Targeted delivery of siRNA using transferrin-coupled lipoplexes specifically sensitizes CD71 high expressing malignant cells to antibody-mediated complement attack

Abstract

The overexpression of membrane-bound complement regulatory proteins (mCRP; CD46, CD55, CD59) preventing opsonization and complement-dependent cytotoxicity (CDC) is considered a major barrier for successful antibody-based cancer immunotherapy. To avoid a potential deleterious effect of mCRP neutralization on normal tissue cells, complement regulation has to be selectively targeted to the malignant cells. In this study, anti-mCRP small interfering RNAs (siRNAs) were encapsulated in transferrin-coupled lipoplexes for the specific delivery to transferrin receptor CD71(high) expressing BT474, DU145, and SW480 as well as corresponding CD71-knockdown (CD71(low)) tumor cells. Targeted delivery with transferrin-siRNA-lipoplexes became possible by charge neutralization and resulted in efficient silencing of all three mCRPs up to 90%, which is dependent on their CD71 expression. The mCRP knockdown led to a significant increase of CDC on CD71(high) tumor cells by 68% in BT474, 58% in DU145, and 40% in SW480 cells but only slightly increased on CD71(low) cells. Downregulation of CD46 and CD55 significantly increased C3 opsonization only on CD71(high) tumor cells. Our results demonstrate for the first time that by specific delivery of anti-mCRP siRNA through transferrin receptor, complement regulation can be selectively neutralized, allowing specific antibody-mediated killing of tumor cells without affecting healthy bystander cells, which appears to be a suited strategy to improve antibody-based cancer immunotherapy.

Targeting peroxiredoxin 1 impairs growth of breast cancer cells and potently sensitises these cells to prooxidant agents

“…One of the theoretical possibilities of tumour-specific downregulation of PRDX1 is the conjugation of anti-PRDX1 siRNA with a tumour-specific targeting agent, e.g., an antibody (reviewed in 40 ). Targeting CD71, 41 acting as a transferrin receptor, would be of special interest, as CD71-overexpressing cancer cells are characterised with an increased iron uptake and, hence, should be more prone to prooxidant actions of ascorbate. 42 Obviously, in HER2-expressing cancers, HER2 molecule itself might be targeted by the siRNA-antibody conjugate.…”

The dual role of complement in cancer and its implication in anti-tumor therapy

“…An alternative approach, down-modulation of mCRPs, has been successfully achieved in vitro by RNA interference (134); however, there are numerous problems (e.g., in vivo stability, tissue specific targeting and unwanted immune system activation) that are currently preventing the use of RNA interference in vivo.…”

Neutralization of membrane complement regulators improves complement-dependent effector functions of therapeutic anticancer antibodies targeting leukemic cells

“…The obstacle of this approach for clinical application is still the targeted delivery of specific siRNA to the tissue of interest. The feasibility of such a strategy has recently been demonstrated by our group showing an improved CDC upon specific delivery of siRNA using transferrin-coated lipoplexes 53 . A successful targeted delivery of mCRPs-specific siRNAs may serve as an adjuvant to improve the efficacy of antibody-based immunotherapy.…”