טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentHila Novak
SubjectCD33 as Target for Immunotherapy: Functional Properties and
Targeting of Cytotoxic T Cells
DepartmentDepartment of Biology
Supervisor Full Professor Reiter Yoram
Full Thesis textFull thesis text - English Version


Abstract

In the present work we show that effector cytotoxic T cells can be specifically recruited to tumor cells via antibody-mediated MHC class I bearing highly antigenic peptide complexes. A recombinant chimeric molecule was created by the genetic fusion of the scFv antibody fragment derived from the anti-EGFR monoclonal antibody C225, to monomeric single-chain HLA-A2 complexes containing immunodominant tumor or viral specific peptides. The fusion protein can induce very efficiently CTL-dependent lysis of EGFR-expressing tumor cells regardless of the expression of self peptide-MHC complexes. Moreover, the molecule exhibited very potent antitumor activity in vivo in nude mice bearing preestablished human tumor xenografts.

The fusion molecule combines the advantages of the well-established tumor targeting capabilities of high affinity recombinant fragments of antibodies with the known efficient, specific and potent killing ability of CD8 T lymphocytes directed against highly antigenic MHC/peptide complexes.

We expended the reported strategy to the hematopoietic system. CD33 was chosen for this challenge. Being recognized as the marker for myeloid progenitor cells and displays a clinical target for hematological malignancies of the myeloid arm. By applying a novel surface assisted phage display method we were able to isolate a recombinant mAb towards CD33. Gold surfaces covered with polylayer and NTA/Ni were coated with recombinant His-tagged CD33 and exposed to a large Fab antibody library. Binding selectivity was shown both on recombinant CD33 and myeloid leukemia cell lines.

The findings of this work include the discovery of a novel T cell subpopulation which expresses CD33. Under polyclonal activation of human peripheral blood T lymphocytes, the CD33 positive T cell subpopulation appears and reaches a rather stable fraction among activated T cells. CD3+CD33+ subpopulation was shown to have a unique cytokine profile. This finding was supported by expression pattern of selected cytokines. We also found that this subpopulation expresses markers related to the inhibitory well known Tregs.

Perhaps the most exquisite phenomena was the suppressive effect of this subpopulation on proliferating T cells. We were able to show CD3+CD33+ T cells actively suppress T cell proliferation in a dose dependent manner.

To our knowledge, these findings define a previously unknown subset of regulatory/inhibitory T cells.