cancers Article Pimavanserin: A Novel Autophagy Modulator for Pancreatic Cancer Treatment Sharavan Ramachandran, Itishree S. Kaushik and Sanjay K. Srivastava *   Citation: Ramachandran, S.; Kaushik, I.S.; Srivastava, S.K. Pimavanserin: A Novel Autophagy Modulator for Pancreatic Cancer Treatment. Cancers 2021, 13, 5661. https://doi.org/10.3390/cancers 13225661 Academic Editors: Subhash C. Chauhan and David Wong Received: 2 June 2021 Accepted: 5 November 2021 Published: 12 November 2021 Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affil- iations. Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/). Department of Immunotherapeutics and Biotechnology, Center for Tumor Immunology and Targeted Cancer Therapy, Texas Tech University Health Sciences Center, Abilene, TX 79601, USA; sharavan.ramachandran@ttuhsc.edu (S.R.); i.kaushik@ttuhsc.edu (I.S.K.) * Correspondence: sanjay.srivastava@ttuhsc.edu; Tel.: +1-325-696-0464; Fax: +1-325-676-3875 Simple Summary: Pimavanserin a novel anti-psychotic drug suppresses the growth of pancreatic tumors in vitro and in vivo and can be developed as a treatment option for pancreatic cancer. Abstract: Pancreatic tumors exhibit high basal autophagy compared to that of other cancers. Several studies including those from our laboratory reported that enhanced autophagy leads to apoptosis in cancer cells. In this study, we evaluated the autophagy and apoptosis inducing effects of Pima- vanserin tartrate (PVT). Autophagic effects of PVT were determined by Acridine Orange assay and Transmission Electron Microscopy analysis. Clinical significance of ULK1 in normal and pancreatic cancer patients was evaluated by R2 and GEPIA cancer genomic databases. Modulation of proteins in autophagy signaling was assessed by Western blotting and Immunofluorescence. Apoptotic effects of PVT was evaluated by Annexin-V/APC assay. Subcutaneous xenograft pancreatic tumor model was used to evaluate the autophagy-mediated apoptotic effects of PVT in vivo. Autophagy was induced upon PVT treatment in pancreatic ducal adenocarcinoma (PDAC) cells. Pancreatic cancer patients exhibit reduced levels of autophagy initiator gene, ULK1, which correlated with reduced patient survival. Interestingly, PVT induced the expression of autophagy markers ULK1, FIP200, Atg101, Beclin-1, Atg5, LC3A/B, and cleavage of caspase-3, an indicator of apoptosis in several PDAC cells. ULK1 agonist LYN-1604 enhanced the autophagic and apoptotic effects of PVT. On the other hand, autophagy inhibitors chloroquine and bafilomycin blocked the autophagic and apoptotic effects of PVT in PDAC cells. Notably, chloroquine abrogated the growth suppressive effects of PVT by 25% in BxPC3 tumor xenografts in nude mice. Collectively, our results indicate that PVT mediated pancreatic tumor growth suppression was associated with induction of autophagy mediated apoptosis. Keywords: cancer; autophagy; cell death; drug repurposing; apoptosis 1. Introduction Several stress elements including drug-induced stress activate intra-cellular survival mechanisms such as autophagy to tolerate the harsh conditions or trigger cell death path- ways [1,2]. Autophagy occurs in three forms such as; macroautophagy, microautophagy, and chaperone-mediated autophagy. Macroautophagy, hereafter referred to as autophagy, is the most prevalent form of autophagy in eukaryotes, and play a pivotal role in maintain- ing tissue homeostasis. Autophagy is considered as an evolutionarily conserved catabolic process, where the sequestered cellular organelles are delivered to lysosomes by the au- tophagosomes for degradation. This ubiquitous process happens in five distinct stages: initiation, vesicle nucleation, vesicle elongation, autophagosome expansion, and fusion of autophagosome with lysosome for degradation. ULK1 orchestrates the initiation of autophagy, where it assembles with Atg13, Atg101, and FIP200 to form the autophagy initiation complex. Phosphorylation of ULK1 at Ser 757 prevents ULK1 in forming the autophagy initiation complex, thereby inhibits autophagy. The ULK1 complex guides the assembly of class-III PI3K or VPS34 complex comprising VPS34, Atg14, UVRAG and Cancers 2021, 13, 5661. https://doi.org/10.3390/cancers13225661 https://www.mdpi.com/journal/cancers