A novel bispecific siRNA concept: Efficient dual knockdown of YAP1 and WWTR1 with a single guide strand

Yes-associated protein (YAP, encoded by <i>YAP1</i>) and transcriptional coactivator with PDZ-binding motif (TAZ, encoded by <i>WWTR1</i>) are master transcriptional regulators in the Hippo pathway. Because of their complementary roles, simultaneously inhibiting these proteins can efficiently attenuate downstream signaling. Their aberrant activation contributes to serious diseases, including cancer and fibrosis, making them key therapeutic targets. However, no approved therapeutic agents directly target YAP/TAZ because of technical challenges in designing small-molecule inhibitors for transcription regulators. Here, we present a novel approach using a bispecific small interfering RNA (siRNA) that can simultaneously suppress both <i>YAP1</i> and <i>WWTR1</i> with a single guide strand by targeting the consensus sequence of the two genes. Our lead bispecific siRNA, bs<i>YW</i>-61, can potently knock down <i>YAP1</i>/<i>WWTR1</i> across human, cynomolgus monkey, mouse, and rat cell lines. Dual knockdown of <i>YAP1</i>/<i>WWTR1</i> synergistically suppressed downstream gene expression patterns and cancer cell proliferation. Favorable target specificity and off-target profile were confirmed by luciferase reporter assays and RNA sequencing. Additionally, a chemically modified bispecific siRNA showed effective knockdown <i>in vivo</i>. Our data demonstrate the potential for using a bispecific siRNA targeting <i>YAP1</i>/<i>WWTR1</i> as a therapeutic agent for liver diseases, presenting a novel approach for dual targeting with a simple conventional siRNA structure.