title: Cytokine-modified Mycobacterium smegmatis as a novel anticancer immunotherapy.
abstract: Intravesical administration of live M. bovis BCG organisms for carcinoma in situ of the urinary bladder is the most successful immunotherapy for solid malignancy. Nevertheless BCG-therapy is associated with significant toxicity and is ineffective in 30-40% of cases. Recently it has been proposed that cytokine-modified mycobacteria may give greater efficacy. As any immunotherapy involving administration of live BCG organisms (wild-type or recombinant) is likely to have associated toxicity (notably in the immunocompromised), we examined the anti-tumour potential of the closely related nonpathogenic organism, Mycobacterium smegmatis, and a TNFalpha gene-modified recombinant M. smegmatis. When wild-type M. smegmatis were delivered to immunocompetent C57Bl/6 mice bearing the transplantable MB49 bladder tumour, efficacy comparable to live BCG was observed with 10-20% long-term survival. However, this effect was lost in both Nude and Beige mice, lacking functional T and NK cells, respectively. Recombinant M. smegmatis secreting TNFalpha, however, gave a 70% durable tumour-free survival. Lymphocytes from draining lymph-nodes and spleens of these mice exhibited pronounced IFNgamma production to mycobacterial-antigen and tumour-lysate, indicating a bias towards cell-mediated immunity. This was further supported by histopathological examination of the tumour site, which revealed significantly increased numbers of CD3+ lymphocytes in animals receiving the recombinant vaccine, but not in those receiving wild-type bacteria. Importantly, tumour rejection following M. smegmatis/TNFalpha was independent of T lymphocytes, as athymic Nude mice efficiently eradicated MB49 tumours. In contrast, the therapeutic efficacy of M. smegmatis/TNFalpha was reduced in animals deficient in NK cytolytic function, suggesting a role for NK-cells in initial tumour destruction. Furthermore the absence of NK-function in Beige mice did not prevent the establishment of tumour-protective memory. No toxicity was observed with wild-type or recombinant M. smegmatis in immunocompetent, T-deficient or NK-deficient models. We demonstrate for the first time that recombinant mycobacteria expressing mammalian cytokines have markedly increased anti-tumour properties. The lack of toxicity suggests that M. smegmatis is a "safe" vehicle for use in immunocompromised patients.
authors.affiliation: Applied Immunology Laboratory and Clinical Biotherapy Programme, Cancer Research UK Clinical Centre, St. James's University Hospital, Leeds, United Kingdom.
authors.forename: Sarah L
authors.initials: SL
authors.lastname: Young
authors.forename: Michael
authors.initials: M
authors.lastname: Murphy
authors.forename: Xing Wu
authors.initials: XW
authors.lastname: Zhu
authors.forename: Patricia
authors.initials: P
authors.lastname: Harnden
authors.forename: Michael A
authors.initials: MA
authors.lastname: O'Donnell
authors.forename: Keith
authors.initials: K
authors.lastname: James
authors.forename: Poulam M
authors.initials: PM
authors.lastname: Patel
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Selby
authors.forename: Andrew M
authors.initials: AM
authors.lastname: Jackson
mesh_terms: D000818:Animals; D018572:Disease-Free Survival; D005818:Genetic Engineering; D006801:Humans; D007121:Immunocompetence; D007167:Immunotherapy / Q000379:methods*; D007694:Killer Cells, Natural / Q000502:physiology; D051379:Mice; D008810:Mice, Inbred C57BL; D020102:Mycobacterium smegmatis / Q000235:genetics* / Q000276:immunology* / Q000472:pathogenicity; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis / Q000235:genetics* / Q000494:pharmacology; D001749:Urinary Bladder Neoplasms / Q000276:immunology / Q000628:therapy* / Q000662:veterinary
chemical_list: D014409:Tumor Necrosis Factor-alpha