The therapeutic potential of anticancer antibodies is limited by the resistance 
of tumor cells to complement-mediated attack, primarily through the 
over-expression of membrane complement regulatory proteins (mCRPs: CD46, CD55 
and CD59). Trastuzumab, an anti- HER2 monoclonal antibody, approved for the 
treatment of HER2-positive breast and gastric cancers, exerts only minor 
complement-mediated cytotoxicity (CDC). Pertuzumab is a novel anti-HER2 
monoclonal antibody, which blocks HER2 dimerization with other ligand-activated 
HER family members. Here, we explored the complement-mediated anti-tumor effects 
of trastuzumab and pertuzumab on HER2-positive tumor cells of various 
histological origins. Delivery of chemically stabilized anti-mCRP siRNAs using 
cationic lipoplexes, AtuPLEXes, to HER2-over-expressing BT474, SK-BR-3 (breast), 
SKOV3 (ovarian) and Calu-3 (lung) cancer cells reduced mCRPs expression by 
85-95%. Knockdown of individual complement regulators variably led to increased 
CDC only upon combined treatment with trastuzumab and pertuzumab. The combined 
down-regulation of all the three regulators augmented CDC by 48% in BT474, 46% 
in SK-BR-3 cells, 78% in SKOV3 cells and by 30% in Calu-3 cells and also 
increased complement-induced apoptosis and caspase activity on mCRP neutralized 
tumor cells. In addition, antibody-induced C3 opsonization of tumor cells was 
significantly enhanced after mCRP silencing and further augmented tumor cell 
killing by macrophages. Our findings suggest that siRNA-induced inhibition of 
complement regulator expression clearly enhances complement- and 
macrophage-mediated anti-tumor activity of trastuzumab and pertuzumab on 
HER2-positive tumor cells. Thus - if selectively targeted to the tumor - 
siRNA-induced inhibition of complement regulation may serve as an innovative 
strategy to potentiate the efficacy of antibody-based immunotherapy.