Targeting the EGF-receptor and the CD38/NADase in solid and hematological malignancies with nanobody-based heavy chain antibodies and AAV vectors

Abstract

Targeted tumor therapy with monoclonal antibodies (mAbs) is based on targeting mutated, selectively expressed or overexpressed cell surface antigens such as the epidermal growth factor receptor (EGFR) or the NAD-hydrolase CD38. However, not all patients respond to targeted therapy. Moreover, many patients develop resistance towards the therapeutic mAb. Patients with head and neck cancer (HNSCC) or metastatic colorectal cancer (mCRC) may develop mutations of the EGFR that result in loss of binding by the mAbs cetuximab and panitumumab. In patients with Multiple Myeloma (MM), loss of CD38 from the cell surface may render tumor cells inaccessible for the αCD38-specific mAb daratumumab and consequently resistant against antibody-mediated effector functions, such as natural killer cell (ADCC)- and complement-dependent (CDC) lysis.

The goal of this thesis was to improve the targeting of EGFR and CD38 overexpressing tumor cells with nanobody-based heavy chain antibodies (hcAb) and Adeno-associated viral (AAV) vectors. Nanobodies comprise a single variable domain of 15 kilo Dalton (kDa). The results show that nanobody 7D12, in contrast to cetuximab, can bind to EGFR epitope escape variants and subsequently inhibit the proliferation of EGFR variant cells. In case of CD38, some nanobodies inhibited the enzymatic activities of CD38 more effectively than daratumumab. To allow Fc-mediated effector functions, selected αEGFR and αCD38 nanobodies were fused to the hinge, CH2, and CH3 domains of human IgG1. These chimeric llama/human heavy chain antibodies (hcAb) are about half the size of conventional mAbs (75 kDa vs. 150 kDa). The Fc- optimized 7D12-hcAb mediated effective ADCC and CDC also against cells expressing EGFR ECD variants that did not respond to cetuximab or panitumumab. In primary bone marrow samples from MM patients the combination of daratumumab with αCD38 hcAbs mediated more effective CDC than daratumumab alone. Finally, CD38-specific nanobodies mediated more effective transduction of CD38+ MM cells by recombinant AAVs.

The results of this thesis pave the way for novel therapeutic approaches against EGFR or CD38 overexpressing solid and hematological malignancies.