cancers Article IGF2 Mediates Resistance to Isoform-Selective-Inhibitors of the PI3K in HPV Positive Head and Neck Cancer Mai Badarni 1,2 , Manu Prasad 1,2 , Artemiy Golden 3 , Baisali Bhattacharya 1,2 , Liron Levin 4,5 , Ksenia M. Yegodayev 1,2 , Orr Dimitstein 2,6 , Ben-Zion Joshua 2,7 , Limor Cohen 1,2 , Ekaterina Khrameeva 3 , Dexin Kong 8 , Angel Porgador 1,2 , Alex Braiman 1,2 , Jennifer R. Grandis 9 , Barak Rotblat 5,10, * and Moshe Elkabets 1,2, *   Citation: Badarni, M.; Prasad, M.; Golden, A.; Bhattacharya, B.; Levin, L.; Yegodayev, K.M.; Dimitstein, O.; Joshua, B.-Z.; Cohen, L.; Khrameeva, E.; et al. IGF2 Mediates Resistance to Isoform-Selective-Inhibitors of the PI3K in HPV Positive Head and Neck Cancer. Cancers 2021, 13, 2250. https://doi.org/10.3390/ cancers13092250 Academic Editor: Miriam Martini Received: 25 March 2021 Accepted: 29 April 2021 Published: 7 May 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/). 1 The Shraga Segal Department of Microbiology, Immunology and Genetics, Faculty of Health Science, Ben-Gurion University of the Negev, Beer-Sheva 84105, Israel; maibda@post.bgu.ac.il (M.B.); manupras@post.bgu.ac.il (M.P.); baisali@post.bgu.ac.il (B.B.); Kseniay@post.bgu.ac.il (K.M.Y.); ayashli@post.bgu.ac.il (L.C.); angel@bgu.ac.il (A.P.); braiman@bgu.ac.il (A.B.) 2 Faculty of Health Sciences, Ben-Gurion University of the Negev, Beer-Sheva 84105, Israel; orrdi@clalit.org.il (O.D.); benzionj@bmc.gov.il (B.-Z.J.) 3 Center of Life Sciences, Skolkovo Institute of Science and Technology, 121205 Moscow, Russia; golden.artemiy@gmail.com (A.G.); E.Khrameeva@skoltech.ru (E.K.) 4 Bioinformatics Core Facility, National Institute for Biotechnology in the Negev, Ben-Gurion University of the Negev, Beer-Sheva 84105, Israel; levinl@post.bgu.ac.il 5 The National Institute for Biotechnology in the Negev, Ben-Gurion University of the Negev, Beer Sheva 84105, Israel 6 Department of Otolaryngology—Head and Neck Surgery, Soroka University Medical Center, Beer-Sheva 84105, Israel 7 Department of Otorhinolaryngology and Head & Neck Surgery, Barzilay Medical Center, Ashkelon 7830604, Israel 8 School of Pharmaceutical Sciences, Tianjin Medical University, Tianjin 300070, China; kongdexin@tmu.edu.cn 9 Department of Otolaryngology—Head and Neck Surgery, University of California San Francisco, San Francisco, CA 94143, USA; Jennifer.Grandis@ucsf.edu 10 Department of Life Sciences, Faculty of Life Science, Ben-Gurion University of the Negev, Beer-Sheva 84105, Israel * Correspondence: rotblat@bgu.ac.il (B.R.); moshee@bgu.ac.il (M.E.); Tel.: +972-(0)8-6428806 (B.R.); +972-86428846 (M.E.) Simple Summary: In the current study, we delineate the molecular mechanisms of acquisition of resistance to two isoform-selective inhibitors of PI3K (isiPI3K), alpelisib and taselisib, in human papillomavirus positive head and neck cell lines. By comparing RNA sequencing of isiPI3K-sensitive tumor cells and their corresponding isiPI3K-acquired-resistant tumor cells, we found that overex- pression of insulin growth factor 2 (IGF2) is associated with the resistance phenotype. We further demonstrated by gain and loss of function studies that IGF2 plays a causative role in limiting the sensitivity of human papillomavirus-positive head and neck cell lines. Moreover, we show that blocking IGF2 stimulation activity, using an inhibitor of the IGF1 receptor (IGF1R), enhances isiPI3K efficacy and displays a synergistic anti-tumor effect in vitro and superior anti-tumor activity ex vivo and in vivo. Abstract: Over 50% of human papilloma positive head-and-neck cancer (HNC HPV+ ) patients harbor genomic-alterations in PIK3CA, leading to hyperactivation of the phosphatidylinositol-4, 5-bisphosphate 3-kinase (PI3K) pathway. Nevertheless, despite PI3K pathway activation in HNC HPV+ tumors, the anti-tumor activities of PI3K pathway inhibitors are moderate, mostly due to the emergence of resistance. Thus, for potent and long-term tumor management, drugs blocking resistance mechanisms should be combined with PI3K inhibitors. Here, we delineate the molecular mechanisms of the acquisition of resis- tance to two isoform-selective inhibitors of PI3K (isiPI3K), alpelisib (BYL719) and taselisib (GDC0032), in HNC HPV+ cell lines. By comparing the transcriptional landscape of isiPI3K-sensitive tumor cells with that of their corresponding isiPI3K-acquired-resistant tumor cells, we found upregulation of insulin growth factor 2 (IGF2) in the resistant cells. Mechanistically, we show that upon isiPI3K treatment, Cancers 2021, 13, 2250. https://doi.org/10.3390/cancers13092250 https://www.mdpi.com/journal/cancers