Blocking HIF signaling via novel inhibitors of CA9 and APE1/Ref-1 dramatically affects pancreatic cancer cell survival

Abstract

Pancreatic ductal adenocarcinoma (PDAC) has reactive stroma that promotes tumor signaling, fibrosis, inflammation, and hypoxia, which activates HIF-1α to increase tumor cell metastasis and therapeutic resistance. Carbonic anhydrase IX (CA9) stabilizes intracellular pH following induction by HIF-1α. Redox effector factor-1 (APE1/Ref-1) is a multifunctional protein with redox signaling activity that converts certain oxidized transcription factors to a reduced state, enabling them to upregulate tumor-promoting genes. Our studies evaluate PDAC hypoxia responses and APE1/Ref-1 redox signaling contributions to HIF-1α-mediated CA9 transcription. Our previous studies implicated this pathway in PDAC cell survival under hypoxia. We expand those studies, comparing drug responses using patient-derived PDAC cells displaying differential hypoxic responses in 3D spheroid tumor-stroma models to characterize second generation APE1/Ref-1 redox signaling and CA9 inhibitors. Our data demonstrates that HIF-1α-mediated CA9 induction differs between patient-derived PDAC cells and that APE1/Ref-1 redox inhibition attenuates this induction by decreasing hypoxia-induced HIF-1 DNA binding. Dual-targeting of APE1/Ref-1 and CA9 in 3D spheroids demonstrated that this combination effectively kills PDAC tumor cells displaying drastically different levels of CA9. New APE1/Ref-1 and CA9 inhibitors were significantly more potent alone and in combination, highlighting the potential of combination therapy targeting the APE1-Ref-1 signaling axis with significant clinical potential.

Architecture of The Human Ape1 Interactome Defines Novel Cancers Signatures

“…These 11 gene sets were then subjected to survival analysis for a better understanding of their role according to a clinical perspective. LIHC, LUAD and PAAD networks, which were composed of bad prognostic genes in high percentages, confirmed previous studies on the important role of APE1 in these cancers [52][53][54][55][57][58][59][60][61][62][63][64][65] . Kaplan-Meier estimation was used to calculate survival probability associated with these genes; the ones having significantly lower survival prognosis (p < 0.05) were used to create cancer-specific PPI subnetworks ( Fig.…”

“…In order to focus our attention on the most relevant cancer types, we based our bioinformatic analysis on the eleven datasets having the highest number of bad prognostic genes. Among those, we were interested in particular in liver (LIHC) [52][53][54][55][56] , lung (LUAD) [57][58][59][60] and pancreatic (PAAD) [61][62][63][64][65] cancer datasets, as the essential role of APE1 in the tumorigenic processes of these cancer types is already well established. Notably, the LUAD dataset was the third having the highest number of bad prognostic genes (n = 153).…”

Differential expression of APE1 in hepatocellular carcinoma and the effects on proliferation and apoptosis of cancer cells

“…On the one hand, APE1 functions as an endonuclease involved in DNA base excision repair, it is closely related to tumor cell proliferation; on the other hand, it regulates the activity of redox-sensitive transcription factors (12). Many studies showed APE1 over-expression in a variety of tumors (13)(14)(15). Some scholars used enzyme-linked immunosorbent assay to detect serum APE1 in patients suffering from bladder cancer without chemotherapy or radiotherapy (51 cases) and in a non-tumor control group (55 cases).…”

Clinical and Pre-Clinical Evidence of Carbonic Anhydrase IX in Pancreatic Cancer and Its High Expression in Pre-Cancerous Lesions

“…Moreover, a combination treatment with APE1/Ref-1 inhibitor E3330 and CAIX inhibitor SLC-0111 (Clinical Trial NCT02215850) was shown to reduce proliferation and size of three-dimensional spheroids derived from PDAC patients [64]. In line with these findings, the application of novel analogues of both APE1/Ref-1 and CAIX inhibitors (namely APX2009, APX2014, and FC12-531A) with improved potency significantly affected pancreatic cancer cell survival [65].…”

“…SLC-0111-mediated CAIX inhibition has proven to be effective in patient-derived 2D and 3D PDAC cell cultures [64,65], followed by an SLC-0111 safety study (NCT02215850) in advanced solid tumors. Currently, a Phase 1/2 clinical trial (NCT03450018) uses the selective, small molecule inhibitor SLC-0111 in CAIX-positive patients diagnosed with metastatic PDAC.…”