Evaluation of &ggr;-Retroviral Vectors That Mediate the Inducible Expression of IL-12 for Clinical Application

The clinical application of interleukin-12 (IL-12) has been hindered by the toxicity associated with its systemic administration. To potentially overcome this problem, we developed a promoter designed to direct IL-12 expression within the tumor environment using an inducible composite promoter containing binding motifs for the nuclear factor of activated T cells (NFAT) linked to a minimal IL-2 promoter. In this study, the NFAT promoter was coupled to a single-chain human IL-12 gene and inserted into 2 &ggr;-retroviral self-inactivating vectors (SERS.NFAT.hIL12 and SERS.NFAT.hIL12.PA2) and 1 &ggr;-retroviral vector (MSGV1.NFAT.hIL.12 PA2). Peripheral blood lymphocytes (PBLs) were double transduced with an antigen-specific T-cell receptor and the 3 NFAT.hIL12 vectors. Evaluation of inducible IL-12 expression, transduction efficiency, and vector production considerations led to the choice of the MSGV1.NFAT.hIL12.PA2 vector for clinical application. MSGV1.NFAT.hIL12.PA2 PG13 retroviral vector producer cell clones were screened by transduction of tumor antigen-specific PBLs. On the basis of expression studies in PBL, clone D3 was chosen to produce clinical-grade viral vector supernatant and was demonstrated to efficiently transduce young tumor-infiltrating lymphocytes (TIL). The vector-transduced young TIL with known tumor recognition demonstrated specific inducible IL-12 production after coculture with HLA-matched tumor targets and had augmented effector function as demonstrated by increased IFN-&ggr; secretion. These results support the clinical application of adoptive transfer of young TIL engineered with the NFAT.hIL12 vector as a new approach for cancer immunotherapy.

[1]  S. Rosenberg,et al.  Improving adoptive T cell therapy by targeting and controlling IL-12 expression to the tumor environment. , 2011, Molecular therapy : the journal of the American Society of Gene Therapy.

[2]  A. Schambach,et al.  Improving transcriptional termination of self-inactivating gamma-retroviral and lentiviral vectors. , 2007, Molecular therapy : the journal of the American Society of Gene Therapy.

[3]  R. Herzog.  Gene therapy for SCID-X1: round 2. , 2010, Molecular therapy : the journal of the American Society of Gene Therapy.

[4]  S. Rosenberg,et al.  CD8+ Enriched “Young” Tumor Infiltrating Lymphocytes Can Mediate Regression of Metastatic Melanoma , 2010, Clinical Cancer Research.

[5]  S. Rosenberg,et al.  Adoptive cell therapy for patients with metastatic melanoma: evaluation of intensive myeloablative chemoradiation preparative regimens. , 2008, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[6]  S. Rosenberg,et al.  Transfer of a TCR gene derived from a patient with a marked antitumor response conveys highly active T-cell effector functions. , 2005, Human gene therapy.

[7]  A. Bakker,et al.  NFAT-controlled expression of GFP permits visualization and isolation of antigen-stimulated primary human T cells. , 2000, Blood.

[8]  M. Del Vecchio,et al.  Interleukin-12: Biological Properties and Clinical Application , 2007, Clinical Cancer Research.

[9]  Christof von Kalle,et al.  and insertional genotoxicity Cell culture assays reveal the importance of retroviral vector design for , 2006 .

[10]  G. Trinchieri,et al.  Tumor-specific CD8+ T cells expressing interleukin-12 eradicate established cancers in lymphodepleted hosts. , 2010, Cancer research.

[11]  R. Morgan,et al.  A simple and reliable method for screening retroviral producer clones without selectable markers. , 1997, Human gene therapy.

[12]  S. Rosenberg,et al.  Adoptive cell transfer therapy. , 2007, Seminars in oncology.

[13]  F. Ozturk,et al.  Expression levels of the PiT‐2 receptor explain, in part, the gestational age‐dependent alterations in transduction efficiency after in utero retroviral‐mediated gene transfer , 2012, The journal of gene medicine.

[14]  M. Eiden,et al.  Evaluation of residual promoter activity in γ-retroviral self-inactivating (SIN) vectors. , 2012, Molecular therapy : the journal of the American Society of Gene Therapy.

[15]  B. Shalmon,et al.  Clinical Responses in a Phase II Study Using Adoptive Transfer of Short-term Cultured Tumor Infiltration Lymphocytes in Metastatic Melanoma Patients , 2010, Clinical Cancer Research.

[16]  K. Cornetta,et al.  Rapid production of clinical-grade gammaretroviral vectors in expanded surface roller bottles using a "modified" step-filtration process for clearance of packaging cells. , 2011, Human gene therapy.

[17]  J. Leonard,et al.  Effects of Single-Dose Interleukin-12 Exposure on Interleukin-12–Associated Toxicity and Interferon-γ Production , 1997 .

[18]  J. Leonard,et al.  Effects of single-dose interleukin-12 exposure on interleukin-12-associated toxicity and interferon-gamma production. , 1997, Blood.

[19]  S. Riddell,et al.  The use of anti-CD3 and anti-CD28 monoclonal antibodies to clone and expand human antigen-specific T cells. , 1990, Journal of immunological methods.

[20]  W. Burns,et al.  Both CD4 and CD8 T Cells Mediate Equally Effective In Vivo Tumor Treatment When Engineered with a Highly Avid TCR Targeting Tyrosinase , 2010, The Journal of Immunology.

[21]  J Cohen,et al.  IL-12 Deaths: Explanation and a Puzzle , 1995, Science.