<div>AbstractPurpose:<p>Only one chemical class of topoisomerase I (TOP1) inhibitors is FDA approved, the camptothecins with irinotecan and topotecan widely used. Because of their limitations (chemical instability, drug efflux-mediated resistance, and diarrhea), novel TOP1 inhibitors are warranted. Indenoisoquinoline non-camptothecin topoisomerase I (TOP1) inhibitors overcome chemical instability and drug resistance that limit camptothecin use. Three indenoisoquinolines, LMP400 (indotecan), LMP776 (indimitecan), and LMP744, were examined in a phase I study for lymphoma-bearing dogs to evaluate differential efficacy, pharmacodynamics, toxicology, and pharmacokinetics.</p>Experimental Design:<p>Eighty-four client-owned dogs with lymphomas were enrolled in dose-escalation cohorts for each indenoisoquinoline, with an expansion phase for LMP744. Efficacy, tolerability, pharmacokinetics, and target engagement were determined.</p>Results:<p>The MTDs were 17.5 mg/m<sup>2</sup> for LMP 776 and 100 mg/m<sup>2</sup> for LMP744; bone marrow toxicity was dose-limiting; up to 65 mg/m<sup>2</sup> LMP400 was well-tolerated and MTD was not reached. None of the drugs induced notable diarrhea. Sustained tumor accumulation was observed for LMP744; γH2AX induction was demonstrated in tumors 2 and 6 hours after treatment; a decrease in TOP1 protein was observed in most lymphoma samples across all compounds and dose levels, which is consistent with the fact that tumor response was also observed at low doses LMP744. Objective responses were documented for all indenoisoquinolines; efficacy (13/19 dogs) was greatest for LMP744.</p>Conclusions:<p>These results demonstrate proof-of-mechanism for indenoisoquinoline TOP1 inhibitors supporting their further clinical development. They also highlight the value of the NCI Comparative Oncology Program (<a href="https://ccr.cancer.gov/Comparative-Oncology-Program" target="_blank">https://ccr.cancer.gov/Comparative-Oncology-Program</a>) for evaluating novel therapies in immunocompetent pets with cancers.</p></div>
<p>Figure SF2: cfDNA mutational analysis cfDNA from plasma collected upon enrollment was analyzed for mutations with a 74-gene Guardant panel (1). Alterations were identified in 30 genes as shown in the figure. Patient IDs: Green=Stable Disease, Red=Progressive Disease, and Black=non-evaluable. Specific patients are discussed below and in the main text.</p>
To determine the maximum-tolerated dose (MTD) of the histone deacetylase inhibitor vorinostat combined with fixed doses of cytarabine (ara-C or cytosine arabinoside) and etoposide in patients with poor-risk or advanced acute leukemia, to obtain preliminary efficacy data, describe pharmacokinetics, and in vivo pharmacodynamic effects of vorinostat in leukemia blasts.In this open-label phase I study, vorinostat was given orally on days one to seven at three escalating dose levels: 200 mg twice a day, 200 mg three times a day, and 300 mg twice a day. On days 11 to 14, etoposide (100 mg/m(2)) and cytarabine (1 or 2 g/m(2) twice a day if ≥65 or <65 years old, respectively) were given. The study used a standard 3+3 dose escalation design.Eighteen of 21 patients with acute myelogenous leukemia (AML) treated on study completed planned therapy. Dose-limiting toxicities [hyperbilirubinemia/septic death (1) and anorexia/fatigue (1)] were encountered at the 200 mg three times a day level; thus, the MTD was established to be vorinostat 200 mg twice a day. Of 21 patients enrolled, seven attained a complete remission (CR) or CR with incomplete platelet recovery, including six of 13 patients treated at the MTD. The median remission duration was seven months. No differences in percentage S-phase cells or multidrug resistance transporter (MDR1 or BCRP) expression or function were observed in vivo in leukemia blasts upon vorinostat treatment.Vorinostat 200 mg twice a day can be given safely for seven days before treatment with cytarabine and etoposide. The relatively high CR rate seen at the MTD in this poor-risk group of patients with AML warrants further studies to confirm these findings.
<p>Table S1: Schedule of pharmacokinetic analysis; Table S2: Overview of study dose levels (N=50); Table S3. Adverse events reported at any time during study therapy: grade 3-4 adverse events; Table S4: Veliparib pharmacokinetic (PK) parameters by cohort; Table S5: Total and ultrafilterable platinum pharmacokinetic parameters by cohort; Table S6: Immunohistochemistry results for 28 pre-therapy samples; Figure S1: Veliparib Cmax (ng/mL, panel A) and veliparib AUC0-6 (hr*ng/mL, panel B) by dose Cohort; Figure S2: Veliparib pharmacokinetic parameters (A) Cmax and (B) AUC0-6 at cycles 1 and 4 by cohort; Figure S3: Ultrafilterable platinum pharmacokinetic parameters Cmax and AUC0-24 at cycles 1 and 4 by cohort; Figure S4: Consort 2010 flow diagram for exploratory efficacy analysis comparing germline BRCA mutation positive to wild type; Figure S5: PAR assay results (pg/mL) for 14 patients in dose cohorts 6-8 (120-200 mg)</p>
<p>Table S1. Changes in Proportion (%) of γH2AX-Positive Cells in Peripheral Blood upon Veliparib and Veliparib/Temozolomide Treatment Figure S1. FANCD2 monoubiquitination following ex vivo melphalan treatment of patient bone marrow cells Figure S2.Pretreatment levels of PARP1 and MGMT Figure S3.Pretreatment MGMT promoter methylation in patient peripheral blood (PB) or bone marrow (BM) AML cells Figure S4. PARP inhibition in peripheral blood cells</p>
To support a phase III randomized trial of the multi-targeted tyrosine kinase inhibitor cabozantinib in neuroendocrine tumors, we developed a high-performance liquid chromatography mass spectrometry method to quantitate cabozantinib in 50 μL of human plasma. After acetonitrile protein precipitation, chromatographic separation was achieved with a Phenomenex synergy polar reverse phase (4 μm, 2 × 50 mm) column and a gradient of 0.1% formic acid in acetonitrile and 0.1% formic acid in water over a 5-min run time. Detection was performed on a Quattromicro quadrupole mass spectrometer with electrospray, positive-mode ionization. The assay was linear over the concentration range 50-5000 ng/mL and proved to be accurate (103.4-105.4%) and precise (<5.0%CV). Hemolysis (10% RBC) and use of heparin as anticoagulant did not impact quantitation. Recovery from plasma varied between 103.0-107.7% and matrix effect was -47.5 to -41.3%. Plasma freeze-thaw stability (97.7-104.9%), stability for 3 months at -80°C (103.4-111.4%), and stability for 4 h at room temperature (100.1-104.9%) were all acceptable. Incurred sample reanalysis of (N = 64) passed: 100% samples within 20% difference, -0.7% median difference and 1.1% median absolute difference. External validation showed a bias of less than 1.1%. This assay will help further define the clinical pharmacokinetics of cabozantinib.
