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Loss of treatment-induced ovarian carcinoma (OC) growth suppression poses a major clinical challenge because it leads to disease recurrence. Therefore, new and well-tolerated approaches to maintain OC suppression after standard-of-care treatment are needed. We have profiled ascites as OC tumor microenvironments (TMs) to search for potential components that would exert growth suppression on OC cell cultures. Our investigations revealed that low levels of taurine, a non-proteogenic sulfonic amino acid, were present within OC ascites. Taurine supplementation, beyond levels found in ascites, induced growth suppression without causing cytotoxicity in multiple OC cell cultures, including patient-derived chemotherapy-resistant spheroid and organoid cultures. Suppression of proliferation by taurine was associated with the inhibition of glycolysis, mitochondrial function, and the activation of p21 and TIGAR, theTP53-dependent and independent tumor suppression regulatory pathways. Expression of p21 or TIGAR in various OC cells, in part, mimicked taurine-induced inhibition of OC cell proliferation. Our data support the potential therapeutic value of taurine supplementation in OC after chemotherapy.

High-Grade Serous Carcinoma (HGSC) is the most common and lethal ovarian cancer subtype. PARP-inhibitors (PARPi) have become the mainstay of HGSC targeted therapy, given that these tumors are driven by a high degree of genomic instability and Homologous Recombination (HR) defects. Nonetheless, only ∼30% of patients initially respond to treatment, ultimately relapsing with resistant disease. Thus, despite recent advances in drug development and increased understanding of genetic alterations driving HGSC progression, mortality has not declined, highlighting the need for novel therapies. Using a Small Molecule Activator of Protein Phosphatase 2A (PP2A) (SMAP-061), we investigated the mechanism by which PP2A stabilization induces apoptosis in Patient-Derived HGSC cells and Xenograft (PDX) models alone or in combination with PARPi. We uncovered that PP2A genes essential for transformation (B56α,B56γ and PR72) and basal phosphatase activity (PP2A-A and -C) are heterozygously lost in the majority of HGSC. Moreover, loss of these PP2A genes correlates with worse overall patient survival. We show that SMAP-061 stabilization of PP2A inhibits the HR output by targeting RAD51, leading to chronic accumulation of DNA damage and ultimately apoptosis. Furthermore, combination of SMAP-061 and PARPi leads to enhanced apoptosis in both HR-proficient and -deficient cells and in patient-derived xenograft models. Our studies identify PP2A as novel regulator of HR and introduces PP2A activators as a potential treatment for HGSC tumors. Our studies further emphasize the potential of PP2A modulators to overcome PARPi insensitivity, given that targeting RAD51 has presented benefits in overcoming PARPi-resistance driven by BRCA1/2 mutation reversions.

Dysregulation of cap-dependent translation is a hallmark of cancer, with key roles in supporting the transformed phenotype. The eIF4E binding proteins (4E-BP1, 2, 3) are major negative regulators of cap-dependent translation that are inactivated in tumors through inhibitory phosphorylation by oncogenic kinases (e.g., mTOR) or by downregulation. Previous studies from our group and others have linked tumor suppressive PP2A family serine/threonine phosphatases to activation of 4E-BP1. Here, we leveraged novel small molecule activators of PP2A (SMAPs) (e.g., DT-061, DT-1154) that are being developed as antitumor agents to (a) explore the role of a subset of B56-PP2As in regulation of 4E-BP activity, and (b) to evaluate the potential of B56-PP2A reactivation for restoring translation control in tumor cells. We show that SMAPs promote PP2A-dependent hypophosphorylation of 4E-BP1/4EBP2 in the presence of active upstream inhibitory kinases (mTOR, ERK, AKT), supporting a role for B56-PP2As as 4E-BP phosphatases. Unexpectedly, DT-061 also led to robust PP2A-dependent upregulation of 4E-BP1, but not 4E-BP2 or 4E-BP3. Cap-binding assays and eIF4E immunoprecipitation showed that SMAP/B56-PP2A blocks the formation of the eIF4F translation initiation complex. Bicistronic reporter assays that directly measure cap-dependent translation activity confirmed the translational consequences of these effects. siRNA knockdown pointed to B56α-PP2A as a mediator of SMAP effects on 4E-BPs, although B56β- and/or B56ε-PP2A may also play a role. 4E-BP1 upregulation involved ATF4-dependent transcription of the 4E-BP1 gene (EIF4EBP1) and the effect was partially dependent on TFE3/TFEB transcription factors. Thus, B56-PP2A orchestrates a translation repressive program involving transcriptional induction and hypophosphorylation of 4E-BP1, highlighting the potential of PP2A-based therapeutic strategies for restoration of translation control in cancer cells.

Metastasis results from a complex set of traits acquired by tumor cells, distinct from those necessary for tumorigenesis. Here, we investigate the contribution of enhancer elements to the metastatic phenotype of osteosarcoma. Through epigenomic profiling, we identify substantial differences in enhancer activity between primary and metastatic tumors in human patients as well as nearisogenic pairs of high and low lung-metastatic osteosarcoma cells. We term these regions Metastatic Variant Enhancer Loci (Met-VELs). We demonstrate that these Met-VELs drive coordinated waves of gene expression during metastatic colonization of the lung. Met-VELs cluster non-randomly, indicating that activity of these enhancers and their associated gene targets are positively selected. As evidence of this causal association, osteosarcoma lung metastasis is inhibited by global interruptions of Met-VEL-associated gene expression via pharmacologic BET inhibition, by knockdown of AP-1 transcription factors that occupy Met-VELs, and by knockdown or functional inhibition of individual genes activated by Met-VELs, such as F3. We further show that genetic deletion of a single Met-VEL at the F3 locus blocks metastatic cell outgrowth in the lung. These findings indicate that Met-VELs and the genes they regulate play a functional role in metastasis and may be suitable targets for anti-metastatic therapies.