Abstract B018: Targeting Notch4 and HIF2α signaling reduces the ability of cells to survive chemotherapy by undergoing the polyaneuploid transition
Authors: Carroll Christopher P. , Hammarlund Emma , Pienta Kenneth J. , Amend Sarah R. 
Published: May 2022

Abstract
Giant cancer cells have been ignored for 200 years, but recently coupled to a role in metastasis and therapeutic resistance. Some cancer cells appear to survive environmental stress, such as chemotherapy or radiotherapy, via a polyaneuploid transition. This results in a PolyAneuploid Cancer Cell (PACC) state with up to 10-fold larger cell sizes. It is proposed that PACCs utilize a conserved evolutionarily or developmental program for polyploidization, which is commonly observed in other organisms and tissues. After a stress-induced formation, the PACC state first undergoes a period of quiescence and eventually undergoes reductive cell division to give rise to proliferative progeny. We hypothesize that PACCs play a role in tumor recurrence and therapy resistance by surviving the initial treatment and subsequently reseeding the tumor via pseudohypoxic HIF signaling. Dosing with LD50 cisplatin for 72h results in the induction of the PACC state in cancer cell lines from breast (HCC1806), renal (786-0), colon (HCT116), and lung (U1690) tumors. Cells in the PACC state had 3-fold larger nuclei after 3 days of treatment and 6-fold larger nuclei after 10 days. Transcriptomic data showed an overall decrease in gene expression across all 4 cell lines. However, there was an increased expression of HIF2 (2.7-fold), Notch4 (2-fold), Plekhs1 (2.3-fold) in PACCs relative to untreated control. Moreover, there was an increase in the expression of replication dependent histones. Overall, ATACseq indicated a reduction in open chromatin in PACCs, as measured by reduced levels of histone 4 and histone 4 modifications (H4k5ac, h4k16ac, H4R3me2). However, H3k27me3 was increased 20% and AP1 binding sites were enriched in open chromatin regions. HIF2 and AP1 cooperate to regulate gene expression and confer therapy resistance. We hypothesize that AP1 is activated in response to treatment and stabilizes HIF2 under oxygenated conditions. HIFs regulate regulators of the cell cycle to induce polyploidy and downregulate β-catenin, a known inducer of cytokinesis. Small molecule inhibition of Notch and HIF-2α reduced PACC formation by 50% and 65% respectively at day 3 and 60% and 75% at day 10. The surviving PACCs were smaller than those untreated by Notch and HIF-2α inhibitors. Large metabolic changes must be undertaken to generate and sustain the larger cellular mass accompanied by polyploidy. HIF signaling metabolically rewires cells to a glycolytic phenotype that provides a carbon source for increased de novo generation of nucleotides, lipids, and proteins. Indicative of this rewiring is that PACCs have increased expression of Hexokinase (2.5-fold), CPT1 (2.3-fold), and GP6D (2-fold). In conclusion, tumor cells that survive chemotherapy are polyaneuploid, larger, and in an altered metabolic state. This process appears driven by Notch4 and HIF-2α signaling. Combining conventional chemotherapy with Notch or HIF2α inhibitors presents an opportunity to reduce tumor recurrence by inhibiting a survival mechanism.
Citation Format: Christopher P. Carroll, Emma Hammarlund, Kenneth J. Pienta, Sarah R. Amend. Targeting Notch4 and HIF2α signaling reduces the ability of cells to survive chemotherapy by undergoing the polyaneuploid transition [abstract]. In: Proceedings of the AACR Special Conference on the Evolutionary Dynamics in Carcinogenesis and Response to Therapy; 2022 Mar 14-17. Philadelphia (PA): AACR; Cancer Res 2022;82(10 Suppl):Abstract nr B018.