Comprehensive Target Engagement by the EZH2 Inhibitor Tulmimetostat Allows for Targeting of ARID1A Mutant Cancers.

Recurrent somatic mutations in the BAF chromatin remodeling complex subunit ARID1A occur frequently in advanced urothelial carcinoma, endometrial cancers, and ovarian clear cell carcinoma, creating an alternative chromatin state that may be exploited therapeutically. The histone methyltransferase EZH2 has previously been identified as targetable vulnerability in the context of ARID1A mutations. Here, we describe the discovery of tulmimetostat, an orally available, clinical stage EZH2 inhibitor and elucidate its therapeutic potential for treating ARID1A mutant tumors. Tulmimetostat administration achieved efficacy in multiple ARID1A mutant bladder, ovarian, and endometrial tumor models and improved cisplatin response in chemotherapy-resistant models. Consistent with its comprehensive and durable level of target coverage, tulmimetostat demonstrated greater efficacy than other PRC2-targeted inhibitors at comparable or lower exposures in a bladder cancer xenograft mouse model. Tulmimetostat mediated extensive changes in gene expression in addition to a profound reduction in global H3K27me3 levels in tumors. Phase I clinical pharmacokinetic and pharmacodynamic data indicated that tulmimetostat exhibits durable exposure and profound target engagement. Importantly, a tulmimetostat controlled gene expression signature identified in whole blood from a cohort of 32 cancer patients correlated with tulmimetostat exposure, representing a pharmacodynamic marker for the assessment of target coverage for PRC2-targeted agents in the clinic. Collectively, this data suggests that tulmimetostat has the potential to achieve clinical benefit in solid tumors as a monotherapy but also in combination with chemotherapeutic agents and may be beneficial in various indications with recurrent ARID1A mutations.

Utilization and toxicity patterns of 2-weekly (Q2W) versus 4-weekly (Q4W) nivolumab for treatment of adjuvant and metastatic melanoma at BC cancer.

BACKGROUND: Nivolumab, an immune checkpoint inhibitor used to treat several malignancies, is associated with immune-related adverse events (IrAEs). Original dosing for melanoma was 3 mg/kg (maximum 240 mg) every 2 weeks (Q2W). Based on simulation studies depicting similar efficacy and toxicity to original dosing, extended interval dosing of 6 mg/kg (maximum 480 mg) every 4 weeks (Q4W) was introduced. OBJECTIVE: This study will compare safety between Q2W and Q4W dosing at BC Cancer in melanoma patients. METHODS: Retrospective chart review for reported incidence, onset, and severity of IrAEs in melanoma patients treated with nivolumab Q2W and Q4W dosing was completed. Fisher's test was conducted for first incidence IrAEs using Microsoft Excel. RESULTS: Seventy-one patients were identified (Q2W n = 35, Q4W n = 36). Baseline characteristics were similar in both groups. No statistically significant difference was found in incidence of IrAEs between Q2W and Q4W dosing (Q2W 40% vs Q4W 50%, p = 0.477). Rash was most common (Q2W 79% vs Q4W 50%) followed by hypothyroidism (Q2W 33% vs Q4W 20%). Median onset of IrAEs seemed later with Q4W dosing (Q2W cycle 1 vs Q4W cycle 4). Regardless of dosing, most IrAEs were grade 1-2 in severity (Q2W 100% vs Q4W 89%). CONCLUSION: Q4W dosing is associated with comparable incidence and potentially later onset of IrAEs compared to Q2W dosing. Most IrAEs in both dosing groups were similar and mild. Therefore, Q4W dosing offers a safe alternative to Q2W dosing while providing benefits including decreased workload for staff, decreased clinic visits, and viral exposure by patients.

Farnesoid X receptor enhances epithelial ACE2 expression and inhibits virally induced IL-6 secretion: implications for intestinal symptoms of SARS-CoV-2.

Intestinal inflammation and diarrhea are often associated with SARS-CoV-2 infection. The angiotensin converting enzyme 2 (ACE2) receptor plays a key role in SARS-CoV-2 pathogenesis, facilitating entry of the virus into epithelial cells, while also regulating mucosal inflammatory responses. Here, we investigated roles for the nuclear bile acid receptor farnesoid X receptor (FXR) in regulating ACE2 expression and virally mediated inflammatory responses in intestinal epithelia. Human colonic or ileal enteroids and cultured T84 and Caco-2 monolayers were treated with the FXR agonists, obeticholic acid (OCA) or GW4064, or infected with live SARS-CoV-2 (2019-nCoV/USA_WA1/2020). Changes in mRNA, protein, or secreted cytokines were measured by qPCR, Western blotting, and ELISA. Treatment of undifferentiated colonic or ileal enteroids with OCA increased ACE2 mRNA by 2.1 ± 0.4-fold (n = 3; P = 0.08) and 2.3 ± 0.2-fold (n = 3; P < 0.05), respectively. In contrast, ACE2 expression in differentiated enteroids was not significantly altered. FXR activation in cultured epithelial monolayers also upregulated ACE2 mRNA, accompanied by increases in ACE2 expression and secretion. Further experiments revealed FXR activation to inhibit IL-6 release from both Caco-2 cells infected with SARS-CoV-2 and T84 cells treated with the viral mimic, polyinosinic:polycytidylic acid, by 46 ± 12% (n = 3, P < 0.05) and 35 ± 6% (n = 8; P < 0.01), respectively. By virtue of its ability to modulate epithelial ACE2 expression and inhibit virus-mediated proinflammatory cytokine release, FXR represents a promising target for the development of new approaches to prevent intestinal manifestations of SARS-CoV-2.NEW & NOTEWORTHY Activation of the nuclear bile acid receptor, farnesoid X receptor (FXR), specifically upregulates ACE2 expression in undifferentiated colonic epithelial cells and inhibits virus-induced proinflammatory cytokine release. By virtue of these actions FXR represents a promising target for the development of new approaches to prevent intestinal manifestations of SARS-CoV-2 infection.

Long-term outcomes associated with a modified versus traditional closed anal sacculectomy for treatment of canine anal sac neoplasia.

OBJECTIVE: To report the short-term and long-term outcomes of dogs that underwent the modified closed and traditional closed anal sacculectomy procedures for the treatment of anal sac neoplasia. ANIMALS: 90 client-owned dogs. Methods: The medical records of 2 tertiary referral hospitals were reviewed to identify dogs that underwent anal sacculectomy for treatment of anal sac neoplasia between January 2016 and December 2020. Data collected included signalment and preoperative diagnostic findings. The occurrence of intraoperative and postoperative complications, short-term outcomes, and long-term outcomes were also collected. Descriptive statistics were calculated to summarize dog signalment information, and recurrence, metastasis, and survival proportions were compared between techniques using Fisher exact tests. RESULTS: 35 and 55 dogs, respectively, underwent the modified or traditional closed anal sacculectomy procedure. Minor postoperative complications that resolved with minimal intervention occurred in 5 of 35 (14.3%) modified approach dogs and 12 of 55 (21.8%) traditional approach dogs. Tumor recurrence was confirmed in 8 of 35 (22.9%) modified and 8 of 55 (26.4%) traditional approach dogs and was suspected in 3 of 35 (8.6%) and 6 of 55 (13.2%; P = .68), respectively. Confirmed metastatic disease was identified in 8 of 35 (22.9%) and 14 of 53 (26.4%) modified and traditional approach dogs, respectively, and was suspected in 4 of 35 (11.4%) and 7 of 53 (13.2%). Sixty-three (70%) dogs survived to study conclusion. CLINICAL RELEVANCE: No benefits in complication rate or local recurrence were identified in dogs following the modified approach as opposed to the traditional closed anal sacculectomy technique.

Active enterohepatic cycling is not required for the choleretic actions of 24-norUrsodeoxycholic acid in mice.

The pronounced choleretic properties of 24-norUrsodeoxycholic acid (norUDCA) to induce bicarbonate-rich bile secretion have been attributed to its ability to undergo cholehepatic shunting. The goal of this study was to identify the mechanisms underlying the choleretic actions of norUDCA and the role of the bile acid transporters. Here, we show that the apical sodium-dependent bile acid transporter (ASBT), organic solute transporter-α (OSTα), and organic anion transporting polypeptide 1a/1b (OATP1a/1b) transporters are dispensable for the norUDCA stimulation of bile flow and biliary bicarbonate secretion. Chloride channels in biliary epithelial cells provide the driving force for biliary secretion. In mouse large cholangiocytes, norUDCA potently stimulated chloride currents that were blocked by siRNA silencing and pharmacological inhibition of calcium-activated chloride channel transmembrane member 16A (TMEM16A) but unaffected by ASBT inhibition. In agreement, blocking intestinal bile acid reabsorption by coadministration of an ASBT inhibitor or bile acid sequestrant did not impact norUDCA stimulation of bile flow in WT mice. The results indicate that these major bile acid transporters are not directly involved in the absorption, cholehepatic shunting, or choleretic actions of norUDCA. Additionally, the findings support further investigation of the therapeutic synergy between norUDCA and ASBT inhibitors or bile acid sequestrants for cholestatic liver disease.

Secondary (iso)BAs cooperate with endogenous ligands to activate FXR under physiological and pathological conditions.

IsoBAs, stereoisomers of primary and secondary BAs, are found in feces and plasma of human individuals. BA signaling via the nuclear receptor FXR is crucial for regulation of hepatic and intestinal physiology/pathophysiology. AIM: Investigate the ability of BA-stereoisomers to bind and modulate FXR under physiological/pathological conditions. METHODS: Expression-profiling, luciferase-assays, fluorescence-based coactivator-association assays, administration of (iso)-BAs to WT and cholestatic mice. RESULTS: Compared to CDCA/isoCDCA, administration of DCA/isoDCA, UDCA/isoUDCA only slightly increased mRNA expression of FXR target genes; the induction was more evident looking at pre-mRNAs. Notably, almost 50% of isoBAs were metabolized to 3-oxo-BAs within 4 h in cell-based assays, making it difficult to study their actions. FRET-based real-time monitoring of FXR activity revealed that isoCDCA>CDCA stimulated FXR, and isoDCA and isoUDCA allowed fully activated FXR to be re-stimulated by a second dose of GW4064. In vivo co-administration of a single dose of isoBAs followed by GW4064 cooperatively activated FXR, as did feeding of UDCA in a background of endogenous FXR ligands. However, in animals with biliary obstruction and concomitant loss of intestinal BAs, UDCA was unable to increase intestinal Fgf15. In contrast, mice with an impaired enterohepatic circulation of BAs (Asbt-/-, Ostα-/-), administration of UDCA was still able to induce ileal Fgf15 and repress hepatic BA-synthesis, arguing that UDCA is only effective in the presence of endogenous FXR ligands. CONCLUSION: Secondary (iso)BAs cooperatively activate FXR in the presence of endogenous BAs, which is important to consider in diseases linked to disturbances in BA enterohepatic cycling.

Cannabinoid receptor 2 signaling in peripheral immune cells modulates disease onset and severity in mouse models of Huntington's disease.

Peripheral immune cells and brain microglia exhibit an activated phenotype in premanifest Huntington's disease (HD) patients that persists chronically and correlates with clinical measures of neurodegeneration. However, whether activation of the immune system contributes to neurodegeneration in HD, or is a consequence thereof, remains unclear. Signaling through cannabinoid receptor 2 (CB(2)) dampens immune activation. Here, we show that the genetic deletion of CB(2) receptors in a slowly progressing HD mouse model accelerates the onset of motor deficits and increases their severity. Treatment of mice with a CB(2) receptor agonist extends life span and suppresses motor deficits, synapse loss, and CNS inflammation, while a peripherally restricted CB(2) receptor antagonist blocks these effects. CB(2) receptors regulate blood interleukin-6 (IL-6) levels, and IL-6 neutralizing antibodies partially rescue motor deficits and weight loss in HD mice. These findings support a causal link between CB(2) receptor signaling in peripheral immune cells and the onset and severity of neurodegeneration in HD, and they provide a novel therapeutic approach to treat HD.