Lipoplex mediated silencing of membrane regulators (CD46, CD55 and CD59) enhances complement‐dependent anti‐tumor activity of trastuzumab and pertuzumab

Abstract

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.

Highlights

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  Delivery of 2′‐O‐methyl modified siRNAs to CD46, CD55 and CD59 (mCRPs) using lipoplex.

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  mCRPs inhibition sensitizes tumor cells to trastuzumab and pertuzumab induced complement attack.

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  Enhanced C3 opsonization of HER2‐positive tumor cells upon mCRP silencing.

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  Increased killing of opsonized tumor cells by macrophages.

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  Enhanced anti‐tumor effect of trastuzumab and pertuzumab upon mCRP inhibition.