED containing sequences of two ICP34.5 flanking regions and IL-15 ED. p34.5 CMV RSV IL-15ED contains a double expression cassette (RSV-pA, CMV pA). Figure 7 shows a plasmid map of p34.5 CMV FLIC BP RSV IL-15ED, containing sequences of two ICP34.5 flanking regions, IL-15 ED and FLIC BP. p34.5 CMV FLIC BP RSV IL-15ED contains a double expression cassette (RSV-pA, CMV pA). Figure 8 shows the inactivation of HSV following exposure to 300 J of UV light. Figure 9 shows the viability of four TCC bladder cancer cell lines (T24, TCCSUP, Ku19-19 and VMCUB), treated with HSV / HSV5-15 with and without UV inactivation. Figure 10 shows TLR2 Stimulation determined using HEK-Blue™ hTLR-2 Detection: a) Plasmid DNA (containing FLiC BP ligand); and b) FLiC BP ligand expressed from Oncolytic HSV (HSV5). Examples The inventors designed and tested novel oncolytic viruses for treating non-muscle invasive bladder cancer (NMIBC). In view of the problems associated with BCG (i.e. non-responders and / or toxicity), the inventors sought to identify a single TLR agonist that would provide an alternative therapeutic option to BCG via non-toxic stimulation of NK and NKT cells shown to play an important role in the anti-tumour responses induced by BCG in the treatment of NMIBC. The inventors carried out an extensive screening exercise of bacterial TLR ligands with the aim of identifying one that is suitable for expression in an oncolytic Herpes Simplex virus (oHSV). As a result of their screening, they adopted the TLR5 ligand, Flagellin from Burkholderia pseudomallei, i.e. FliC BP. In addition to the TLR ligand, the inventors explored the effects of the oHSV also encoding the cytokine, IL-15. The genes encoding FliC BP and IL15 were harboured on an HSV strain which had had two genes deleted, i.e. ICP34.5 and ICP47. Materials and Methods Production of Oncolytic HSV-1 viruses All oncolytic viruses constructed and used were based on Human Herpes Simplex Virus 1 strain 17+ (Genbank X14112) obtained from ECACC. Knockout of HSV-1 ICP47 gene To knockout ICP47 from the HSV-1 genome, a plasmid (pUC57 del ICP47) was designed and synthesized, containing sequences of two HSV-1 flank regions (Ϫ47 143671-145261bp, 145581-146461bp Genbank X14112) on either side of ICP47 (see Figure 4). A PCR generated CMV EGFP pA cassette was generated from pcDNA3.1+ EGFP using primers CGFPA F (cloning primer 5) GAGGAAGACGAGGAGACGGGCCAGATATACGCGTTG [SEQ ID No: 19] and AGCTCCTCGTCTTCCCAATCCTCCCCCTTG [SEQ ID No: 20]. The PCR product was cut with Bbs-1 and then inserted between flanking regions of ICP47 to produce pUC57 del ICP47 CMV EGFP pA. This viral shuttle vector was linearized (ssp-1) and added to HSV-1 infected cellular BHK DNA and transfected with Calcium Phosphate into BHK cells. The resulting homologous recombinant was plaque purified resulting in clones of HSV-117+ 47- EGFP. The CMV EGFP pA cassette was removed from the virus (HSV-1 17+ 47- EGFP) by homologous recombination using pUC57 del ICP47, which resulted in clones of HSV-117+ 47-. Knockout of HSV-1 ICP34.5 gene To knockout ICP34.5 gene from HSV-117+ 47-, a plasmid (p34.5- DC) containing sequences of two HSV-1 flanking regions (Ϫ34.5123462-124958bp, 125713-126790bp Genbank X14112) either side of ICP34.5 was generated (see Figure 5). p34.5- DC contains a double expression cassette (RSV-pA, CMV pA). Under the CMV promoter, the inventors cloned (BamHI, MfeI) synthetic green fluorescent gene (EGFP) Genbank AF188479.1 from pcDNA3.1+ EGFP to produce p34.5 DC EGFP. This viral shuttle vector was linearized (ssp-1) and added to HSV-1 infected cellular BHK DNA and transfected with Calcium Phosphate into BHK cells. The resulting homologous recombinant was plaque purified resulting in clones of HSV-117+ 34.5-EGFP 47-. Insertion of recombinant Flagella (Burkholderia pseudomallei Bp) sequence into HSV- 117+ 34.5-47- To express Flagella from Burkholderia pseudomallei (Genbank ABA48561.1, a TLR5 ligand) in Oncolytic HSV-117+ 34.5-47-, a plasmid (pcDNA3.1+S-FLIC BP-M3) was designed and synthesized. Like all bacterial proteins, Flagella is normally non- glycosylated, when produced by Burkholderia pseudomallei. Glycosylation of the Bp sequences may inhibit the TLR5 ligand function. To try to check for N-glycosylation sites in the Bp Flagella, the sequences were run through Net N Glyc server website software. The software showed no signalling peptide to engage with the N-glycosylation machinery, but 3x N-glycosylation sites were found (AA No 18, 270, 358). To avoid N- glycosylation sites, Asparagine's (N) on the glycosylation site were changed to glutamines (Q). To secrete the flagella protein out of the cells, a CD33 secrete signal was added. This recombinant sequence was synthesized then cloned into pcDNA3.1+(BamH, EcoRI), resulting in pcDNA3.1+S-FLIC BP-M3. To produce a viral shuttle vector, S-FLIC BP-M3 was subcloned (BamHI, MfeI) from pcDNA3.1+S-FLIC BP-M3 into p34.5- DC under the CMV promoter, resulting in p34.5- DC S-FLIC-BP- M3. To obtain a recombinant virus, p34.5- DC S-FLIC-BP-M3 was linearized (ssp-1) and added to HSV-1 infected cellular BHK DNA and transfected with Calcium Phosphate into BHK cells. The resulting homologous recombinant was plaque purified resulting in clones of HSV-117+ 34.5- S-FLIC BP-M347-. Insertion of recombinant IL-15 sequence into HSV-117+ 34.5-47- To express Human Interleukin 15 (GenBank: AF031167.1) in Oncolytic HSV-117+ 34.5- 47-, a plasmid (pcDNA3.1+IL-15ED) was designed and synthesized (see Figure 6). This recombinant sequence was synthesized then cloned into pcDNA3.1+(BamH, EcoRI). To produce a viral shuttle vector containing IL-15ED, the cytokine sequence was subcloned (PmeI, Xcm-I) from pcDNA3.1+IL-15ED into p34.5- DC plasmid under the RSV promoter. To obtain a recombinant virus, p34.5- DC IL-15ED was linearized (ssp-1) and added to HSV-1 infected cellular BHK DNA and transfected with Calcium Phosphate in BHK cells. The resulting homologous recombinant was plaque purified resulting in clones of HSV-117+ 34.5- IL-15ED 47-. Insertion of both recombinant IL-15 and Flagella (Burkholderia pseudomallei Bp) sequences into HSV-117+ 34.5-47- To express Human Interleukin 15 and Flagella (Bp) sequences from Oncolytic HSV-1 17+ 34.5-47-, the viral shuttle plasmid containing IL-15ED was cut with BamHI, MfeI, and Flagella was inserted under the CMV promoter (see Figure 7). To obtain a recombinant virus, p34.5- DC IL-15ED / S-FLIC-BP-M3 was linearized (ssp-1) and added to HSV-1 infected cellular BHK DNA and transfected with Calcium Phosphate in BHK cells. The resulting homologous recombinant was plaque purified resulting in clones of HSV-117+ 34.5- IL-15ED / S-FLIC-BP-M347-. Example 1 – Screening candidate TLR ligands The inventors first set out to screen several TLR ligands (i.e. TLR2, 4 or 5) to determine which was optimum for use in an oHSV for treating cancer. Each TLR ligand was inserted into the mammalian expression vector, pcDNA3.1. It has previously been shown that N glycosylation sites inhibit ligand receptor interactions. Therefore, each gene was mutated to remove N glycosylation sites, thus turning the asparagine amino acid to glutamine. Also, to allow TLR ligand protein release from mammalian cells, a sequence containing a secretion signal from CD33 was added. To screen the recombinant TLR ligands, the inventors exposed them to NF-KB reporter systems that express their target TLR receptors. These systems test the ability of TLR ligands to activate a NF-KB promoter via a marker gene. TLR2 ligands were tested on Human TLR2 / NF-κB -AP-1 / SEAP reporter in HEK293 cells (Invivogen) (Figure 2b). TLR4 and TLR5 ligands were screened on 293 cells transiently transfected with pNL3.2 Luc NF-KB RE / pGL4.54 Luc2 / TK plasmids (Promega) (Figure 2a). From this screening of TLR ligands, the inventors identified FliC_BP as the best candidate (Figure 2a). This result was consistent with the ability of FliC_BP to interact with TLR5 on HEK cells and trigger activation of transcription factor NF-κB. This TLR5 ligand offers the opportunity to enhance antitumour immune responses seen with oncolytic viruses with an innate bacterial response. Example 2 – Design and generation of an expression cassette and oHSV comprising the TLR5 ligand and / or IL-15 The inventors then constructed a backbone oncolytic virus based on human HSV-1 (strain 17+) in which previously described tumour selective and immune controlling deletions have been included. Referring to Figure 1, there is shown (in the upper panel), a schematic layout of HSV-1 wild type (WT) showing the corresponding positions of the genes, ICP34.5 and ICP47, which are functionally deleted, as discussed below. The inventors then inserted the candidate TLR ligand, FliC_BP, into this backbone oncolytic HSV virus (oHSV or oHSV GFP) to create a series of vectors expressing either FliC_BP (oHSV-FliC_BP), or IL-15 (oHSV_IL-15), or both genes (oHSV-FliC_BP-IL- 15). ICP34.5 The HSV-1 neurovirulence protein, infected cell protein 34.5 (ICP34.5), is necessary for HSV-1 infection of neurons and other healthy cells, because it binds to, and blocks PKR, allowing for viral replication. ICP34.5 blocks PKR pathway activation by binding to, and activating, the PP1α phosphatase, which dephosphorylates eIF-2α, thereby preventing the shutdown of protein translation by eIF-2. Therefore, for the infection of cancer cells, as is the case here when treating bladder cancer, the inventors believed that it would be beneficial for the ICP34.5 gene to be absent, disrupted or non- functional. ICP47 ICP47 is involved in the lytic activity of the HSV-1, and so rendering it non-functional improves its lytic activity. In many cancer cells, the type I IFN signalling pathways are disrupted, and in the absence of ICP34.5, viral replication occurs in cancer cells lacking normal IFN signalling, whereas normal cells will be less permissive because IFN signalling is intact. As such, the inventors believe that this feature of HSV-1 ICP47 / − mutants provides an important safety mechanism, because infection of normal cells will be aborted in the absence of PKR and type I IFN signalling defects. Thus, for the infection of cancer cells, for example when treating bladder cancer, the inventors believe that it would also be advantageous for the ICP47 gene to be absent, disrupted or non-functional. Referring now to the lower panel of Figure 1, there is shown a schematic layout of an expression cassette which is integrated into the site of the disrupted ICP34.5 gene. The expression cassette includes a nucleotide sequence encoding the TLR5 ligand, Flagella from Burkholderia pseudomallei (i.e. FliC BP), the expression of which is controlled by a CMV promoter. Expression extends in the 5’ to 3’ direction (left to right shown in the Figure). The inventors found that FliC_BP is a particularly effective candidate for enhancing antitumour immune responses seen with oncolytic viruses with an innate bacterial response. In addition, the expression cassette includes a nucleotide sequence encoding the cytokine (IL-15), the expression of which is controlled by an RSV promoter. Expression extends in the 3’ to 5’ direction (right to left shown in the Figure). The IL-15 is a pleiotropic cytokine which plays a major role in the development of inflammatory and protective immune responses to microbial invaders and parasites by modulating immune cells of both the innate and adaptive immune systems. The expression of IL-15 promotes the proliferation of NK cells and cytotoxic functions of CD8 T cells. FliC_BP is a bacterial protein that stimulates a strong innate response, and together, IL-15 and FliC_BP act synergistically. The expression cassette is flanked by two polyA tails. The cassette encoding IL-15 and FliC_BP was inserted into the oHSV which was functionally deleted for the genes, ICP34.5 and ICP47, to create the vector, HSV-117+ 34.5- IL-15ED / S-FLIC-BP-M3 47-. As described in the following examples, oHSV-117+ 34.5- IL-15ED / S-FLIC-BP-M3 47- was then tested for its efficacy to treat cancer, such as bladder cancer. Example 3 – Characterising oHSV expressing FliC_BP and / or IL-15 in vitro. To verify viral expression of FliC_BP, the bladder cancer cell line, 5637 was infected with oHSV-FliC_BP and after 24hrs was immuno-stained with a mouse polyclonal serum generated against flagellin bacteria protein from BP. This staining showed expression of FliC_BP in the cytoplasm and strongly at the membrane of infected cells (Figure 2f). No staining signal was detected in 5637 bladder cancer cells infected with oHSV (data not shown) or secondary antibody alone (Figure 2f). To test whether virally expressed FliC_BP can activate the MyD88-NF-KB pathway, the inventors used a TLR receptor NF-KB reporter assay. Supernatant from infected bladder cancer cells was added to 293 cells, which express TLR5 receptor, and which are transiently transfected with a plasmid containing a NF-KB promoter upstream of a luciferase gene. The oHSV-FliC_BP virus showed 10-18-fold increase in luciferase expression compared to oHSV (Figure 2c). This showed that secreted FliC_BP produced after only 4hrs of infection of cancer bladder cells with oHSV-FliC_BP was able to activate NF-KB in the reporter system (Figure 2c). Using a soluble ectodomain of human TLR5 - TLR5 inhibitor (α Hu TLR5-Fc, InvivoGen), the inventors were able to partly block MyD88-NF-KB pathway activation, demonstrating that the activity of the virally produced FliC_BP was at least, in part, via TLR5 binding, showing that the virally expressed protein can interact in a similar manner to its bacterial counterpart and would therefore be expected to stimulate an innate immune response (Figure 2e). To understand the effect of virally expressed FliC_BP alone on a macrophage model, THP-1 cells (+PMA) were treated with oHSV or oHSV FLIC BP which had been UV inactivated. The viral treated THP-1 cells were incubated for 24hrs and qPCR for innate cytokine transcripts was carried out. The data indicates an enhanced innate immune response is seen with oHSV FLIC BP compared to oHSV (Figure 2g). In order to enhance the activation of NK cells by the novel FLiC_BP virus, the inventors aimed to include the cytokine IL-15. The inventors first constructed a virus expressing IL-15 (oHSV IL-15). Expression of IL-15 from oHSV IL-15 was detected using an IL-15 cell bioassay (Promega). A bladder cancer cell line, 5637 was infected with oHSV-IL-15 virus at MOI 0.1 for 48hrs. Supernatant from these infected cells was added to IL-15 Bioassay cells which have been engineered to express luciferase in response to IL-15 signalling. This assay demonstrated that biologically active IL-15 was produced from the virus at approximately 30ng / ml (Figure 2d). The added IL-15 to the virus containing FliC_BP to create oHSV-FliC_BP-IL-15 and demonstrated expression in an IL-15 cell bioassay. Example 4 – In vivo bladder cancer efficacy of oHSV-FliC_BP-IL-15 The data demonstrates that the oncolytic virus according to the invention is effective at curing up to 50% of mice in an in vivo subcutaneous model of bladder cancer (Figure 3a, b). After 60 days of remission these mice were rechallenged with the same bladder cancer cell lines MB49 and remained cancer free (Figure 2c). Splenocytes including CD4, CD8 and NK cells from cured mice showed activation of tumour killing cytokines such as TNF-α in the presence of MB49 tumours cells (Figure 3e, f, g), when infected with oHSV-FliC_BP-IL-15. Splenocytes from naive mice lack this activation. Example 5 – Live replicating HSV enhances the lysis of bladder tumour cells Methodology 4.2 x 106293 cells were plated in 25 cm3flasks and incubated overnight at 37°C. The cells were then infected with HSV or HSV5-15 at a MOI of 1 in 2 ml of 2% FCS medium per flask for 1 hour. HSV5 or HSV5-15 refers to the inventors’ own nomenclature and the insertion of FliC_BP into either HSV-1 or HSV-2. After incubation, the virus was removed, and 5 ml of 10% FCS medium was added per flask. The cells were then incubated for an additional 24 hours. The supernatant was collected, centrifuged at 1500 rpm for 5 minutes at room temperature, and transferred to a 10 cm diameter dish. This cell-free supernatant was exposed to 300 J of UV light (fractionated dose 3x), after which it was stored at -80°C. Four different transitional cell carcinoma (TCC) bladder cancer cell lines were used as per the table below. KU19-19, VMCUB, and TCCSUP-G cells were plated at 1 x 104cells per well (100μL) of a 96-well plate and T24 cells were plated at 8 x 103cells per 96-well plate. All cells were incubated at 370C overnight. Cells were then treated with HSV / HSV5-15 with and without UV inactivation in fresh 2% FCS medium (50μL at MOI 0.5) and incubated for 2 hours.100μl of fresh 10% FCS medium was added and incubated for 72hrs. The medium was removed and 100μl of MTS Reagent (CellTiter 96® AQueous Assay, Promega) was added for a 1-hour incubation at 370C. The absorbance was recorded at 490nm using an ELISA plate reader. Results HSV was successfully inactivated by UV irradiation, although a small amount of residual activity remained (see Figure 8). The four TCC bladder cancer cell lines (T24, TCCSUP, Ku19-19, VMCUB) were seen to be sensitive to oncolysis from HSV5-15 and backbone virus HSV. UV inactivation of HSV5-15 and backbone virus HSV strongly inhibited oncolysis in both virus and all TCC bladder cell lines tested (see Figure 9). These results demonstrate that HSV and HSV5-15 achieve maximal oncolysis of bladder tumour cells when they are in a live replicating state, facilitating the direct destruction of cancer cells but also promoting the release of therapeutic proteins, such as IL-15 and FliC_BP along with tumour antigens. This release is essential for stimulating both innate and adaptive immune responses against tumour antigens and viral proteins leading to sustained anti-tumour immunity. The death of RU19-19 cells at the highest concentrations of UV-inactivated HSV or HSV5-15 is due to the residual activity of the virus following incomplete inactivation from UV (Figure 8) combined with the sensitivity of RU19-19 cells to HSV-induced lysis. Example 6 – FliC_BP expression from plasmid or oncolytic HSV causes activation of TLR2 The inventors aimed to investigate the effect of FliC_BP, when expressed from both a plasmid and an oncolytic HSV, on the activation of Toll-like receptor 2 (TLR2). Methodology NF-KB response SEAP assay for TLR2 Expression of FliC_BP protein from a mammalian plasmid 293 cells were plated at a density of 1.5x104cells per well in a 96-well plate and incubated overnight until reaching 50-70% confluency. The ViaFect™ Transfection Reagent, DNA (at a 7:1 ratio), and diluent (serum-free medium) were allowed to warm to room temperature and gently vortexed. Plasmid DNA (containing FliC_BP plasmid and pcDNA EGFP) was added to the diluent Opti-MEM® according to the proportions. The mixture was pipetted gently to ensure thorough mixing. The ViaFect™ Transfection Reagent was then pipetted directly into the medium containing the diluted DNA without touching the walls of the plastic tubes. The mixture was incubated for 15- 30 minutes at room temperature. Next, 5 µl of the transfection complex was added to the cells, and the plate was gently shaken on a rotating platform shaker (600 rpm) for 30 seconds at low speed. This process was repeated once more per sample. The cells were then incubated for 48 hours. After incubation, the supernatant was removed from the cells and centrifuged at 2000g for 3 minutes. Expression of FliC_BP protein from oncolytic HSV (HSV5) 293 cells were plated at a density of 1.5x104cells per well in a 96-well plate and incubated overnight until reaching 50-70% confluency. The cells were then infected with HSV5 or HSV GFP at a MOI of 1 in 2% FCS medium per flask for 1 hour. HSV5 refers to the inventors’ own nomenclature and the insertion of FliC_BP into either HSV-1 or HSV-2. After incubation, the virus was removed, and 10% FCS medium was added wells. The cells were then incubated for an additional 24 hours. After incubation, the supernatant was removed from the cells and centrifuged at 2000g for 3 minutes. TLR2 stimulation determined using HEK-Blue™ Detection HEK-Blue™ Detection is a specialised cell culture medium designed for the detection of secreted embryonic alkaline phosphatase (SEAP), serving as the reporter protein produced and secreted by the cells. In the experimental setup, 20µl of each sample was dispensed into the wells of a flat-bottom 96-well plate, with six wells allocated per sample. Additionally, 20µl of 10ng / ml Pam3CSK4 was added to designated control wells as a positive control, while 20µl of sterile growth medium served as the negative control. To prepare HEK-Blue™ hTLR2 cells for the assay, the existing medium was removed, and the cells gently rinsed with 5-10 ml of pre-warmed PBS for a T-75 flask. Subsequently, 2-5 ml of pre-warmed PBS was added to the flask and the cells were incubated at 37°C for 1-2 minutes to facilitate detachment. The cells were then detached either by tapping the flask or by using a cell scraper, ensuring the dissociation of any cell clumps by gentle pipetting up and down. Importantly, trypsin was not used for detaching HEK-Blue™ hTLR2 cells to maintain cell integrity. Following detachment, the cells were resuspended at a concentration of 2.8x105cells per ml in HEK-Blue™ Detection medium. Approximately 180µl of cells, corresponding to approximately 50,000 cells, were immediately added to each sample well. Care was taken to avoid prolonged incubation of cells at room temperature in the HEK-Blue™ Detection medium as it could lead to elevated background or false positive readings. The assembled plate was then incubated at 37°C in a 5% CO2 environment overnight to allow for SEAP production and secretion. Following the incubation, SEAP levels were determined using a plate reader at 600 nm, providing quantitative measurements of SEAP activity indicative of the immune response elicited by the samples tested. This assay setup enables the evaluation of immune activation mediated through TLR2 signalling pathways, offering valuable insights into cellular responses to various stimuli or treatments. Results FliC_BP, expressed from either a plasmid or an oncolytic HSV, successfully activated TLR2 in a HEK-Blue™ hTLR2 assay. This result indicates that FliC_BP can trigger TLR2-mediated signalling pathways, suggesting either direct binding of FliC_BP to the TLR2 receptor or the occurrence of crosstalk between TLR5 (the primary receptor for flagellin) and TLR2. Toll-like receptors (TLRs) are crucial components of the innate immune system, responsible for recognizing and responding to microbial pathogens. Different TLR ligands can induce unique cytokine profiles through pathway-specific signalling, illustrating the complexity of TLR pathway interactions. The result indicates that FliC_BP, a flagellin protein from Burkholderia pseudomallei, can trigger TLR2- mediated signalling pathways, suggesting either direct binding of FliC_BP to the TLR2 receptor or the occurrence of crosstalk between TLR5 (the primary receptor for flagellin) and TLR2. This previously unreported activity for B. pseudomallei flagellin or indeed any bacterial flagellin, may account for the observed enhanced ability of FliC_BP to stimulate innate immune responses. Simultaneous activation of the TLR2 and TLR5 pathways likely amplifies the production of proinflammatory cytokines, thereby contributing to a more robust and immediate immune response. The ability of FliC_BP to activate multiple TLR pathways could be a key factor in its potential use as a therapeutic agent, particularly in the context of oncolytic virotherapy where strong immune activation is desired. Conclusions The inventors have created a novel oncolytic virus that is effective for BCG non- responder models (and ultimately patients). Intravesical delivery of this novel oncolytic virus promises highly selective tumour infection and cancer cell cytotoxicity. Advantageously, the virus retains potent tumour cytotoxicity with enhanced immunogenicity by the inclusion of a novel bacterial protein alongside IL-15, which promotes the proliferation of NK cells and cytotoxic functions of CD8 T cells (Figure 2 and 3). As such, the oncolytic virus is able to selectively kill cancer cells in the bladder at the same time as triggering powerful anti-cancer immunity to prevent the cancer recurring.

Claims

Claims 1. A genetic construct comprising a first coding sequence, which encodes a TLR agonist, and a second coding sequence, which encodes a cytokine.

2. The genetic construct according to claim 1, wherein the TLR agonist is a TLR2 agonist, a TLR4 agonist, or a TLR5 agonist.

3. The genetic construct according to claim 2, wherein: (i) the TLR2 agonist is selected from a group consisting of: LpqH, LprA, LprG, LpqT, PhoS1, MPB83, MPT83, LAM, AraLAM, LM, PIM2 / 6, TDM, HSP70, MymA, PE_PGRS33, EsxL, PPE18, PPE26, PPE32, PPE57, Lrp, GPLs, PILAM, PPE60, OmpA and OmpC; (ii) the TLR4 agonist is FimH; or (iii) the TLR5 agonist is selected from a group consisting of: FliC, CagL, CagY, FllA and FlaB.

4. The genetic construct according to any one of the preceding claims, wherein the TLR agonist is Burkholderia pseudomallei flagellin protein (FliC_BP).

5. The genetic construct according to claim 4, wherein FliC_BP comprises or consists of an amino acid sequence substantially as set out in SEQ ID No: 1 or 3, or a fragment or variant thereof, and / or wherein FliC_BP is encoded by the nucleotide sequence substantially as set out in SEQ ID No: 2 or 4, or a fragment or variant thereof.

6. The genetic construct according to any one of the preceding claims, wherein the first coding sequence encodes a CD33 secretion signal.

7. The genetic construct according to claim 6, wherein the CD33 secretion signal comprises or consists of an amino acid sequence substantially as set out in SEQ ID No: 5, or a fragment or variant thereof, and / or wherein the CD33 secretion signal is encoded by the nucleotide sequence substantially as set out in SEQ ID No: 6, or a fragment or variant thereof.

8. The genetic construct according to any one of the preceding claims, wherein the cytokine is a chemokine, optionally wherein the chemokine is selected from a group consisting of: CXCL10, CXCL9, CXCL11, CXCL8, CXCL12, CCL2, CCL3 and CCL5.

9. The genetic construct according to any one of the preceding claims, wherein the cytokine is selected from a group consisting of: IL-2, IL-6, IL-10, IL-12, non-secreting IL-12, IL-15, IL-17, IL-21, IL-33, and type I and type II interferons.

10. The genetic construct according to any one of the preceding claims, wherein the cytokine is IL-15.

11. The genetic construct according to claim 10, wherein IL-15 comprises or consists of an amino acid sequence substantially as set out in SEQ ID No: 9, or a fragment or variant thereof, and / or wherein IL-15 is encoded by the nucleotide sequence substantially as set out in SEQ ID No: 10, or a fragment or variant thereof.

12. The genetic construct according to any one of the preceding claims, wherein the genetic construct further comprises a first promoter, operably linked to the first coding sequence encoding the TLR agonist, and / or a second promoter operably linked to the second coding sequence encoding the cytokine.

13. The genetic construct according to claim 12, wherein the or each promoter is selected from a group consisting of: CMV, RSV, T7 polymerase, EF1a, SV40, PGK1, Ac5, UBI, MP-84 and MP-135.

14. The genetic construct according to either claim 12 or claim 13, wherein the first promoter operably linked to the first coding sequence encoding the TLR agonist, is the cytomegalovirus (CMV) promoter.

15. The genetic construct according to claim 14, wherein the first promoter comprises or consists of a nucleic acid sequence substantially as set out in SEQ ID No: 12, or a fragment or variant thereof.

16. The genetic construct according to any one of claims 12 to 15, wherein the second promoter operably linked to the second coding sequence encoding the cytokine, is the RSV promoter.

17. The genetic construct according to claim 16, wherein the second promoter comprises or consists of a nucleic acid sequence substantially as set out in SEQ ID No: 13, or a fragment or variant thereof.

18. The genetic construct according to any one of the preceding claims, wherein the genetic construct comprises a first and second nucleotide sequence encoding a first and second poly-A tail, preferably wherein the first poly-A tail coding sequence is disposed 3’ of the first coding sequence encoding the TLR agonist and / or wherein the second poly-A tail coding sequence is disposed 3’ of the second coding sequence encoding the cytokine.

19. The genetic construct according to claim 18, wherein the first and / or second poly-A tail comprises a nucleic acid sequence substantially as set out in SEQ ID No: 14 or 15, or a fragment or variant thereof.

20. An expression cassette comprising the genetic construct of any one of claims 1 to 19.

21. An oncolytic virus comprising the genetic construct of any one of claims 1 to 19 or the expression cassette of claim 20.

22. An oncolytic virus comprising a nucleotide sequence, which encodes Burkholderia pseudomallei flagellin protein (FliC_BP).

23. The oncolytic virus according to claim 22, wherein the oncolytic virus comprises a comprises a first promoter, operably linked to the nucleotide sequence, which encodes FliC_BP.

24. The oncolytic virus according to any one of claims 21 to 23, wherein the oncolytic virus is selected from a group consisting of: herpes simplex virus (HSV), Coxsackie virus, Maraba virus, measles virus (MV), Newcastle disease virus (NDV), poliovirus, reovirus, retroviruses, Seneca Valley virus (SVV), Alphavirus, Semliki Forest virus (SFV), Sindbis virus (SINV), Vesicular stomatitis virus (VSV), Sindbis virus (SBV), Adenovrius, Poxvirus, Parvovirus, Flavivirus, Zika virus, Paramyxo virus, Picornavirus, and Rhabdovirus.

25. The oncolytic virus according to any one of claims 21 to 24, wherein: (i) the oncolytic virus is herpes simplex virus (HSV), optionally wherein the HSV is HSV-1 or HSV-2; and / or (ii) wherein the oncolytic virus is a live, replicating virus.

26. The oncolytic virus according to any one of claims 21 to 25, wherein the oncolytic virus comprises a functionally deleted ICP34.5 gene, preferably wherein the oncolytic virus comprises a functional deletion of at least two copies of the ICP34.5 gene.

27. The oncolytic virus according to any one of claims 21 to 26, wherein the oncolytic virus comprises a functionally deleted ICP47 gene.

28. The genetic construct according to any one of claims 1 to 19, the expression cassette according to claim 20, or the oncolytic virus according to any one of claims 21 to 27, for use in therapy.

29. The genetic construct according to any one of claims 1 to 19, the expression cassette according to claim 20, or the oncolytic virus according to any one of claims 21 to 27, for use in treating, preventing or ameliorating cancer.

30. The genetic construct, expression cassette, or oncolytic virus, for use according to claim 29, wherein the cancer is bladder cancer.

31. The genetic construct, expression cassette, or oncolytic virus, for use according to either claim 29 or claim 30, wherein the cancer is non-muscle invasive bladder cancer (NMIBC), or BCG-refractory disease bladder cancer.

32. A pharmaceutical composition comprising the genetic construct according to any one of claims 1 to 19, the expression cassette according to claim 20, or the oncolytic virus according to any one of claims 21 to 27, and a pharmaceutically acceptable vehicle.

33. A process for making the pharmaceutical composition according to claim 32, the process comprising combining a therapeutically effective amount of the genetic construct according to any one of claims 1 to 19, the expression cassette according toclaim 20, or the oncolytic virus according to any one of claims 21 to 27, with a pharmaceutically acceptable vehicle.

Images