Precision-activated T-cell engagers targeting HER2 or EGFR and CD3 mitigate on-target, off-tumor toxicity for immunotherapy in solid tumors.

To enhance the therapeutic index of T-cell engagers (TCEs), we engineered masked, precision-activated TCEs (XPAT proteins), targeting a tumor antigen (human epidermal growth factor receptor 2 (HER2) or epidermal growth factor receptor (EGFR)) and CD3. Unstructured XTEN polypeptide masks flank the N and C termini of the TCE and are designed to be released by proteases in the tumor microenvironment. In vitro, unmasked HER2-XPAT (uTCE) demonstrates potent cytotoxicity, with XTEN polypeptide masking providing up to 4-log-fold protection. In vivo, HER2-XPAT protein induces protease-dependent antitumor activity and is proteolytically stable in healthy tissues. In non-human primates, HER2-XPAT protein demonstrates a strong safety margin (>400-fold increase in tolerated maximum concentration versus uTCE). HER2-XPAT protein cleavage is low and similar in plasma samples from healthy and diseased humans and non-human primates, supporting translatability of stability to patients. EGFR-XPAT protein confirmed the utility of XPAT technology for tumor targets more widely expressed in healthy tissues.

First-in-Human Phase I Study to Evaluate the Brain-Penetrant PI3K/mTOR Inhibitor GDC-0084 in Patients with Progressive or Recurrent High-Grade Glioma.

PURPOSE: GDC-0084 is an oral, brain-penetrant small-molecule inhibitor of PI3K and mTOR. A first-in-human, phase I study was conducted in patients with recurrent high-grade glioma. PATIENTS AND METHODS: GDC-0084 was administered orally, once daily, to evaluate safety, pharmacokinetics (PK), and activity. Fluorodeoxyglucose-PET (FDG-PET) was performed to measure metabolic responses. RESULTS: Forty-seven heavily pretreated patients enrolled in eight cohorts (2-65 mg). Dose-limiting toxicities included 1 case of grade 2 bradycardia and grade 3 myocardial ischemia (15 mg), grade 3 stomatitis (45 mg), and 2 cases of grade 3 mucosal inflammation (65 mg); the MTD was 45 mg/day. GDC-0084 demonstrated linear and dose-proportional PK, with a half-life (∼19 hours) supportive of once-daily dosing. At 45 mg/day, steady-state concentrations exceeded preclinical target concentrations producing antitumor activity in xenograft models. FDG-PET in 7 of 27 patients (26%) showed metabolic partial response. At doses ≥45 mg/day, a trend toward decreased median standardized uptake value in normal brain was observed, suggesting central nervous system penetration of drug. In two resection specimens, GDC-0084 was detected at similar levels in tumor and brain tissue, with a brain tissue/tumor-to-plasma ratio of >1 and >0.5 for total and free drug, respectively. Best overall response was stable disease in 19 patients (40%) and progressive disease in 26 patients (55%); 2 patients (4%) were nonevaluable. CONCLUSIONS: GDC-0084 demonstrated classic PI3K/mTOR-inhibitor related toxicities. FDG-PET and concentration data from brain tumor tissue suggest that GDC-0084 crossed the blood-brain barrier.

Alternative dosing regimens for atezolizumab: an example of model-informed drug development in the postmarketing setting.

PURPOSE: To determine the exposure-response (ER) relationships between atezolizumab exposure and efficacy or safety in patients with advanced non-small cell lung cancer (NSCLC) or urothelial carcinoma (UC) and to identify alternative dosing regimens. METHODS: ER analyses were conducted using pooled NSCLC and UC data from phase 1 and 3 studies (PCD4989g, OAK, IMvigor211; ClinicalTrials.gov IDs, NCT01375842, NCT02008227, and NCT02302807, respectively). Objective response rate, overall survival, and adverse events were evaluated vs pharmacokinetic (PK) metrics. Population PK-simulated exposures for regimens of 840 mg every 2 weeks (q2w) and 1680 mg every 4 weeks (q4w) were compared with the approved regimen of 1200 mg every 3 weeks (q3w) and the maximum assessed dose (MAD; 20 mg/kg q3w). Phase 3 IMpassion130 (NCT02425891) data were used to validate the PK simulations for 840 mg q2w. Observed safety data were evaluated by exposure and body weight subgroups. RESULTS: No significant ER relationships were observed for safety or efficacy. Predicted exposures for 840 mg q2w and 1680 mg q4w were comparable to 1200 mg q3w and the MAD and consistent with observed PK data from IMpassion130. Observed safety was similar between patients with a Cmax above and below the predicted Cmax for 1680 mg q4w and between patients in the lowest and upper 3 body weight quartiles. CONCLUSION: Atezolizumab regimens of 840 mg q2w and 1680 mg q4w are expected to have comparable efficacy and safety as the approved regimen of 1200 mg q3w, supporting their interchangeable use and offering patients greater flexibility.

Investigation of the absolute bioavailability and human mass balance of navoximod, a novel IDO1 inhibitor.

AIMS: Navoximod (GDC-0919, NLG-919) is a small molecule inhibitor of indoleamine-2,3-dioxygenase 1 (IDO1), developed to treat the acquired immune tolerance associated with cancer. The primary objectives of this study were to assess navoximod's absolute bioavailability (aBA), determine the mass balance and routes of elimination of [14 C]-navoximod, and characterize navoximod's metabolite profile. METHODS: A phase 1, open-label, two-part study was conducted in healthy volunteers. In Part 1 (aBA), subjects (n = 16) were randomized to receive oral (200 mg tablet) or intravenous (5 mg solution) navoximod in a crossover design with a 5-day washout. In Part 2 (mass balance), subjects (n = 8) were administered [14 C]-navoximod (200 mg/600 µCi) as an oral solution. RESULTS: The aBA of navoximod was estimated to be 55.5%, with a geometric mean (%CV) plasma clearance and volume of distribution of 62.0 L/h (21.0%) and 1120 L (28.4%), respectively. Mean recovery of total radioactivity was 87.8%, with 80.4% detected in urine and the remainder (7.4%) in faeces. Navoximod was extensively metabolized, with unchanged navoximod representing 5.45% of the dose recovered in the urine and faeces. Glucuronidation was identified as the primary route of metabolism, with the major glucuronide metabolite, M28, accounting for 57.5% of the total drug-derived exposure and 59.7% of the administered dose recovered in urine. CONCLUSIONS: Navoximod was well tolerated, quickly absorbed and showed moderate bioavailability, with minimal recovery of the dose as unchanged parent in the urine and faeces. Metabolism was identified as the primary route of clearance and navoximod glucuronide (M28) was the most abundant metabolite in circulation with all other metabolites accounting for <10% of drug-related exposure.

A phase Ib study of pictilisib (GDC-0941) in combination with paclitaxel, with and without bevacizumab or trastuzumab, and with letrozole in advanced breast cancer.

BACKGROUND: This phase Ib study (NCT00960960) evaluated pictilisib (GDC-0941; pan-phosphatidylinositol 3-kinase inhibitor) plus paclitaxel, with and without bevacizumab or trastuzumab, or in combination with letrozole, in patients with locally recurrent or metastatic breast cancer. METHODS: This was a three-part multischedule study. Patients in parts 1 and 2, which comprised 3 + 3 dose escalation and cohort expansion stages, received pictilisib (60-330 mg) plus paclitaxel (90 mg/m2) with and without bevacizumab (10 mg/kg) or trastuzumab (2-4 mg/kg). In part 3, patients received pictilisib (260 mg) plus letrozole (2.5 mg). Primary objectives were evaluation of safety and tolerability, identification of dose-limiting toxicities (DLTs) and the maximum tolerated dose (MTD) of pictilisib, and recommendation of a phase II dosing regimen. Secondary endpoints included pharmacokinetics and preliminary antitumor activity. RESULTS: Sixty-nine patients were enrolled; all experienced at least one adverse event (AE). Grade ≥ 3 AEs, serious AEs, and AEs leading to death were reported in 50 (72.5%), 21 (30.4%), and 2 (2.9%) patients, respectively. Six (8.7%) patients reported a DLT, and the MTD and recommended phase II pictilisib doses were established where possible. There was no pictilisib-paclitaxel drug-drug interaction. Two (3.4%) patients experienced complete responses, and 17 (29.3%) patients had partial responses. CONCLUSIONS: Combining pictilisib with paclitaxel, with and without bevacizumab or trastuzumab, or letrozole, had a manageable safety profile in patients with locally recurrent or metastatic breast cancer. The combination had antitumor activity, and the additive effect of pictilisib supported further investigation in a randomized study. TRIAL REGISTRATION: ClinicalTrials.gov, NCT00960960 . Registered on August 13, 2009.

Phase Ia study of the indoleamine 2,3-dioxygenase 1 (IDO1) inhibitor navoximod (GDC-0919) in patients with recurrent advanced solid tumors.

BACKGROUND: Indoleamine-2,3-dioxygenase 1 (IDO1) catalyzes the oxidation of tryptophan into kynurenine and is partially responsible for acquired immune tolerance associated with cancer. The IDO1 small molecule inhibitor navoximod (GDC-0919, NLG-919) is active as a combination therapy in multiple tumor models. METHODS: This open-label Phase Ia study assessed safety, pharmacokinetics (PK), pharmacodynamics (PD), and preliminary anti-tumor activity of navoximod in patients with recurrent/advanced solid tumors, administered as 50-800 mg BID on a 21/28 day and at 600 mg on a 28/28 day schedule. Plasma kynurenine and tryptophan were longitudinally evaluated and tumor assessments were performed. RESULTS: Patients (n = 22) received a median of 3 cycles of navoximod. No maximum tolerated dose was reached. One dose-limiting toxicity of Grade 4 lower gastrointestinal hemorrhage was reported. Adverse events (AEs) regardless of causality in ≥20% of patients included fatigue (59%), cough, decreased appetite, and pruritus (41% each), nausea (36%), and vomiting (27%). Grade ≥ 3 AEs occurred in 14/22 patients (64%), and were related to navoximod in two patients (9%). Navoximod was rapidly absorbed (Tmax ~ 1 h) and exhibited dose-proportional increases in exposure, with a half-life (t1/2 ~ 11 h) supportive of BID dosing. Navoximod transiently decreased plasma kynurenine from baseline levels with kinetics consistent with its half-life. Of efficacy-evaluable patients, 8 (36%) had stable disease and 10 (46%) had progressive disease. CONCLUSIONS: Navoximod was well-tolerated at doses up to 800 mg BID decreasing plasma kynurenine levels consistent with its half-life. Stable disease responses were observed. TRIAL REGISTRATION: ClinicalTrials.gov identifier: NCT02048709 .

Combining "Bottom-up" and "Top-down" Approaches to Assess the Impact of Food and Gastric pH on Pictilisib (GDC-0941) Pharmacokinetics.

Pictilisib, a weakly basic compound, is an orally administered, potent, and selective pan-inhibitor of phosphatidylinositol 3-kinases for oncology indications. To investigate the significance of high-fat food and gastric pH on pictilisib pharmacokinetics (PK) and enable label recommendations, a dedicated clinical study was conducted in healthy volunteers, whereby both top-down (population PK, PopPK) and bottom-up (physiologically based PK, PBPK) approaches were applied to enhance confidence of recommendation and facilitate the clinical development through scenario simulations. The PopPK model identified food (for absorption rate constant (Ka )) and proton pump inhibitors (PPI, for relative bioavailability (Frel ) and Ka ) as significant covariates. Food and PPI also impacted the variability of Frel . The PBPK model accounted for the supersaturation tendency of pictilisib, and gastric emptying physiology successfully predicted the food and PPI effect on pictilisib absorption. Our research highlights the importance of applying both quantitative approaches to address critical drug development questions.

Genome-wide discovery of drug-dependent human liver regulatory elements.

Inter-individual variation in gene regulatory elements is hypothesized to play a causative role in adverse drug reactions and reduced drug activity. However, relatively little is known about the location and function of drug-dependent elements. To uncover drug-associated elements in a genome-wide manner, we performed RNA-seq and ChIP-seq using antibodies against the pregnane X receptor (PXR) and three active regulatory marks (p300, H3K4me1, H3K27ac) on primary human hepatocytes treated with rifampin or vehicle control. Rifampin and PXR were chosen since they are part of the CYP3A4 pathway, which is known to account for the metabolism of more than 50% of all prescribed drugs. We selected 227 proximal promoters for genes with rifampin-dependent expression or nearby PXR/p300 occupancy sites and assayed their ability to induce luciferase in rifampin-treated HepG2 cells, finding only 10 (4.4%) that exhibited drug-dependent activity. As this result suggested a role for distal enhancer modules, we searched more broadly to identify 1,297 genomic regions bearing a conditional PXR occupancy as well as all three active regulatory marks. These regions are enriched near genes that function in the metabolism of xenobiotics, specifically members of the cytochrome P450 family. We performed enhancer assays in rifampin-treated HepG2 cells for 42 of these sequences as well as 7 sequences that overlap linkage-disequilibrium blocks defined by lead SNPs from pharmacogenomic GWAS studies, revealing 15/42 and 4/7 to be functional enhancers, respectively. A common African haplotype in one of these enhancers in the GSTA locus was found to exhibit potential rifampin hypersensitivity. Combined, our results further suggest that enhancers are the predominant targets of rifampin-induced PXR activation, provide a genome-wide catalog of PXR targets and serve as a model for the identification of drug-responsive regulatory elements.

Expanding the role of the dynein regulatory complex to non-axonemal functions: association of GAS11 with the Golgi apparatus.

The mammalian GAS11 gene is a candidate tumor suppressor of unknown function that was previously identified as one of several genes upregulated upon growth arrest. Interestingly, although GAS11 homologs in Trypanosoma brucei (trypanin) and Chlamydomonas reinhardtii (PF2) are integral components of the flagellar axoneme and are necessary for regulating flagellar beat, the GAS11 gene was discovered based on its expression in cells that do not assemble a motile cilium. This suggests that GAS11 function might not be restricted to the cilium. To investigate this possibility, we generated GAS11-specific antibodies and demonstrate here that GAS11 is expressed in a variety of mammalian cells that lack a motile cilium. In COS7 cells, GAS11 is associated with the detergent-insoluble cytoskeleton and exhibits a juxtanuclear localization that overlaps with the pericentrosomal Golgi apparatus. This localization is dependent upon intact microtubules and is cell-cycle regulated, such that GAS11 is dispersed throughout the cytoplasm as cells progress through mitosis. GAS11 remains associated with Golgi fragments following depolymerization of cytoplasmic microtubules but is dispersed upon disruption of the Golgi with brefeldin A. These data suggest that GAS11 is associated with the Golgi apparatus. In support of this, recombinant GAS11 binds Golgi membranes in vitro. In growth-arrested mIMCD3 cells, GAS11 co-localizes with gamma-tubulin at the base of the primary cilium. The pericentrosomal Golgi apparatus and base of the cilium both represent convergence points for microtubule minus ends and correspond to sites where dynein regulation is required. The algal GAS11 homolog functions as part of a dynein regulatory complex (DRC) in the axoneme (Rupp and Porter. J Cell Biol 2003;162:47-57) and our findings suggest that components of this axonemal dynein regulatory system have been adapted in mammalian cells to participate in non-axonemal functions.