Development of Cancer-Stem-Cell Targeting Pan-Aldehyde Dehydrogenase Inhibitors as an Adjunct to Chemotherapy in the Treatment of Ovarian Cancer

Aldehyde Dehydrogenase (ALDH) activity is commonly used as a marker to identify cancer stem-like cells. The three ALDH1A isoforms have all been individually implicated in cancer stem-like cells and in chemoresistance; however, which isoform is preferentially expressed varies considerably between various cell lines and tumors. An inhibitor of these three isoforms could provide a useful broad-spectrum tool to study ALDH1A biology in any of these cells and could potentially be used as a therapeutic. The redundant expression and function of the ALDH1A isoforms also suggests that cells could develop resistance to a single isoform selective inhibitor by simply expressing an alternate isoform. Lead compound 673A, a pan ALDH1A inhibitor, selectively depleted cancer stem cells and reversed chemosensitivity in vivo. Based on this compelling activity we set out to optimize 673A and another HTS hit CM39 for ALDH1A potency and selectivity, cellular potency, and pharmacokinetic properties. Although progress in the development of 673A-based inhibitors has been limited, an affinity probe has been synthesized to uncover new targets of the potentially promiscuous compound and to search for protein-protein interactions involving ALDH. Our campaign to optimize CM39 with ALDH1A1 afforded first-in-class pan-inhibitors of ALDH1A1, 1A2, and 1A3 with excellent selectivity over ALDH2. We have developed compounds with up to 1000-fold improvement in inhibition of specific ALDH isoforms and 5-10 fold improved solubility and metabolic stability. Exemplary compounds exhibited potent cellular inhibition of ALDH, depleted the CD133+ putative cancer stem cell population, and were highly synergistic with cisplatin in patient derived ovarian cancer spheroids. We have also developed the most potent, selective ALDH1A3 inhibitor to date and have begun to define the structural features determining selectivity between the three ALDH1A isoforms.PHDMedicinal ChemistryUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttps://deepblue.lib.umich.edu/bitstream/2027.42/147713/1/bchuddle_1.pd

Structure-Based Optimization of a Novel Class of Aldehyde Dehydrogenase 1A (ALDH1A) Subfamily-Selective Inhibitors as Potential Adjuncts to Ovarian Cancer Chemotherapy

Aldehyde dehydrogenase (ALDH) activity is commonly used as a marker to identify cancer stem-like cells. The three ALDH1A isoforms have all been individually implicated in cancer stem-like cells and in chemoresistance; however, which isoform is preferentially expressed varies between cell lines. We sought to explore the structural determinants of ALDH1A isoform selectivity in a series of small-molecule inhibitors in support of research into the role of ALDH1A in cancer stem cells. An SAR campaign guided by a cocrystal structure of the HTS hit CM39 (7) with ALDH1A1 afforded first-in-class inhibitors of the ALDH1A subfamily with excellent selectivity over the homologous ALDH2 isoform. We also discovered the first reported modestly selective single isoform 1A2 and 1A3 inhibitors. Two compounds, 13g and 13h, depleted the CD133+ putative cancer stem cell pool, synergized with cisplatin, and achieved efficacious concentrations in vivo following IP administration. Compound 13h additionally synergized with cisplatin in a patient-derived ovarian cancer spheroid model

Tuning the Baird aromatic triplet-state energy of cyclooctatetraene to maximize the self-healing mechanism in organic fluorophores

Bright, photostable, and nontoxic fluorescent contrast agents are critical for biological imaging. "Self-healing" dyes, in which triplet states are intramolecularly quenched, enable fluorescence imaging by increasing fluorophore brightness and longevity, while simultaneously reducing the generation of reactive oxygen species that promote phototoxicity. Here, we systematically examine the self-healing mechanism in cyanine-class organic fluorophores spanning the visible spectrum. We show that the Baird aromatic triplet-state energy of cyclooctatetraene can be physically altered to achieve order of magnitude enhancements in fluorophore brightness and signal-to-noise ratio in both the presence and absence of oxygen. We leverage these advances to achieve direct measurements of large-scale conformational dynamics within single molecules at submillisecond resolution using wide-field illumination and camera-based detection methods. These findings demonstrate the capacity to image functionally relevant conformational processes in biological systems in the kilohertz regime at physiological oxygen concentrations and shed important light on the multivariate parameters critical to self-healing organic fluorophore design

A Pan-ALDH1A Inhibitor Induces Necroptosis in Ovarian Cancer Stem-like Cells

Summary: Ovarian cancer is typified by the development of chemotherapy resistance. Chemotherapy resistance is associated with high aldehyde dehydrogenase (ALDH) enzymatic activity, increased cancer “stemness,” and expression of the stem cell marker CD133. As such, ALDH activity has been proposed as a therapeutic target. Although it remains controversial which of the 19 ALDH family members drive chemotherapy resistance, ALDH1A family members have been primarily linked with chemotherapy resistant and stemness. We identified two ALDH1A family selective inhibitors (ALDH1Ai). ALDH1Ai preferentially kills CD133+ ovarian cancer stem-like cells (CSCs). ALDH1Ai induce necroptotic CSC death, mediated, in part, by the induction of mitochondrial uncoupling proteins and reduction in oxidative phosphorylation. ALDH1Ai is highly synergistic with chemotherapy, reducing tumor initiation capacity and increasing tumor eradication in vivo. These studies link ALDH1A with necroptosis and confirm the family as a critical therapeutic target to overcome chemotherapy resistance and improve patient outcomes. : Chefetz et al. identify ALDH1A enzyme family inhibitors (ALDH1Ai) that trigger necroptosis in CD133+ ovarian CSCs. Necroptosis is mediated in part by the induction of mitochondrial uncoupling proteins and reduction of oxidative phosphorylation. ALDH1Ai is highly synergistic with chemotherapy, reducing tumor initiation and growth and increasing tumor eradication rates. Keywords: ALDH, CSC, necroptosis, CD133+, cell death, ovarian cancer, resistanc