Download A phase I trial to ... and antitumor activity of ursolic acid

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A phase I trial to evaluate the multiple-dose tolerability safety
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and
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patientssubjects with advanced solid tumors
antitumor
activity
of
ursolic
acid
liposomes
in
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Zhengzi Qian†, Xianhuo Wang†, Zheng Song, Huilai Zhang, Shiyong Zhou, Jing Zhao
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and Huaqing Wang*
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E- mail address:
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Zhengzi Qian
[email protected]
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Xianhuo Wang
[email protected]
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Zheng Song
[email protected]
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Huilai Zhang
[email protected]
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Shiyong Zhou
[email protected]
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Jing Zhao
[email protected]
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Huaqing Wang
[email protected]
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†
The authors contributed equally to this work.
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Department of Lymphoma, Sino-US Center for Lymphoma and Leukemia, Tianjin
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Medical University Cancer Institute and Hospital, National Clinical Research Center
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of Cancer, Key Laboratory of Cancer Prevention and Therapy, Tiyuanbei, Huanhuxi
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Road, Hexi District, Tianjin 300060, China.
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*
E-mail address for correspondence: [email protected].
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Abstract
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Background: Ursolic acid liposome (UAL), a new antitumor drug, has potential
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therapeutic value. However, limited clinical data exists regarding multiple-dose
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tolerabilitysafety, antitumor activity, and the recommended dose (RD) of UAL in a
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phase II trial of patientssubjects with advanced solid tumors.
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Methods: All patientssubjects were intravenously administered UAL for 14
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consecutive days of a 21-day treatment cycle. Twenty-one patientssubjects were
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enrolled in 1 of 3 sequential cohorts (56, 74, and 98 mg/m2 ) to evaluate multiple-dose
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tolerability and efficacy. Eight additional patientssubjects were treated with UAL (74
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mg/m2 ) to evaluate multiple-dose pharmacokinetics.
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Results: Treatment-related adverse events included elevated aspartate transaminase,
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alanine transaminase, gamma- glutamyl transpeptidase, and triglycerides levels;
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abdominal distention; pruritus; arthralgia; pyrexia; hypokalemia; and microscopic
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hematuria and proteinuria. However, no ≥grade 3 adverse events (NCI-CTC) were
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observed. Sixty percent patientssubjects achieved stable disease after 2 treatment
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cycles. Multiple-dose pharmacokinetic analysis revealed no significant differences in
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the elimination half- life, maximum plasma concentration, area under the plasma
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concentration time curve (AUC 0→t ), and AUC0→∞ values between days 1–14
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(P > 0.05), suggesting UAL does not accumulate in the body.
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Conclusions: This trial demonstrates that UAL was tolerable, had manageable
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toxicity, could potentially improve patient remission rates, and did not accumulate in
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the body. A large phase II study is recommended to confirm these results (i.e. RD of
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98 mg/m2 ).
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Trial registration: Chinese Clinical Trial Registry: ChiCTR-ONC-12002385.
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Keywords: Ursolic acid liposome; Phase I; Multiple-dose administration; Tolerability;
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Pharmacokinetics; Antitumor activity
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Introduction
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Ursolic acid (UA) is a natural hydroxy pentacyclic triterpene compound (Figure 1)
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isolated from Chinese herbs including Eriobotrya japonica, Rosmarinus officinalis,
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and Glechoma hederaceae [1, 2]. Previous studies have indicated that UA can induce
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apoptosis [3-5] and cell differentiation [6, 7], inhibit invasion and metastasis [8], and
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inhibit angiogenesis [9-11] in various tumors. UA treatment is also safe [12]. Thus,
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UA is a potentially valuable compound. However, the poor solubility of UA in
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hydrous solutions greatly limits its applications.
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Liposomes have been utilized as a drug delivery system to overcome the poor
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solubility of UA, increase the therapeutic efficiency, reduce the side-effects, and
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enhance the bioavailability of drugs that have been broadly applied [13-15]. Currently,
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ursolic acid liposomes (UALs) have been studied successfully and have been
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approved by the State Food and Drug Administration (SFDA) of China to enter
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clinical trials (NO. 2009L00634). We have previously published data regarding the
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maximum tolerated dose, dose- limiting toxicity (DLT), and pharmacokinetics of UAL
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in a single-dose administration study. The recommended doses in multiple-dose
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administration trials of UAL are 56, 74, and 98 mg/m2 [16]. In actuality, multiple-dose
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administration is usually adopted for most of drugs in clinic. Therefore, it is more
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important to study the effects of UALs in a multiple-dose administration trial.
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The primary objective of this study was to evaluate the tolerability of UAL treatment
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and the recommended dose (RD) in a multiple-dose administration phase II trial
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consisting of patientssubjects with advanced solid tumors. The second objective was
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to perform a preliminary assessment of the antitumor activity of UALs.
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Materials and Methods
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We performed a phase I, open-label, single center trial in patientssubjects with
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advanced solid tumors. The SFDA of China and the Hospital Medical Ethics
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Committees approved the trial and it was conducted in accordance with the
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Declaration of Helsinki and the applicable local regulatory requirements and laws. A
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signed, written informed consent of the legal representatives and the consent of each
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patient were obtained before any study procedure was performed. UALs were
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supplied by Wuhan Li Yuanheng Medicine Technology Co. Ltd (Wuhan, China) as a
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freeze-dried powder for infusion. Each glass vial contained 3 mg of active drug. It
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was uniformly dispersed in 250 mL of 5% glucose solution before administration.
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Patient eligibility
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Eligible patientssubjects were aged 18–75 years with cytologically or histologically
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confirmed advanced solid tumors; they either refused standard therapies or standard
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effective therapies did not exist; they had an Eastern Cooperative Oncology Group
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(ECOG) performance status (PS) of 0–2; a Karnofsky score ≥ 60%; a life
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expectancy ≥ 3 months; practiced adequate contraception; had adequate hematological
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function [white blood cell (WBC) ≥ 4.0 × 109 /L; absolute neutrophil count
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(ANC) ≥ 2.0 × 109 /L; platelet count ≥ 100 × 109 /L; hemoglobin ≥ 100 g/L]; had
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adequate hepatic and renal function [alanine transaminase (ALT), aspartate
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transaminase (AST), and alkaline phosphatase (ALP) ≤ 2.5 the upper limit of normal
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(ULN) (or 5 × ULN for hepatic cancer/metastatic hepatic cancer); total bilirubin
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(TBIL) ≤ 1.5 × ULN; serum creatinine (CRE) levels of ≤ 1.5 × ULN; a creatinine
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clearance rate of ≤ 1.5 × ULN; normal urea]; and had normal pulmonary function.
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Study design and treatment
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PatientsSubjects were assigned to 1 of 3 sequential dose cohorts of UAL: 56, 74, or
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98 mg/m2 , administered via a 14-day consecutive, intravenous 4 h infusion, and given
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a rest for 7-day per 21-day cycle. Each cohort consisted of at least 3 patientssubjects.
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Once all enrolled patientssubjects had been monitored for 2 weeks and had no higher
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than grade 3 non-hematological toxicity or grade 4 hematological toxicity, the next
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dose was administered. The trial was terminated when ≥1/3 of the patientssubjects
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experienced DLT, a severe adverse event (AE), or tumor progression. The DLT was
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defined as grade 4 thrombocytopenia, grade 4 neutropenia lasting for ≥7 days, febrile
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neutropenia, grade 4 anemia, or grade 3/4 non-hematological toxicity. Evaluated
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patientssubjects were required to complete at least 1 cycle of treatment. After that, if
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patientssubjects needed to continue treatment because they could not gain any benefit
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from other treatments, additional cycles were administered until disease progression
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or unacceptable toxicity occurred, or if the patient refused further treatment.
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Additional patientssubjects were recruited in order to evaluate the pharmacokinetics
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of UAL treatment. These patientssubjects were administered a dose of 74 mg/m2 of
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UAL via a consecutive, 14-day, intravenous 4 h infusion.
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Tolerability and toxicity
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Tolerability and toxicity were evaluated in all patientssubjects treated with at least 1
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cycle of UAL therapy. Vital signs including body temperature, respiration, pulsation,
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and blood pressure were examined at screening and once a day thereafter.
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Hematological parameters (red blood cell, WBC, hemoglobin, ANC, and platelet),
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urine routines (urinary protein, glucose, erythrocyte, leukocyte and urine bilirubin),
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and stool routines (fecal erythrocyte and fecal leukocyte) were tested, and an
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electrocardiogram performed at screening and on the 14th day of the cycle. Blood
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biochemistries including ALT, AST, ALP, gamma-glutamyl transpeptidase (GGT),
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TBIL, direct-reacting bilirubin, total protein, GLU, lactate dehydrogenase, creatine
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kinase, bun urea nitrogen, CRE, UA, cholesterol, triglyceride (TG), high-density
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lipoprotein, low-density lipoprotein, K +, Na+, Ca2+ and Cl- were examined at screening
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and then once a week thereafter. Fibrinogen (Fbg) and prothrombin time (PT) were
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examined at screening and during the 3rd week. To further evaluate the immune
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functions of patientssubjects after UAL administration, we measured CD4/CD8 and
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natural killer (NK) cell activity in the circulation both at screening and on the 14th day.
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AEs were evaluated according to the National Cancer Institute Common Terminology
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Criteria for AEs (NCI-CTCAE) version 3.0.
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Response evaluation
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Serial randomly patientssubjects treated with at least 2 cycles were selected to
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evaluate the therapeutic efficacy of UALs. The tumor response was examined by
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using computerized tomography, magnetic resonance imaging, chest radiography, or
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ultrasonography according to the response evaluation criteria in solid tumors
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(RECIST) at the scheduled times (baseline and 2 cycles later) either until the tumor
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progressed or until the final visit. Complete response (CR), partial response (PR),
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stable disease (SD), and progressive disease (PD) were defined according to
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RECIST.
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Multiple-dose pharmacokinetics
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Blood samples for pharmacokinetic analysis were collected into heparinized tubes on
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the 1st and 14th days of the study, at various time points including 0, 0.5, 1, 2 and 4 h
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during infusion, 5, 15 and 30 min, 1, 1.5, 2, 3, 4, 6, 8 and 12 h after the end of
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infusion. Plasma was separated using centrifugation and then stored at –20 °C until
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analysis.
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UAL concentrations were measured using validated ultra-performance liquid
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chromatography/tandem mass spectroscopy (UPLC/MS/MS) methods as described
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previously [17]. In brief, chromatography was performed using a Waters Acquity
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UPLCT M BEH C8 column (100 × 2.1 mm, 1.7 μm). The mobile phase consisted of
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acetonitrile and 10 mM ammonium formate (9:1, v/v) at a flow rate of 0.2 mL/min.
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The elution time was 3 m. Multiple-reaction monitoring was performed at m/z
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455.1→455.0 and m/z 469.3→425.2 for UAL and glycyrrhetinic acid (internal
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standard) respectively in negative ion mode with an electrospray ionization source.
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Estimates of pharmacokinetic parameters for UAL were derived from individual
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concentration-time data sets by non-compartmental analysis.
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Statistical conside rations
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Tolerability, toxicity, efficacy, and pharmacokinetic characteristics were explored and
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analyzed in detail. Non-compartmental pharmacokinetic parameters were determined
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from individual plasma concentration-time data using DAS version 2.1.1.
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Results
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Patient characte ristics
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Twenty-one patientssubjects (7 men and 14 women), aged 19–68 years (median age:
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54 years), were enrolled in the study, and their characteristics at baseline are listed in
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Table 1. Twenty patientssubjects (95%) had an ECOG performance status of 0–1. All
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patientssubjects were treated with surgery (43%), radiotherapy (52%), chemotherapy
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(14%), and/or other therapies (67%). The study included 5 (24%) patientssubjects
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with non-Hodgkin lymphoma, 5 (24%) patientssubjects with Hodgkin lymphoma, 1
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(5%)
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hepatomahepatocellular carcinoma, 1 (5%) patient with gallbladder carcinoma, 2 (9%)
patientssubjects
with
renal
carcinoma,
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1
(5%)
patient
with
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patientssubjects with breast cancer, 2 (9%) patientssubjects with lung cancer, and 4
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(19%) patientssubjects with other cancer.
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Tolerability and toxicity
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Tolerability and toxicity were evaluated for all patientssubjects. The vital sign data
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showed that all values fluctuated within the normal range at every time point among
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the 3 cohorts (Figure 2). All hematological parameters, (Fbg, PT) and results of
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electrocardiography and routine stool test were normal. Only 1 patient experienced
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grade 1 microscopic hematuria, while 2 patientssubjects developed grade 1 proteinuria
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after 2 cycles of treatment with UAL (74 mg/m2 ).
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Immune function tests showed no significant differences in CD4/CD8 at screening
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and on the 14th day (0.60 ± 0.31 and 0.82 ± 0.24, P > 0.05, 56 mg/m2 ; 0.82 ± 0.48 and
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0.61 ± 0.24, P > 0.05, 74 mg/m2 ; 1.39 ± 0.96 and 1.23 ± 0.23, P > 0.05, 98 mg/m2 ).
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Significant differences in the NK cells were also not observed (18.40 ± 7.66 and
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22.60 ± 5.97, P > 0.05, 56 mg/m2 ; 17.52 ± 11.57 and 20.87 ± 8.58, P > 0.05,
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74 mg/m2 ; 17.91 ± 10.02 and 18.40 ± 7.50, P > 0.05, 98 mg/m2 ). These results
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suggested that the UAL did not affect patient immune function.
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In addition, 3 (14%) patientssubjects treated with 56 mg/m2 UAL developed a
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low-grade fever (grade 1) but then recovered after 2 h without any treatment (Table 2).
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Three (14%) patientssubjects treated with 56, 74 and 98 mg/m2 UAL experienced
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grade 2 GGT elevation. Two (10%) patientssubjects treated with 56 and 74 mg/m2
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UAL experienced grade 1 abdominal distention. Finally, 1 (5%) patient had grade 2
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ALT elevation. Other mild symptoms including AST and TG elevation, pruritus,
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arthralgia, and hypokalemia were also observed. However, no National Cancer
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Institute common toxicity criteria (NCI-CTC) ≥ grade 3 treatment-related AEs were
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observed. The most frequent AEs included pyrexia, GGT elevation, and abdominal
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distention. These results suggested that UAL was tolerable and safe among 3 dose
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cohorts after administration via a consecutive 14-day intravenous 4 h infusion every
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21 days. Therefore, a UAL dose of 98 mg/m2 was considered the RD for a phase II
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trial.
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Efficacy
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As only 5 of 21 (23.8%) patientssubjects preferred to receive and finish at least 2
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cycles of UAL treatment, the evaluation of preliminary antitumor efficacy was limited.
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Three (60%) patientssubjects achieved stable disease. One of these patientssubjects
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had advanced renal carcinoma and had no significant change in the lesion after 2
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cycles of treatment with 56 mg/m2 UAL. Another patient that had advanced
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hepatomahepatocellular carcinoma had no significant change in the lesion after 2
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cycles of treatment with 74 mg/m2 UAL. Finally, the third patient had advanced lung
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cancer in which the lesion shrunk from 9.6–7.5 cm after 2 cycles of treatment with 98
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mg/m2 UAL.
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Two additional patientssubjects, 1 with primary non-Hodgkin lymphoma and the other
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with breast cancer, showed PD after 2 cycles treatment with 74 mg/m2 UAL. No CR
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and PR were observed, which could be because the patientssubjects had advanced
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stage tumors and did not benefit from other prior treatment schemes. Another possible
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explanation is that the number of patientssubjects that could be evaluated was too
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small. Regardless, UAL does have the potential to improve the patient remission rate.
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A phase II study of a large number of patientssubjects is recommended to confirm this
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finding.
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Multiple-dose pharmacokinetics
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Eight additional patientssubjects were enrolled in the trial in order to investigate the
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pharmacokinetics of UAL therapy. The pharmacokinetic data (Table 3) following
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multiple-dose administration showed that the values of the elimination half- life (t1/2 ),
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maximum plasma concentration (C max ), area under the plasma concentration time
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curve (AUC0→t ), and
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1589 ± 635 ng/mL, 5172 ± 1136 ng·h/mL, and 5498 ± 1525 ng·h/mL respectively.
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They were
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4834 ± 933 ng·h/mL respectively during the 14th day. There were no significant
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differences in the values of t1/2 , Cmax, AUC0→t , and AUC0→∞ (P > 0.05) between days
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1–14, suggesting that the pharmacokinetics were unaltered with multiple-daily dosing
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and that the UAL did not accumulate in the body. In addition, we found that there was
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a close relationship between the values of C max or AUC and AEs. The value of C max or
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AUC0→∞ during the 1st day were 4.58 ± 2.04
h,
1211 ± 204
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ng/mL,
4705 ± 873
ng·h/mL,
h,
and
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AUC increased as the AEs (including hepatotoxicity and abdominal distension)
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increased in seriousness.
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Discussion
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This study demonstrated that UAL treatment of patientssubjects with advanced solid
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tumors via multiple-dose and consecutive 14-day intravenous infusion every 21 days
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at doses of 56, 74, and 98 mg/m2 was safe. The results are consistent with preclinical
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information [12]. In addition, multiple-dose pharmacokinetics showed that the value
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of Cmax or AUC was associated with AEs. The value of Cmax or AUC was greater when
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the risk of AEs occurring in patientssubjects was elevated. The reasons for this might
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be the following: when the value of C max is elevated, hepatocytes would be exposed to
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a high concentration of drug and would be stimulated to release serial enzymes
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including AST, ALT, and GGT. If the value of AUC was high simultaneously, the time
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of stimulation would be prolonged. Therefore, the risk of hepatotoxicity and
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gastrointestinal toxicity would become elevated. These results suggested that the
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manageable toxicity associated with UAL treatment could be further controlled via
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kinetic monitoring.
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UA has been widely reported to have antitumor activities in preclinical studies [3-11,
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18-21]. However, the clinical antitumor effects of UA or UAL have not been reported
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previously. In our study, the preliminary antitumor activity of UAL was evaluated for
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the first time in 5 patientssubjects. Although no CR or PR occurred, SD was observed
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in 3 (60%) patientssubjects with advanced solid tumors. Specifically, 1 lung cancer
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patient showed significant improvement and the lesion decreased in size (range,
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9.6–7.5 cm) after 2 cycles of treatment with a UAL dose of 98 mg/m2 . These results
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indicate UAL can potentially improve patient remission.
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The pharmacokinetic data of UA in animals showed that T1/2 was about 4.3 h [22]. In
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this clinical trial, the mean T1/2 of UAL was 4.00-4.58 h, suggesting the T1/2 value was
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low so that it could rapidly eliminate from blood. This phenomenon suggested that
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UAL did not accumulate in the body, and that UAL must be infused repeatedly to
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keep the plasma-drug concentration steady and further enhance its antitumor effect.
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Conclusions
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In summary, the multiple-dose administration of UAL was tolerable with manageable
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toxicity. Further, the UAL did not accumulate in the body. We conclude that UAL has
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the potential to improve the patient remission rates. The recommended dose of UAL
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for a phase II clinical trial is 98 mg/m2 .
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Abbreviations
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UA: Ursolic acid; UAL: Ursolic acid liposome; RD: Recommended dose ; SFDA:
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State Food and Drug Administration; MTD: Maximum tolerated dose; DLT:
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Dose- limiting
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Performance status; AEs: AEs; NCI-CTCAE: National Cancer Institute Common
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Terminology Criteria for AEs; CT: Computerized tomography; MRI: Scan or
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magnetic resonance imaging; CR: Complete response; PR: Partial response; SD:
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Stable disease; PD: Progressive disease; UPLC/MS/MS: Ultra-performance liquid
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chromatography/tandem mass spectroscopy; EIS: Electrospray ionization source; Fbg:
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Fibrinogen; PT: Prothrombin time; t1/2: Elimination half- life; Cmax : Maximum plasma
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concentration; AUC: Area under the plasma concentration time curve
toxicity; ECOG: Eastern Cooperative Oncology Group; PS:
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Competing interests
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The author(s) declare(s) that there is no conflict of interests regarding the publication
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of this paper.
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Author Contributions
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The individual author contributions are following: ZQ contributed to the design and
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implementation of the study protocol. XW contributed to data analysis and to writing
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the manuscript. ZY contributed to data acquisition. ZS, HZ, SZ, LQ, JZ, and XM
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participated in patient recruitment. PW and XH contributed to the design of the
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protocol. HW designed the study and revised the manuscript. All the authors read and
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approved the final manuscript.
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Acknowledgme nts
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The authors greatly appreciate financial support from the Tianjin Medical University
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Cancer Hospital Doctoral Fund and the Tianjin Medical University Science Fund at
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our institution. In addition, the authors thank Wuhan Li Yuanheng Medicine
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Technology Co. Ltd (Wuhan, China) providing the UAL.
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the traditional chinese medicinal preparation Lu-Ying extract. Yakugaku
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Zasshi 2005, 125(6): 509-515.
411
412
413
21
414
Figures
415
Figure 1. The che mical structure of ursolic acid.
416
417
Figure 2. Vital sign data for the 3 cohorts at screening and throughout the
418
infusion. (A) Body temperature, (B) respiration, (C) pulsation, (D) diastolic pressure,
419
and (E) systolic pressure at the 3 different doses: 56 mg/m2 (n = 3), 74 mg/m2 (n = 14),
420
and 98 mg/m2 (n = 4), (mean ± SD).
421
422
423
424
425
426
427
22
428
Tables
429
Table 1. Patient characteristics at baseline.
PatientsSubjects
Characteristic
56 mg/m2 (n = 3) 74 mg/m2 (n = 14) 98 mg/m2 (n = 4)
Gender, n
Male
1
4
2
Female
2
10
2
Median age (range)
57 (49–59)
40.5 (19–68)
53.5 (42–59)
ECOG PS, n
0
2
9
1
1
1
4
3
2
1
Type of tumor, n
Non-Hodgkin
lymphoma
1
3
1
Hodgkin lymphoma
5
Renal carcinoma
1
HepatomaHepatocellular
carcinoma
1
Gallbladder carcinoma
1
Breast cancer
1
1
Lung cancer
2
Other
4
Prior therapy, n
Surgery
0
7
2
Radiotherapy
3
7
1
Chemotherapy
0
2
1
Other therapy
0
11
3
ECOG: Eastern Cooperative Oncology Group; PS: performance status
430
431
432
433
434
435
23
436
437
Table 2. Incidence of treatment-related adverse events.
Nu mber of patientssubjects
AE, N
2
56 mg/ m (n = 3)
2
74 mg/ m (n = 14)
98 mg/ m2 (n = 4)
Total (n = 21)
G1
G2
≥G3
G1
G2
≥G3
G1
G2
≥ G3
G1
G2
≥ G3
AST
-
-
-
1
-
-
-
-
-
1 (5%)
-
-
ALT
-
1
-
-
-
-
-
-
-
-
1 (5%)
-
GGT
-
1
-
-
1
-
-
1
-
-
3 (14%)
-
TG
-
-
-
1
-
-
-
-
-
1 (5%)
-
-
distention
1
-
-
1
-
-
-
-
-
2 (10%)
-
-
Pruritus
-
-
-
1
-
-
-
-
-
1 (5%)
-
-
Arthralgia
-
-
-
1
-
-
-
-
-
1 (5%)
-
-
Low-grade fever
3
-
-
-
-
-
-
-
-
3 (14%)
-
-
Hypokalemia
-
-
-
1
-
-
-
-
-
1 (5%)
-
-
Hepatotoxicity
Abdominal
G1, G2, and G3 represent grade 1, grade 2, and grade 3 respectively, according to NCI-CTC grades.
AE: adverse event; -: no occurrence.
438
439
440
441
442
443
444
445
446
447
448
449
24
450
451
Table 3. Ursolic acid liposome pharmacokinetic parameters for the 1st and 14th
452
days (mean ± standard deviation, SD; n = 8).
Parameter
Day 1
Mean ± SD
4.58
±
2.04
Unit
t1/2
h
Vd
L/m2
2
Day 14
Mean ± SD
4.00
±
1.27
88.60
±
31.80
89.90
±
28.10
CL
L/(h·m )
14.40
±
3.94
15.80
±
3.05
AUC(0-t)
ng·h/mL
5172
±
1136
4705
±
873
AUC(0-∞)
ng·h/mL
5498
±
1525
4834
±
933
MRT(0-t)
h
3.34
±
0.55
3.30
±
0.31
MRT(0-∞)
h
4.31
±
1.89
3.78
±
0.70
Tmax
h
3.00
±
1.41
3.63
±
1.06
Cmax
ng/mL
1589
±
635
1211
±
204
453
25