Improvement of ACK1-targeted therapy efficacy in lung adenocarcinoma using chloroquine or bafilomycin A1

Background: Activated Cdc42-associated kinase 1 (Ack1) is a promising druggable target for Cancer, but its inhibitors only showed moderate effects in clinical trials. The study aimed to investigate the underlying mechanisms and improve the antitumor efficacy of Ack1 inhibitors.

Methods: RNA-seq was performed to determine the downstream pathways of ACK. Using Lasso COX regression analysis, we built a risk signature with ACK1-related Autophagy genes in the lung adenocarcinoma (LUAD) patients from The Cancer Genome Atlas (TCGA) project. The performance of the signature in predicting the tumor immune environment and response to immunotherapy and chemotherapy were assessed in LUAD. CCK8, mRFP-GFP-LC3 assay, western blot, colony formation, wound healing, and transwell migration assays were conducted to evaluate the effects of the Ack1 Inhibitor on lung Cancer cells. A subcutaneous NSCLC xenograft model was used for in vivo study.

Results: RNA-seq revealed the regulatory role of Ack1 in Autophagy. Furthermore, the risk signature separated LUAD patients into low- and high-risk groups with significantly different prognoses. The two groups displayed different tumor immune environments regarding 28 immune cell subsets. The low-risk groups showed high immune scores, high CTLA4 expression levels, high immunophenoscore, and low DNA mismatch repair capacity, suggesting a better response to immunotherapy. This signature also predicted sensitivity to commonly used chemotherapy and targeted drugs. In vitro, the Ack1 inhibitors (AIM-100 and Dasatinib) appeared to trigger adaptive autophagy-like response to protect lung Cancer cells from Apoptosis and activated the AMPK/mTOR signaling pathway, partially explaining its moderate antitumor efficacy. However, blocking lysosomal degradation with chloroquine/Bafilamycine A1 or inhibiting AMPK signaling with compound C/shPRKAA1 enhanced the Ack1 inhibitor's cytotoxic effects on lung Cancer cells. The efficacy of the combined therapy was also verified using a mouse xenograft model.

Conclusions: The resulting signature from ACK1-related Autophagy genes robustly predicted survival and drug sensitivity in LUAD. The lysosomal degradation inhibition improved the therapeutic effects of the Ack1 Inhibitor, suggesting a potential role for Autophagy in therapy evasion.

ACK1; AMPK; Bafilomycin A1; Chloroquine; LUAD; Prognosis.