Clinical
`Cancer
`Research
`
`Cancer Therapy: Clinical
`
`Weekly nab-Rapamycin in Patients with Advanced
`Nonhematologic Malignancies: Final Results of a Phase I Trial
`
`Ana M. Gonzalez-Angulo1, Funda Meric-Bernstam2,3, Sant Chawla4, Gerald Falchook3, David Hong3,
`Argun Akcakanat2, Huiqin Chen1, Aung Naing3, Siqing Fu3, Jennifer Wheler3, Stacy Moulder1,
`Thorunn Helgason3, Shaoyi Li6, Ileana Elias6, Neil Desai6, and Razelle Kurzrock5
`
`Abstract
`
`Purpose: This dose-finding phase I study investigated the maximum-tolerated dose (MTD) and safety of
`weekly nanoparticle albumin-bound rapamycin (nab-rapamycin) in patients with untreatable advanced
`nonhematologic malignancies.
`Experimental Design: nab-Rapamycin was administered weekly for 3 weeks followed by 1 week of rest,
`with a starting dose of 45 mg/m2. Additional doses were 56.25, 100, 150, and 125 mg/m2.
`Results: Of 27 enrolled patients, 26 were treated. Two dose-limiting toxicities (DLT) occurred at 150
`mg/m2 [grade 3 aspartate aminotransferase (AST) elevation and grade 4 thrombocytopenia], and two DLTs
`occurred at 125 mg/m2 (grade 3 suicidal ideation and grade 3 hypophosphatemia). Thus, the MTD was
`declared at 100 mg/m2. Most treatment-related adverse events (TRAE) were grade 1/2, including
`thrombocytopenia (58%), hypokalemia (23%), mucositis (38%), fatigue (27%), rash (23%), diarrhea
`(23%), nausea (19%), anemia (19%), hypophosphatemia (19%), neutropenia (15%), and hypertrigly-
`ceridemia (15%). Only one grade 3 nonhematologic TRAE (dyspnea) and one grade 3 hematologic event
`(anemia) occurred at the MTD. One patient with kidney cancer had a partial response and 2 patients
`remained on study for 365 days (patient with mesothelioma) and 238 days (patient with neuroendocrine
`tumor). The peak concentration (Cmax) and area under the concentration–time curve (AUC) of rapamycin
`increased with dose between 45 and 150 mg/m2, except for a relatively low AUC at 125 mg/m2. nab-
`Rapamycin significantly inhibited mTOR targets S6K and 4EBP1.
`Conclusions: The clinical dose of single-agent nab-rapamycin was established at 100 mg/m2 weekly (3 of
`4 weeks) given intravenously, which was well tolerated with preliminary evidence of response and stable
`disease, and produced a fairly dose-proportional pharmacokinetic profile in patients with unresectable
`advanced nonhematologic malignancies. Clin Cancer Res; 19(19); 5474–84. Ó2013 AACR.
`
`Introduction
`The prognosis for patients with advanced solid tumors is
`poor, as most malignancies are not responsive to standard
`treatments at the advanced stage. mTOR, a serine/threonine-
`specific protein kinase, is downstream of the phosphoinosi-
`tide 3-kinase (PI3K)/Akt pathway, and a key regulator of cell
`
`Authors' Affiliations: Departments of 1Breast Medical Oncology and Sys-
`tems Biology, 2Surgical Oncology, and 3Investigational Cancer Therapeu-
`tics, The University of Texas MD Anderson Cancer Center, Houston, Texas;
`4Sarcoma Oncology Center, Santa Monica; 5Division of Hematology-Oncol-
`ogy, University of California, San Diego, California; and 6Celgene, Summit,
`New Jersey
`
`Note: Supplementary data for this article are available at Clinical Cancer
`Research Online (http://clincancerres.aacrjournals.org/).
`
`Corresponding Author: Ana M. Gonzalez-Angulo, Department of Breast
`Medical Oncology and Department of Systems Biology, The University of
`Texas MD Anderson Cancer Center, 1515 Holcombe Blvd., Unit 1354,
`Houston, TX 77030. Phone: 713-792-2817; Fax: 713-794-4385; E-mail:
`agonzalez@mdanderson.org
`
`doi: 10.1158/1078-0432.CCR-12-3110
`Ó2013 American Association for Cancer Research.
`
`survival, proliferation, stress, and metabolism (1). mTOR
`inhibition with rapamycin and rapalogs (everolimus and
`temsirolimus) has proven to be effective in various solid
`tumors including renal cell carcinoma, neuroendocrine
`tumors, and breast cancer (2–12).
`Although rapamycin is an efficacious allosteric inhibitor of
`mTOR complex 1 (mTORC1), it has low oral bioavailability,
`poor solubility, and dose-limiting intestinal toxicity (13, 14).
`Other rapalogs, including everolimus and ridaforolimus, are
`also oral preparations and are often associated with signif-
`icant stomatitis (15). Temsirolimus, a prodrug of rapamycin,
`requires conversion by the CYP3A enzyme and also carries a
`significant risk for developing skin rash and stomatitis (16).
`Because none of the rapalogs are highly water soluble, they
`require surfactants and solvents in an intravenous formula-
`tion, such as polysorbate 80 for temsirolimus (17). The use of
`surfactants can potentially cause irritation, local inflamma-
`tion, and potential reduction of drug efficacy due to micellar
`sequestration, and the need for premedication to avoid
`potential hypersensitivity reactions (17). The nanoparticle
`albumin-bound rapamycin (nab-rapamycin; Celgene Inc.),
`
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`.At(~ AmericanAssoci,atwn for Cancer Research
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`Translational Relevance
`In this first clinical evaluation of nanoparticle albu-
`min-bound rapamycin (nab-rapamycin), an mTOR
`inhibitor, it was well tolerated given intravenously in
`patients with unresectable advanced solid tumors. Most
`rapalogs are oral preparations requiring toxic surfactants
`for intravenous formulation due to poor water solubil-
`ity. The nab-technology exploits the natural properties of
`human albumin to achieve a solvent-free drug delivery.
`Dose-limiting toxicities (DLT) including mucositis/sto-
`matitis that were observed with mTOR inhibitors were
`not dose-limiting with nab-rapamycin. Notably, 27% of
`patients were 65 years or older, a frail population that are
`more prone to toxicities and receive lesser benefits than
`younger patients from everolimus/temsirolimus. Pre-
`liminary proof-of-efficacy was observed in this phase I
`study. nab-Rapamycin produced a fairly dose-propor-
`tional peak concentration (Cmax) and area under the
`concentration–time curve (AUC) increase of rapamycin,
`and significantly inhibited mTOR targets.
`
`with a mean particle size of about 100 nm, is freely dis-
`persible in saline and is suitable for intravenous adminis-
`tration, and may be an advantageous alternative to oral
`rapamycin or oral rapalogs. Human albumin has broad
`binding affinity and accumulates in tumors, making it an
`ideal candidate for drug delivery (18, 19). In preclinical
`studies, nab-rapamycin was safe and highly effective in
`multiple tumor types; it reduced cell viability and decreased
`downstream signaling in various xenograft cancer models,
`including pancreatic, colorectal, multiple myeloma, and
`breast cancer (20–23). In addition, in human breast xeno-
`graft models, nab-rapamycin alone produced 75% tumor
`growth inhibition without weight loss and antitumor activ-
`ity was further enhanced with the combination of doxoru-
`bicin (a topoisomerase inhibitor), SAHA [an histone dea-
`cetylase (HDAC) inhibitor], erlotinib (an EGF tyrosine
`kinase inhibitor), and perifosine (an Akt inhibitor) with
`15% or less weight loss, indicating high tolerability in the
`combination regimens (24).
`On the basis of the promising preclinical results, this dose-
`finding phase I study investigated the maximum-tolerated
`dose (MTD) and safety of intravenous single-agent weekly
`nab-rapamycin in patients with untreatable advanced non-
`hematologic malignancies.
`
`Patients and Methods
`This study was conducted at MD Anderson Cancer Center
`(Houston, TX), and the Sarcoma Oncology Center (Santa
`Monica, CA). The study was approved by the Institutional
`Review Board of both participating medical institutions and
`was conducted in compliance with the World Medical
`Association Declaration of Helsinki and Good Clinical
`Practice, Guidelines of the International Conference on
`
`nab-Rapamycin in Advanced Nonhematologic Malignancies
`
`Harmonization (25). Written informed consent was obt-
`ained from all patients before study initiation.
`
`Patients
`Eligible patients were 18 years or older, had histologically
`or cytologically confirmed diagnosis of stage IV cancer that
`was not amenable to curative therapy. Advanced disease was
`defined as metastatic disease or locally advanced disease
`that was surgically unresectable and considered unmanage-
`able with standard therapies such as radiation or systemic
`therapies. Patients had a measurable disease by Response
`Evaluation Criteria in Solid Tumor (RECIST) v1.0, life
`expectancy 3 or more months, an Eastern Cooperative
`Oncology Group (ECOG) performance status of 0–1, ade-
`quate renal function (serum creatinine <1.5 mg/dL and/or
`creatinine clearance 60 mL/min), and were off all therapy
`for at least 4 weeks before study drug administration.
`Patients were excluded from the study if they had brain
`metastasis, history of interstitial lung disease and/or pneu-
`monitis, or a history of allergy or hypersensitivity to the
`study drug or any compounds of similar chemical or bio-
`logic composition.
`
`Study design
`This dose-finding study evaluated MTD and dose-limit-
`ing toxicities (DLT) of nab-rapamycin in patients with
`advanced nonhematologic malignancies. Following base-
`line evaluations, patients entered into the treatment period.
`nab-Rapamycin was administered by intravenous infusion
`for 30 minutes weekly for 3 weeks followed by 1 week of rest
`(28-day cycle), with a starting dose of 45 mg/m2. The
`starting dose of nab-rapamycin was chosen on the basis of
`nonclinical toxicology data of nab-rapamycin. Additional
`dose levels were 56.25, 100, 150, and 125 mg/m2. The
`original protocol was amended to add the 125 mg/m2 dose
`cohort for refinement of MTD.
`The first cycle was considered the treatment interval for
`determination of DLTs and the MTD. The MTD for nab-
`rapamycin was determined using a standard 3þ3 design,
`where 3 patients were enrolled at each dose level. The
`protocol was amended to ensure that all patients at a given
`dose level complete one cycle of therapy before patients
`were enrolled at the next dose level. If no DLT was observed,
`3 additional patients were enrolled at the next dose level. If
`one DLT was observed, the dose level was expanded to 6
`patients. If two DLTs were observed at a given dose level, the
`MTD was considered to be exceeded. Of the 6-patient
`expanded cohort, if 1 of 6 patients experienced a DLT,
`this was defined as the MTD. All patients at a given dose level
`completed one cycle of
`therapy before patients were
`enrolled at the next dose level.
`A DLT was defined [using the National Cancer Institute
`Common Terminology Criteria of Adverse Events (NCI
`CTCAE) v3.0] as any grade 3/4 nonhematologic toxicity,
`grade 3/4 nausea, or vomiting that occurred despite tre-
`atment, grade 4 thrombocytopenia of any duration and
`grade 4 uncomplicated neutropenia (i.e., without fever
`or infection) lasting more than 7 days, grade 4 febrile
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`Gonzalez-Angulo et al.
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`neutropenia that required hospitalization, and any grade 3
`hematologic toxicity that required treatment delay beyond
`3 weeks.
`Throughout the study, patients were routinely assessed
`for toxicities, response, and possible need for a dose mod-
`ification. Patients continued on treatment until they expe-
`rienced progressive disease or unacceptable toxicity, with-
`drew consent, or their physician felt it was no longer in their
`best
`interest
`to continue on treatment. Discontinued
`patients completed the end of study evaluation and entered
`into a 30-day follow-up period.
`
`Assessments and statistical methods
`All patients who received at least one dose of study drug
`(treated population) were evaluated for safety. Safety and
`tolerability endpoints included the incidence of treatment-
`related adverse events (TRAE) by NCI CTCAE v3.0 and the
`percentage of patients experiencing TRAEs that required
`dose delays/modifications, and/or premature discontinua-
`tion of the study drug.
`The exploratory efficacy analysis included the summary
`of percentage of patients who achieved an objective with
`confirmed complete or partial tumor response (CR or PR,
`respectively) and the percentage of patients with confirmed
`stable disease for at least 12 weeks, using RECIST v1.0. The
`objective tumor responses of target or nontarget lesions
`were classified individually based on RECIST v1.0. The
`overall tumor response was determined by taking into
`account the responses of target lesions and nontarget
`lesions as well as the presence of new lesions.
`Tumor response assessments were carried out every 12
`weeks. A waterfall plot was used to illustrate the percentage
`change of target lesion from baseline for all patients with
`target tumor evaluation. The corresponding objective target
`lesion responses, dose level cohorts, and tumor types were
`also provided in the graph.
`
`Molecular analyses
`Evaluation of PTEN loss was carried out with immu-
`nohistochemistry (IHC) using monoclonal mouse anti-
`human PTEN antibody clone 6H2.1 from Dako at 1:100
`dilution, as described by Gonzalez-Angulo and colleagues
`(26). Briefly, both cytoplasmic and nuclear PTEN staining
`in the tumor and non-neoplastic ductal epithelium and
`stroma were quantified. PTEN expression level was scored
`semiquantitatively on the basis of staining intensity (SI)
`and distribution using the immunoreactive score (IRS) as
`follows: IRS ¼ SI  percentage of positive cells. Staining
`intensity was determined as 0, negative; 1, weak; 2,
`moderate; and 3, strong. Percentage of positive cells was
`defined as 0, <1%; 1, 1%–10%; 2, 11%–50%; 3, 51%–
`80%; and 4, >80% positive cells. Tumors with IRS of 0
`were considered to have PTEN loss. A mass spectroscopy–
`based approach evaluating single-nucleotide polymorph-
`isms (SNP) was used to detect known mutations in
`members of the PI3K pathway. Molecular analysis was
`conducted in patients who showed clinical benefit using
`archival tissue.
`
`Pharmacokinetics
`Whole-blood samples (4 mL each) were collected in
`vacutainer tubes containing EDTA as the anticoagulant for
`determination of rapamycin. Samples were obtained only
`during cycle 1 and were taken immediately predose (before
`infusion), during the infusion (15 and 30 minutes before
`end of the infusion), and postinfusion at 1.0, 1.5, 2, 4, 6, 8,
`24, 48, 72, 96, and 168 hours. The samples were stored
`frozen at a temperature between 20
`C and 80
`
`
`C until
`shipment for analysis to St. George’s Hospital at the Uni-
`versity of London (London, United Kingdom).
`The whole-blood samples were analyzed for total (free þ
`bound) rapamycin using high-performance liquid chroma-
`tography–tandem mass
`spectrometry (HPLC/MS-MS).
`Rapamycin concentrations in whole blood were validated
`from 10 to 2,000 ng/mL with 32-desmethoxyrapamycin
`used as an internal standard. Analytes were extracted using
`a solvent mixture and detected and quantified by reverse
`phase HPLC with detection via turbo ion-spray mass
`spectrometry.
`The concentration-versus-time data for rapamycin in
`whole blood were analyzed using a noncompartmental
`analysis technique and WinNonlin software. Pharmacoki-
`netic analysis was based on whole-blood concentrations
`due to the known instability of rapamycin in plasma.
`Calculated parameters included peak concentration (Cmax),
`half-life (t1/2), area under the concentration–time curve
`(AUC), clearance (CL), and steady-state volume of distri-
`bution (Vss). A simple regression model was applied to
`assess the relationship of the pharmacokinetic parameters
`with dose.
`
`Peripheral blood mononuclear cells and reverse phase
`protein arrays
`Whole blood for pharmacodynamics evaluation was
`collected only during cycle 1 at four time points: C1 D1
`(pretreatment), C1 D2, C1 D4, and C1 D8 (immediately
`before next dose) in an 8-mL cell preparation tube with
`sodium citrate (Becton, Dickinson and Company). Sepa-
`ration of peripheral blood mononuclear cells (PBMC) from
`whole blood was accomplished through density gradient
`centrifugation using Ficoll following the manufacturer’s
`recommendations. After centrifugation, plasma compo-
`nent from the upper half of the tube was transferred to
`cryotubes and snap-frozen. The layer containing the cells
`was transferred to a fresh tube, washed, and centrifuged.
`After removal of the supernatant, PBMC pellet was also
`snap-frozen.
`Reverse phase protein array (RPPA) was conducted in the
`MD Anderson Cancer Center Functional Proteomics RPPA
`Core Facility as described previously (27). PBMC samples
`were resuspended in RPPA lysis buffer containing 0.25%
`sodium deoxycholate. Protein concentrations were deter-
`mined using BCA method (Pierce) and 4 SDS sample
`buffer was added. Final protein concentration was adjusted
`to 3 mg/mL. Samples were probed with antibodies that were
`validated for RPPA. A total of 135 proteins and 21 replicates
`were analyzed,
`including S6 S240/244, S6 S235/236,
`
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`nab-Rapamycin in Advanced Nonhematologic Malignancies
`
`Table 1. Baseline patient demographics and
`characteristics
`
`MTD
`n ¼ 7
`57 (36, 76)
`4 (57)
`3 (43)
`
`5 (71)
`2 (29)
`
`0
`0
`6 (86)
`1 (14)
`
`2 (29)
`5 (71)
`0
`
`7 (100)
`
`Age, median years (range)
`<65 years, n (%)
`65 years, n (%)
`Gender, n (%)
`Male
`Female
`Race
`Asian, n (%)
`African heritage, n (%)
`Caucasian, n (%)
`Hispanic, Latino, n (%)
`ECOG, n (%)
`0
`1
`2
`Stage at current diagnosis, n (%)
`IV
`Site of primary diagnosis, n (%)
`1 (14)
`Bladder
`0
`Breast
`2 (29)
`Colorectal
`1 (14)
`Esophagus
`1 (14)
`Head and neck
`0
`Kidney
`1 (14)
`Lung/thoracic
`0
`Prostate
`0
`Stomach
`1 (14)
`Uterus
`0
`Other
`Histology of primary diagnosis, n (%)
`Carcinoma/adenocarcinoma
`5 (71)
`Sarcoma/sarcomatoid
`2 (29)
`Site of metastasis, n (%)
`Visceral
`Nonvisceral
`
`S6KT389, 4EBP1 T37/46, and 4EBP1 T70. Proteomics
`assessment of S6 S240/244 and 4EBP1 T37/46 was carried
`out using Meso Scale discovery (MSD) phosphoprotein
`assays (Meso Scale Discovery).
`The RPPA spot signal intensity data obtained from Micro-
`Vigene automated RPPA module (VigeneTech, Inc.) were
`analyzed using the R package SuperCurve (version 1.4.3;
`ref. 28), available at "http://bioinformatics.mdanderson.
`org/OOMPA". RPPA raw data were treated with median
`centering across samples, and then a centering by the
`sample median was undertaken on the treated data and
`the final normalized data were obtained by applying medi-
`an absolute deviation (MAD) scaling to the data. Linear
`mixed models and ANOVA tests were developed and
`applied to test the pre- versus posttreatment and inhibition
`effects at each dose level and each pair of time points. Tukey
`tests were also used for pairwise comparisons. To test the
`association of proteins expression on patients’ response,
`patients with stable disease and progressive disease were
`also compared using logistic models adjusted by time
`points, and their interactions were also taken into account.
`
`Results
`Patients
`Twenty-seven patients were enrolled in the study and 26
`patients were treated of which 19 have evaluable tumor
`assessment data. Specifically, 7 patients were treated in the
`45 mg/m2 arm, 1 additional patient was added after a pat-
`ient did not complete a full cycle, 3 in the 56.25 mg/m2, 7
`in the 100 mg/m2, 2 in the 150 mg/m2, and 7 in 125 mg/m2
`arm. Seven patients had no tumor assessments beyond
`the baseline evaluation as a result of loss to follow-up
`(3 patients), patient request (1 patient), drug shortage
`(1 patient), and incomplete tumor evaluation (1 patient).
`All patients had discontinued therapy at the time of this
`analysis. Eighteen (69%) patients discontinued treatment
`because of disease progression, 4 (15%) due to adverse
`events/toxicities, 2 (8%) for patient request, and 2 (8%) for
`drug shortage. Patient baseline demographics and charac-
`teristics were described in Table 1. Briefly, the median age
`was 60.5 years, and with the majority of patients were male
`(62%), Caucasian (81%), and had a baseline ECOG score
`of 1 (73%). The most common sites of primary tumor
`diagnosis were head and neck, colorectal, and kidney
`(12% each). Most patients had a carcinoma/adenocarcino-
`ma (54%) and the rest had sarcoma. All patients had visceral
`metastases. The most common sites of metastases were
`lung/thoracic (69%), liver (46%), lymph node (42%), and
`abdomen/peritoneal (42%).
`
`Treatment exposure
`For all patients, the median number of cycles adminis-
`tered was three (1–11, 15, 29), with 27% of patients having
`more than three cycles of therapy. The median cumulative
`rapamycin dose was 405 mg/m2 (100–2,200), with the
`median dose intensity of 68.9 mg/m2/wk (11.4–150.0).
`At the MTD, the median number of cycles was also three
`
`All treated
`patients
`n ¼ 26
`60.5 (18, 78)
`19 (73)
`7 (27)
`
`16 (62)
`10 (38)
`
`1 (4)
`2 (8)
`21 (81)
`2 (8)
`
`5 (19)
`19 (73)
`2 (8)
`
`26 (100)
`
`1 (4)
`1 (4)
`3 (12)
`2 (8)
`3 (12)
`3 (12)
`2 (8)
`1 (4)
`1 (4)
`1 (4)
`8 (31)
`
`14 (54)
`12 (46)
`
`26 (100)
`0
`
`7 (100)
`0
`
`(1–3, 15, 29), with the median cumulative dose of 800
`mg/m2 (100–900) and median dose intensity of 78.9
`mg/m2/wk (51.1–100.0).
`
`Safety results
`MTD. Following dose escalation to 100 mg/m2, nab-
`rapamycin dose was initially escalated to 150 mg/m2. Two
`DLTs occurred in the 150 mg/m2 cohort: a grade 3 elevation
`of aspartate aminotransferase (AST) and a grade 4 throm-
`bocytopenia. After observing DLTs at the 150 mg/m2 cohort,
`a new dose level of 125 mg/m2 was added for refinement of
`MTD. At the 125 mg/m2 dose level, two DLTs occurred
`(grade 3 suicidal ideation and grade 3 hypophosphatemia);
`therefore, the MTD was reached and declared at 100 mg/m2.
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`Gonzalez-Angulo et al.
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`TRAEs. For all cohorts and all grades, 25 of 26 (96%)
`patients experienced at least one TRAE. The most common
`nonhematologic TRAEs reported were mucosal inflamma-
`tion (10 patients; 38%), fatigue (7 patients; 27%), rash (6
`patients; 23%), diarrhea (6 patients; 23%), and nausea (5
`patients; 19%; see Table 2). Most of these adverse events
`were grade 1/2 events, with only three grade 3 nonhema-
`tologic adverse events (two elevated AST and one dyspnea).
`Specifically, at the MTD (100 mg/m2), all 7 patients expe-
`rienced at least one TRAE of any grades, and the most
`common adverse events were mucositis and fatigue (5
`patients; 71% each). Four (15%) patients experienced at
`least one treatment-related serious adverse event, including
`arrhythmia (grade 2) and mood alteration (grade 3) both in
`the 125 mg/m2 cohort, vomiting (grade 3) in the 45 mg/m2
`cohort, and dyspnea (grade 3) in the 100 mg/m2 cohort.
`The most common hematologic TRAE, for all cohorts and
`grades, were thrombocytopenia (58%), followed by hypo-
`kalemia (23%), anemia and hypophosphatemia (19%
`each), and neutropenia and hypertriglyceridemia (15%
`each; see Table 2). Most of these events were grade 1/2,
`and only one grade 4 hematologic event occurred (throm-
`bocytopenia in the 150 mg/m2 arm). At the MTD, the only
`hematologic adverse event was a grade 3 anemia.
`Treatment-related study drug reductions, delays, and dis-
`Five (19%) patients experienced TRAEs
`continuations.
`that required study drug dose reductions and 50% of dose
`reductions occurred at cycle 2. Only 1 patient at the MTD
`had an adverse event that required a dose reduction, which
`
`occurred at cycle 4. The specific events requiring dose
`reductions were one grade 2 thrombocytopenia and one
`grade 2 dyslipidemia in the 100 mg/m2 cohort, and two
`grade 3 thrombocytopenia and one grade 3 suicidal idea-
`tion in the 125 mg/m2 cohort. The patient who experienced
`suicidal ideation had been on antidepressants before the
`trial. After the onset of grade 3 suicidal ideation (end of cycle
`1), this patient received two cycles of nab-rapamycin at a
`reduced dose (100 mg/m2), during which no suicidal
`ideation was reported. In addition, there was a dose reduc-
`tion for a grade 2 elevated AST in the 45 mg/m2 cohort. The
`dose was reduced to 30 mg/m2, which was not specified in
`the protocol. This patient responded to treatment and the
`physician felt that continuing the treatment at a lower dose
`was in the best interest for this patient.
`Sixteen (62%) patients had TRAEs requiring a dose delay:
`4 (57%) patients in the 45 mg/m2, 1 (33%) in the 56.25
`mg/m2, 4 (57%) in the 100 mg/m2, 2 (100%) in the 150
`mg/m2, and 5 (71%) in the 125 mg/m2 cohort. Specifically
`in the 100 mg/m2 cohort, the treatment-related dose delays
`were due to three grade 2 thrombocytopenia, a grade 2
`elevated triglycerides, a grade 2 mucosal inflammation, and
`a grade 3 dyspnea. Only 1 patient had a TRAE that resulted in
`study drug discontinuation (150 mg/m2 cohort; 1 patient
`with a grade 4 thrombocytopenia and a grade 2 diarrhea).
`
`Pharmacokinetics
`Whole-blood samples obtained during cycle 1 of treat-
`ment at
`the specified time points were analyzed for
`
`Table 2. Treatment-related grade 1–4 hematologic and nonhematologic adverse events reported in 10% or
`more of all treated patients
`
`NCI CTCAE v 3.0
`Hematologic AEs, n (%)
`Anemia
`Hypokalemia
`Hypophosphatemia
`Hypertriglyceridemia
`Neutropenia
`Thrombocytopenia
`Nonhematologic AEs, n (%)
`AST
`Constipation
`Diarrhea
`Dyspnea
`Fatigue
`Infection, oral cavity
`Mucositis/stomatitis
`Nausea
`Rash
`Weight loss
`
`MTD (100 mg/m2)
`n ¼ 7
`G3
`
`G1
`
`G2
`
`G4
`
`G1
`
`G2
`
`All treated patients
`n ¼ 26
`G3
`
`0
`1 (14)
`0
`1 (14)
`1 (14)
`1 (14)
`
`0
`0
`1 (14)
`0
`1 (14)
`1 (14)
`3 (43)
`1 (14)
`1 (14)
`0
`
`0
`0
`1 (14)
`1 (14)
`0
`4 (57)
`
`0
`1 (14)
`0
`1 (14)
`4 (57)
`1 (14)
`2 (29)
`1 (14)
`0
`0
`
`1 (14)
`0
`0
`0
`0
`0
`
`0
`0
`0
`1 (14)
`0
`0
`0
`0
`0
`0
`
`0
`0
`0
`0
`0
`0
`
`0
`0
`0
`0
`0
`0
`0
`0
`0
`0
`
`0
`5 (19)
`1 (4)
`2 (8)
`2 (8)
`5 (19)
`
`1 (4)
`1 (4)
`3 (12)
`1 (4)
`1 (4)
`3 (12)
`7 (27)
`3 (12)
`4 (15)
`1 (4)
`
`3 (12)
`0
`2 (8)
`1 (4)
`1 (4)
`6 (23)
`
`0
`2 (8)
`3 (12)
`2 (8)
`6 (23)
`2 (8)
`3 (12)
`2 (8)
`2 (8)
`2 (8)
`
`2 (8)
`1 (4)
`2 (8)
`1 (4)
`1 (4)
`3 (12)
`
`2 (8)
`0
`0
`1 (4)
`0
`0
`0
`0
`0
`0
`
`G4
`
`0
`0
`0
`0
`0
`1 (4)
`
`0
`0
`0
`0
`0
`0
`0
`0
`0
`0
`
`Abbreviations: AE, adverse event; G, grade.
`
`5478
`
`Clin Cancer Res; 19(19) October 1, 2013
`
`Clinical Cancer Research
`
`

`

`nab-Rapamycin in Advanced Nonhematologic Malignancies
`
`211.11(41.62)
`296.74(20.79)
`215.09(29.03)
`135.70(25.71)
`107.36(11.35)
`(%CV)
`Vss,L/m2
`
`36.03(2.25)
`78.80(20.05)
`44.34(44.72)
`39.35(34.88)
`31.29(15.34)
`(%CV)
`CL,mL/min/m2
`
`462.67(2.25)
`219.39(21.34)
`440.73(40.31)
`455.12(30.01)
`545.87(18.74)
`(%CV)
`ng h m2/mL/mg
`AUCinf/dose,
`
`69,400.48(2.25)
`27,423.48(21.34)
`44,072.64(40.31)
`25,600.46(30.01)
`24,564.19(18.74)
`(%CV)
`AUCinf,ng h/mL
`
`25.52(96.39)
`34.09(41.01)
`32.28(38.00)
`51.23(36.38)
`39.47(40.30)
`(%CV)
`ng m2/mL/mg
`Cmax/dose,
`
`3,828.45(96.39)
`4,261.24(41.01)
`3,227.61(38.00)
`2,881.77(36.38)
`1,776.27(40.30)
`(%CV)
`Cmax,ng/mL
`
`1.10(72.86)
`0.36(37.42)
`0.46(20.35)
`0.44(21.66)
`0.46(20.27)
`tmax,h(%CV)
`
`90.75(5.42)
`54.93(22.74)
`63.10(35.77)
`46.31(33.36)
`39.57(26.79)
`HL,h(%CV)
`
`150(2)
`125(7)
`100(7)
`56.25(3)
`45(7)
`
`mg(n)
`Dose,
`
`Abbreviations:HL,half-life;tmax,timeatpeakplasmaconcentration.
`NOTE:meanvaluesarepresentedforallvariables.
`
`Table3.Summarystatisticsofpharmacokineticvariableestimates
`
`rapamycin concentration and noncompartmental phar-
`macokinetic analyses were conducted. Of 27 enrolled
`patients, 26 were evaluable for pharmacokinetic analyses
`(patient demographics in Table 1). There was a rapid
`decline in whole-blood levels of rapamycin in the first
`2 hours following the 30-minute infusion of nab-rapa-
`mycin, which was followed by a slower elimination phase
`(Table 3 and Fig. 1). The Cmax increased proportionally
`over the dose range of 45 to 150 mg/m2 as did the AUC,
`except for a relatively low AUC in the 125 mg/m2 dose
`cohort (Table 3).
`
`Efficacy results
`Of 19 patients evaluable for efficacy with best overall
`tumor response assessments, which included assessment
`of target, nontarget, and new lesions across all cycles, 1
`patient (5%) in the 45 mg/m2 cohort diagnosed with
`adenocarcinoma of the kidney and with bone and intra-
`thoracic metastases had a confirmed PR. The target lesion
`of this patient was reduced by 35.1% and the duration of
`response lasted 183 days. Two (11%) patients had an
`overall tumor evaluation of stable disease (confirmed): 1
`patient with mesothelioma had stable disease for 365
`days and 1 patient with a neuroendocrine tumor in the
`left axillary node had stable disease for 238 days. Eight
`patients had stable disease that could not be confirmed
`either due to absence of follow-up tumor evaluation after
`the first stable disease, or due to progression after the first
`stable disease.
`The waterfall plot in Fig. 2 illustrates the percentage
`change in the target tumors in 18 evaluable patients with
`various tumor types and histologies. Two (11%) patients
`with adenocarcinoma of the kidney had more than 30%
`decrease in the target lesion, which included the patient
`dosed at 45 mg/m2 (the patient mentioned above who had
`a confirmed PR) and another patient in the 56.25 mg/m2
`cohort, whose target lesion was reduced by 34.7% and a
`duration of response lasting 104 days. As seen in Fig. 2, 13
`(72%) patients had a target lesion objective response
`evaluation of stable disease. These patients had cancer of
`the bladder, colorectal, esophagus, head and neck, pros-
`tate, retroperitoneal, or uterus. It is notable that many of
`these patients with a target tumor evaluation of stable
`disease did not have a confirmed overall tumor evaluation
`of stable disease, which in addition to target lesions also
`accounted for nontarget and new lesions. In addition, 3
`(17%) patients had target lesion objective response of pro-
`gressive disease. As seen in Fig. 2, patients with any decrease
`in the target tumor lesion had carcinoma/adenocarcinoma
`of the kidney, bladder, esophagus, or neuroendocrine can-
`cer. Of note, the molecular analyses of the tumor biopsy
`obtained from the patient with mesothelioma achieving the
`longest clinical benefit revealed no loss in PTEN, or an
`activating mutation in PIK3CA or AKT. However, a SNP was
`observed on PHLPP2 (PH domain leucine-rich repeat pro-
`tein phosphatase 2), a gene that codes for a protein phos-
`phatase that mediates dephosphorylation of serine 473 in
`Akt1 (30).
`
`www.aacrjournals.org
`
`Clin Cancer Res; 19(19) October 1, 2013
`
`5479
`
`

`

`nab-Rapamycin treatment was associated with a signifi-
`cant decrease of S6K T389 on D2 with persistent inhibition
`at D8 (Fig. 3A and B) at all doses. nab-Rapamycin treatment
`was associated with significant decrease of 4EBP1 T70 levels
`on D2 and D4, but with recovery by D8 (Fig. 3A and C). nab-
`Rapamycin at 56.25 dose level was not associated with a
`decrease in 4EBP1 T70 levels, whereas a significant decrease
`in 4EBP1 T70 levels was seen with higher doses (Fig. 3B).
`These results show that nab-rapamycin has a dose-depen-
`dent effect on mTOR signaling, with pathway inhibition
`being seen at 56.25 mg/m2 and higher doses. The duration
`of inhibition differs between downstream targets, and is
`longer for S6K T389 than for 4EBP1 T70.
`Next, we determined whether the pharmacokinetic data
`correlated with pathway inhibition on RPPA. There was a
`moderate negative correlation between PBMC 4EBP1 T70
`levels and serum rapamycin concentrations (r2 ¼ 0.446)
`as well as between S6K T389 and serum rapamycin con-
`centration (r2 ¼ 0.517). Unfortunately, PBMCs were not
`available for the patient who had a PR. There was no
`significant difference in the inhibition 4EBP1 and S6K
`phosphorylation between patients who had stable disease
`and patients who had progressive disease.
`
`Discussion
`The results of this phase I dose-finding study showed that
`the MTD for nab-rapamycin in patients with advanced
`nonhematologic malignancies was 100 mg/m2, which pro-
`duced favorable safety profile without the DLTs typically
`
`45
`56.25
`100
`125
`150
`
`Gonzalez-Angulo et al.
`
`4,000
`
`3,000
`
`2,000
`
`1,000
`
`(ng/mL)
`
`Concentration
`
`0
`0.25 0.5 1
`
`2
`
`16 32 64 128 256
`8
`4
`Time (h)
`
`Figure 1. nab-Rapamycin plasma concentration by time.
`
`Pharmacodynamics
`PBMCs were collected from 18 patients. The effect of nab-
`rapamycin on mTOR signaling was assessed by evaluating
`the phosphorylation of mTOR targets, including 4EBP and
`S6K, and S6K target ribosomal S6 on phosphorylation sites,
`with two different assays (MSD and RPPA) previously used
`to show rapamycin-mediated inhibition of downstream
`signaling (31). The MSD phosphoprotein assays revealed
`very low baseline expression of S6 S240/244 and 4EBP T37/
`46 in pretreatment PBMC samples, thus we were unable to
`assess regulation by nab-rapamycin treatment. Therefore,
`the effect of nab-rapamycin on the functional proteomic
`profile was assessed by RPPA.
`
`Progressive disease, i.e., ≥20% increase
`
`-
`
`Stable disease
`
`-
`
`PR, i.e., ≥30 % decrease
`
`-
`
`30
`
`20
`
`10
`
`0
`
`–10
`
`–20
`
`–30
`
`Figure 2. Waterfall plot showing
`percentage change of target lesion
`from baseline by patient and tumor
`types.
`
`Percentage change of target tumor from baseline
`
`mg/m2
`
`100
`
`125
`
`45
`
`56.25
`
`45
`
`100
`
`45
`
`45
`
`100
`
`125
`
`150
`
`125
`
`100
`
`45
`
`100
`
`56.25
`
`45
`
`Kidney
`
`Kidney
`
`Bladder
`
`Esophagus
`
`Esophagus
`
`Neuroendocrine
`
`Retroperitoneal
`
`x
`
`Colorectal
`
`Colorectal
`
`Retroperitoneal
`
`Pleural mesothelioma
`
`Uterus
`
`Prostate
`
`Head and neck
`
`Head a