Enhancing the Clinical Activity of HER2/neu Specific T Cells William Gwin, MD Internal Medicine, Resident University of Washington
Immunotherapy and Cancer Cancer vaccines were originally used in melanoma as an adjuvant treatment to elicit an immune reaction to cancer cells. Adoptive immune therapy was then performed on Melanoma using CD8 clones. Adoptive T cell therapy involves antigenspecific T cells: Selection Ex vivo expansion Adoptive transfer
The Tumor Vaccine Group Research focusing on targeting HER2/neu expressing malignancies. Original work focused on protein, peptide, and DNA vaccines targeting the HER-2/neu (HER2) protein. Adoptive T Cell therapy strategies focusing on CD4 cell lines Investigating if T Cell adoptive therapy with a mixed population is as effective as a clonal population in inhibiting growth.
HER2/neu ErbB-2 Human Epidermal growth factor Receptor Cell membrane surfacebound receptor tyrosine kinase Normally involved in signal transduction pathways leading to cell growth and differentiation QuickTime and a TIFF (Uncompressed) decompressor are needed to see this picture.
Why Target HER2/neu? Amplification of the HER2/neu gene or its protein product is associated with several cancers. 15-20 percent of breast cancers have an amplification of HER2/neu. Associated with increased disease recurrence and worse prognosis. Overexpression also occurs in: ovarian cancer stomach cancer uterine serous endometrial carcinoma
Adoptive T Cell Therapy and HER2/neu Expressing Tumors Previously the most antigenic sequences on HER2/neu have been identified. These antigenic peptides were shown to increase the HER2/neu specific T cell repertoire in vivo. This enabled isolation of increased numbers of antigen specific T cells for ex vivo expansion. It has remained unclear as to which cytokines are best suited during ex vivo culture to promote the generation of high avidity, functional T cells.
Hypothesis The ex vivo cytokine environment in which antigenspecific T cells are expanded can greatly influence the function and clinical activity of the T cells in vivo. QuickTime and a TIFF (Uncompressed) decompressor are needed to see this picture.
Cytokines of interest Cytokines Properties Critical T cell growth factor; promotes function of effector CD8+ T cells IL-4 Stimulates differentiation of naïve CD4 T cells into Th2 cells IL-7 Regulates homeostatic proliferation of memory T cells IL-12 Stimulates the production of IFN-g and TNF-a from T cells and NK cells IL-15 Promotes proliferation and effector function of CD8+ T cells, NK, and NKT cells IL-18 Induces IFN-g and accelerated memory CD8+ T cell proliferation 1 Elicits concurrent humoral and cellular responses; promotes differentiation of naïve CD8 T cells into effector T cells
Aim: To determine the effects of various culture conditions on the induction of antitumor T cell activity in vivo. Culture conditions evaluated: +4 +7 +12 +15 +18 +21 +anti-il-10 +anti-tgfb QuickTime and a TIFF (Uncompressed) decompressor are needed to see this picture.
Isolation, Expansion and Assessment of T Cells 100 μg peptide s.c. Vaccine 1 Day 0 7 Vaccine 2 14 Vaccine 3 21 SAC HARVEST SPLEENS, ISOLATE AND EXPAND T CELLS (IVS on immunizing antigen) DIFFERENT CULTURE CONDITIONS in vivo efficacy
Culture Conditions Impact T Cell Therapeutic Efficacy Tumor size (mm 3 ) 1500 Naive +IL-4 +IL-7 1000 +IL-12 +IL-15 +IL-18 +1 500 +anti-il-10 +anti-tgf-b 0 0 10 20 30 40 p101tc:livemmc (10:1) Time (days)
Culture Conditions Impact T Cell Therapeutic Efficacy 1500 Tumor size (mm 3 ) 1000 500 Naive +1 +anti-il-10 45% Inhibition p<0.0001 65%, 67% Inhibition p<0.0001 36%, 39% Inhibition p=0.0019, p=0.0007 0 0 10 20 30 40 p101tc:livemmc (10:1) Time (days)
Conclusions Cytokine addition to the ex vivo culture does improve the therapeutic efficacy in this model. Compared to alone, the most effective culture conditions that induced significantly greater therapeutic efficacy were /anti-il-10 and /IL- 21 cultures.
Continued Research What cytokines are being produced by these T cells? What subtypes of T cells are these various culture conditions producing ex vivo? How can we enhance the development of particular T cell subtypes such as Th17 cells?
Culture Conditions Impact T Cell Cytokine Secretion 5000 IL-10 4000 Concentration (pg/ml) 3000 2000 1000 b All secrete IL-10 except where anti-il-10 mab added B a s e l i n e IL-4 IL-7 IL-12 IL-15 IL-18 T cell culture cytokines 1 IL-10 Anti- Anti- TGF
/1 Induce Highly Active Antitumor T Cells Secreting Autoimmune Cytokines IL-17 TNF-a 300 3000 Concentration (pg/ml) 200 100 Concentration (pg/ml) 2000 1000 B a s e l i n e IL-4 IL-7 IL-12 IL-15 IL-18 1 T cell culture c ytokines 0 B a s e l i n e IL-4 IL-7 IL-12 IL-15 IL-18 1 T cell culture cytokines Anti- Anti- IL-10 TGF Anti- Anti- IL-10 TGF Only /1 culture shows high production of IL-17 and TNF-a
Conclusions Only /anti-il-10 culture showed low IL-10 production. Blockade of this immunosuppressive cytokine may contribute to enhance the therapeutic efficacy. Only IL2/1 culture showed high IL-17 and TNF-a production. Therapeutic efficacy may be enhanced by the promotion of a pro-inflammatory cytokine environment at the tumor site. Th17 cells are known to produce IL-17 This subpopulation of T cells has been shown to be important in autoimmunity
Acknowledgements The Tumor Vaccine Group Dr. Nora Disis Dr. Vy Lai Dr. Ekram Gad