Phase I Trial of PT-100 (PT-100), A Cytokine-Inducing
Small Molecule, Following Chemotherapy
for Solid Tumor Malignancy
John Nemunaitis, M.D.,1,2 Svetislava J. Vukelja, M.D.,1,3 Donald Richards, M.D.,1,3 Casey Cunningham, M.D.,1,2 Neil Senzer, M.D.,1,2 John Nugent, M.D.,1,4 Houston Duncan,2 Barry Jones, Ph.D.,5 Eric Haltom,5
and Margaret J. Uprichard, Pharm.D.5
Texas Oncology, Inc.,1 Mary Crowley Medical Research Center, Sammons Cancer Center, Baylor University Hospital, Dallas, Texas, USA,2 Tyler Cancer Center, Tyler, Texas, USA,3 Fort Worth Cancer Center, Fort Worth, Texas, USA,4 and Point Therapeutics, Inc., Boston,
Massachusetts, USA.5
ABSTRACT
PT-100 upregulates cytokine expression competitively inhibiting the dipeptidyl peptidase activity of fibroblast activation protein (FAP) and dipeptidyl peptidase IV (DPP-IV). This dose- escalation study was conducted to evaluate the safety of PT-100 in patients receiving myelo- suppressive chemotherapy and to assess its effects on neutrophil recovery.
PT-100 was administered orally for 7 days as a 200 µg, 400 µg, 800 µg, or 1,200 µg total daily dose (divided twice daily) to 6, 6, 17, and 5 patients, respectively. Patients received 2 cycles of chemotherapy: The first cycle served as each individual patient’s control. Patients had to develop Grade 3+ neutropenia in Cycle 1 in order to receive PT-100 in Cycle 2. Most patients received PT-100 on Days 2–8 of chemotherapy in Cycle 2, except at 800 µg where an additional cohort (n = 8) was treated on a Days 5–11 schedule. Five of 7 patients receiving 800 µg on Days 2–8 experienced a ≥1-day improvement in Grade 3+ neutropenia in Cycle 2 versus Cycle 1. Over- all, PT-100 was well tolerated. A reduction in chemotherapy-related nausea, vomiting, fatigue, alopecia, and diarrhea was noted in patients receiving PT-100. Edema/peripheral swelling, hy- potension, hypovolemia, and dizziness were the most common nonhematologic adverse events considered related to PT-100. Two Grade 3 adverse events were considered related to PT-100: syncope (1,200 µg) and orthostatic hypotension (800 µg). A maximum tolerated dose was not reached. Given the accelerated neutrophil recovery, preclinical evidence of antitumor activity, and tolerable toxicities of PT-100, additional studies to optimize the PT-100 dosing schedule in patients receiving myelosuppressive chemotherapy are needed.
INTRODUCTION
PT-100 (PT-100) is an orally available synthetic dipeptide (valine-proline boronic acid) (1), that specifically inhibits dipep-
The authors would like to acknowledge Michelle Hewitt and Brenda Marr for their competent and knowledgeable assistance in the preparation of this manuscript.
Keywords: PT-100, Small molecule, Chemotherapy, Malignancy. Correspondence to:
John Nemunaitis, M.D. 1717 Main Street
60th Floor
Dallas, Texas 75201 USA
email: [email protected]
tidyl peptidases such as fibroblast activation protein (FAP) (2, 3) and dipeptidyl peptidase IV (DPP-IV)/CD26 (4). Both enzymes cleave amino-terminal dipeptides from polypeptides with the sequence NH2 -Xaa-Pro (Ala), (5) where Xaa can be any nat- ural amino acid. Proteolytic activity of DPP-IV/CD26 appears to be involved in regulating activities of polypeptide hormones and chemokines (6). PT-100 appears to modulate the immune system through enhancing the production and release of a num- ber of cytokines and chemokines and has been shown to induce differentiation of hematopoietic progenitors via upregulation of stimulatory cytokines such as granulocyte colony stimulating factor (G-CSF) and interleukin (IL-6) (7). Data suggest that the hematopoietic activity of PT-100 is mediated through a dipep- tidyl peptidase other than CD26. One possible molecular target appears to be FAP, which has been characterized as a cell-surface
553
membrane protein expressed by cultured bone marrow stro- mal cells, lymphoid organs, reactive fibroblasts of tumor ex- tracellular matrix, ductal carcinoma cells of breast cancer, and melanoma cells (8–12). FAP mRNA also has been detected in lymphoid tissues (13). Substantially accelerated neutrophil re- covery was demonstrated in neutropenic mice following admin- istration of cyclophosphamide (10).
The ability of PT-100 to suppress tumor growth also has been described (7). Treatment of mice with PT-100 inoculated subcu- taneously with various tumors, including WEHI 164 (fibrosar- coma), EL4 (lymphoma), and B16 (melanoma), significantly inhibited tumor growth. Long-lived and specific immunity to secondary rechallenge was also established in PT-100-treated mice that rejected their tumors (7). Examination of mRNA levels indicated that PT-100 increased expression in the tumors and draining lymph nodes of cytokines and chemokines that have the ability to promote antitumor responses involving both innate and adaptive immunity (7). Furthermore, the addition of PT-100 to several chemotherapy agents: cisplatin, gemcitabine, and paclitaxel, was shown to enhance tumor rejection in mice compared to treatment with each agent alone. Addition of PT- 100 to the monoclonal antibodies, rituximab and trastuzumab also resulted in a greater antitumor effect in mouse xenograft models compared to the activity of each agent alone (7, 14).
PT-100 has been administered to 90 healthy male subjects in Phase 1 single- and multiple-dose tolerance study. It was well tolerated at single daily doses up to 1,200 µg administered for 7 days. Adverse events reported more frequently in sub- jects receiving PT-100 compared with placebo were headache, temperature-change sensation, myalgia, nausea, vomiting, pe- ripheral swelling, rigors, arthralgia, and sore throat. Hypoten- sion and syncope were not observed. Dose-related increases in plasma G-CSF and IL-6 were demonstrated.
Given the good tolerability at PT-100 doses ≤1,200 µg/
kg/day and the broad preclinical antitumor/hematopoietic activity profile, we initiated a Phase 1 dose-escalation trial to determine the maximum tolerated dose (MTD) of PT-100 and to characterize its hematopoietic activity in advanced cancer patients receiving myelosuppressive chemotherapy.
PATIENTS AND METHODS
Study design
ThiswasaPhase1,6-week,open-label,nonrandomized,mul- ticenter, dose-escalation study designed to determine the max- imum tolerated dose of PT-100 in patients with solid tumors receiving myelosuppressive chemotherapy (Figure 1). Patients were to receive two 3-week cycles of chemotherapy, with ad- ministration of PT-100 in the second cycle. Cycle 1 served as each patient’s own control to allow for a within-patient compar- ison of the addition of PT-100 to chemotherapy. Patients had to develop Grade 3 neutropenia in Cycle 1 to receive PT-100 in Cycle 2. The standard chemotherapy regimens allowed per pro- tocol were carboplatin/paclitaxel, doxorubicin/docetaxel, dox- orubicin/cyclophosphamide, or single-agent docetaxel.
Figure 1. Schematic of the phase 1 dose-escalating study in sub- jects receiving myelosuppressive chemotherapy. Subjects were studied in the initial 2 cycles of chemotherapy. In Cycle 1, sub- jects were not administered PT-100. In Cycle 2, subjects received PT-100 BID from either Days 2–8 or Days 5–11. In each cycle, daily blood samples were taken on Days 1–21 for determination of ANC. Plasma cytokine levels were determined by ELISA on Days 1, 2, 5, 6, 8, 11, 12, 14, 16 and 18. Sequentially ascending doses of PT-100 were studied in separate patient cohorts.
In the absence of dose-limiting toxicity, defined as Grade 3+ arthralgia, edema, elevations in ALT/AST, or unexpected hematologic study-drug related toxicity, 4 dose levels of PT-100 were planned: 200 µg, 400 µg, 800 µg, and 1,200 µg administered daily as 2 divided doses for 7 days on Days 2–8 of the second cycle of chemotherapy. At the 800 µg/day dose level, a dosing schedule of Days 5–11 also was studied. All doses were to be administered orally and on an empty stomach (i.e., 1 hour before or 2 hours after food). Six patients were planned for each dose cohort.
Patients consented to daily blood sampling for determination of ANC. Blood samples for evaluation of pharmacodynamic and pharmacokinetic evaluations also were collected at specified timepoints throughout the study. Adverse events were collected and coded using the Medical Dictionary for Regulatory Affairs (MedDRA).
PT-100 was supplied as either 100 µg or 200 µg tablets with instructions to take the appropriate number of tablets daily for the assigned dose level.
Patient eligibility
Patients 18 to 70 years old with solid tumors were eligible for this study if they were scheduled for chemotherapy with one of the 4 protocol-specified regimens. A Karnofsky performance status of at least 70, adequate organ and immune function, and an estimated life expectancy of at least 6 months were required. Patients were required to have recovered from prior surgery, and any spinal or pelvic radiation therapy must have been completed at least 4 weeks prior to Cycle 1, Day 1. Female patients of childbearing potential were required to have a negative serum pregnancy test result within 7 days of study enrollment and to use a barrier method of contraception throughout the study.
Patients were ineligible for the study if they had received more than 2 previous cytotoxic regimens; chemotherapy or any investigational therapy within 4 weeks of study entry; prior radi- ation therapy to greater than 25 percent of the bone marrow; his- tory of bone marrow transplant; myocardial infarction or stroke
554 J. Nemunaitis et al.
within 3 months of screening; active and uncontrolled infection; a psychiatric disorder or alcohol or chemical abuse that would interfere with consent or follow-up; pregnancy or lactation; or known hypersensitivity to the study drug or its analogs.
All participants signed an institutional review board- approved, protocol-specific, informed consent form prior to study participation.
Patient evaluation
Baseline evaluations included a complete medical history and physical examination (including height and weight), electrocar- diogram, Karnofsky performance status, vital signs (blood pres- sure,pulserate,respiratoryrate,andtemperature),CBCwithdif- ferential and platelet count, serum chemistries, urinalysis, serum pregnancy test, and documentation of concomitant medications. These evaluations also were performed at the beginning of Cy- cle 2. All clinical laboratory samples were analyzed by a central laboratory (LabCorp, Austin, TX).
During the 6-week study period, home healthcare nurses vis- ited the patients daily (except on clinic days) to collect blood for daily determination of circulating blood cells (CBCs), recording of vital signs, and to query for adverse events. In Cycle 2, vital signs also were measured on Days 2, 5, and 8 prior to PT-100 dosing, and at 2 and 8 hours after the morning dose of PT-100. Patients were to record their doses of PT-100 in a dosing diary provided to them. Blood serum chemistries and urinalysis were determined at the end of each week.
Pharmacodynamics
The protocol was amended mid-study to allow for blood sam- pling for cytokine and chemokine determination in the two PT- 100800 µg/dayandthe1,200 µgdaydosegroups.Sampleswere
Table 1. Baseline demographics and disease characteristics
analyzed by a central laboratory (R & D Systems, Minneapolis, MN) for the evaluation of G-CSF, IL-6, IL-8, and IL-11. Blood samples were collected at the following times: Cycle Days 1, 2, 5, 6, 8, 9, 11, 12, 14, 16, and 18. On Day 2, cytokines and chemokines were measured predose and 8 hours postdose. On Days 5 and 8, cytokines and chemokines were measured predose and at 2 and 8 hours postdose. In the one cohort of patients re- ceiving PT-100 on the Day 5–11 schedule, blood samples were collected at the following timepoints: Days 1, 5, 6, 8, 9, 11, 12, 14, 16, and 18. On Days 5 and 11 cytokines and chemokines were measured predose and at 2 hours and 8 hours postdose.
Pharmacokinetics
Blood samples for analysis of PT-100 pharmacokinetics cir- culating maximum concentration (Cmax), maximum time of de- tection (Tmax) and area under the curve (AUC) were collected at the following times: Days 1 and 7 of PT-100 administration (baseline; 15 and 30 minutes postdose; and 1, 2, 3, 4, 6, 8, 12 hours postdose), on Days 2 and 4 (trough level) of PT-100 ad- ministration, and on Cycle 2, Days 10, 12, 14, 16, 18, and 20. Plasma samples were analyzed for PT-100 concentration by Mi- croconstants, San Diego, CA.
RESULTS
Patient population
A total of 41 patients entered Cycle 1, and 34 patients (8 men and 26 women) received PT-100 in Cycle 2. Seven patients who entered into Cycle 1 did not achieve sufficient neutrophil reduction (n = 5) or developed progressive disease (n = 2) and, thus, were not eligible to receive PT-100. The median age of patients was 60 years (range 28 to 74 years). Disease and prior chemotherapy for their primary cancer are shown in Table 1. A
PT-100 Dose Cohort
Gender n (%)
200 µg (n = 6) 400 µg (n = 6) 800 µg (n = 9) 800 µg∗ (n = 8) 1,200 µg (n = 5)
Male 1 (17) 1 (17) 0 (0) 3 (38) 3 (60)
Female 5 (83) 5 (83) 9 (100) 5 (63) 2 (40)
Age median (range) Race n (%)
54.5 (28–74) 62.5 (47–72) 63 (37–68) 62.5 (39–72) 57 (31–70)
Caucasian 6 (100) 5 (83) 9 (100) 5 (63) 5 (100)
Black
Type of Cancer n (%)
0 1 (17) 0 3 (38) 0
Breast 4 (67) 3 (50) 8 (89) 4 (50) 0
Cervical 0 0 0 0 1 (20)
Colorectal 0 0 0 1 (13) 0
Head and neck 0 0 0 1 (13) 0
Liposarcoma 0 0 0 0 1 (20)
NSCLC 1 (17) 1 (17) 1 (11) 0 1 (20)
Mesothelioma 0 0 0 1 (13) 0
Uterine 1 (17) 0 0 1 (13) 0
Pancreatic 0 2 (33) 0 0 1 (20)
Squamous cell skin cancer
# Prior Chemotherapy Regimens
0 0 0 0 1 (20)
Median (range)
∗ Administered on Days 5–11.
1.5 (1–4) 1 (0–4) 0 (0) 1 (0–2) 2 (0–4)
Cytokine-Inducing Small Molecule 555
Table 2. Summary of concurrent chemotherapy by dose
800 µg 800 µg
200 µg 400 µg (Days 2–8) (Days 5–11) 1,200 µg All PT-100
(n = 6) (n = 6) (n = 7) (n = 6) (n = 4) (n = 34)
Doxorubicin/docetaxel 5 2 3 1 0 11
Doxorubicin/cyclophosphamide 0 1 3 2 0 6
Carboplatin/paclitaxel 1 2 1 3 0 7
Docetaxel (single-agent) 0 1 0 0 4 5
summary of concurrent chemotherapy by dose cohort is shown in Table 2. Factors such as age, sex, race, type of cancer, and chemotherapy regimen did not appear to affect tolerability, tu- mor response, cytokine profile, pharmacodynamics, or phar- macokinetics. Safety data are presented within for the 34 pa- tients treated with PT-100 in Cycle 2. Only patients (n = 29) who received at least 5 days of PT-100 are included in the summaries of the effects of PT-100 on neutrophil recovery, cy- tokine/chemokine upregulation, and pharmacokinetics.
Safety and tolerability
Criteria were not met for dose-limiting toxicity in this study; thus, an MTD was not achieved at daily PT-100 doses up to 1,200 µg. Promising preclinical data on the antitumor activity of PT-100 were generated during the conduct of this study, and it was decided to further explore the dose and dosing regimen of PT-100 in clinical studies to evaluate its antitumor effects; thus, further dose escalation in this study was not attempted.
Adverse events
Overall, PT-100 was well tolerated. Adverse events occurring in ≥5 percent of patients who received PT-100 in Cycle 2 are presented by decreasing frequency and dose in Table 3. Among the most frequently occurring events, the incidence of weakness, pyrexia, and febrile neutropenia were similar between Cycle 1 and Cycle 2.
Adverse events reported considerably more frequently in pa- tients receiving PT-100 in Cycle 2 compared with Cycle 1 were: edema in 15/34 patients (44.1 percent) versus 1/34 (2.9 percent); hypotension (including orthostatic) in 8/34 patients (23.5 per- cent) versus 4/34 (11.8 percent); dizziness 8/34 (23.5 percent) versus 5/34 (14.7 percent); hypovolaemia 5/34 (14.7 percent) versus 0; syncope 4/34 (11.8 percent) versus 1/34 (2.9 percent); and pruritis 3/34 (8.8 percent) versus 0. Edema, hypotension, and syncope were dose-related.
Adverse events occurring more frequently in Cycle 1 (in more than 10 patients) than in Cycle 2 were: nausea in 20/34 patients (58.8 percent) versus 5/34 (14.7 percent); alopecia 16/34 (47.1 percent) versus 2/34 (5.9 percent); fatigue 11/34 (32.4 percent) versus 7/34 (20.6 percent); vomiting 8/34 (32.4 percent) versus 5/34 (14.7 percent); constipation 10/34 (29.4 percent) versus 3/34 (8.8 percent); and diarrhea 10/34 (29.4 percent) versus 3/34 (8.8 percent).
The most frequently reported Grade 3 events were neutrope- nia, febrile neutropenia, leucopenia, infection, hypotension (in-
cluding orthostatic), and syncope. Two patients experienced a Grade 3 adverse event considered related to PT-100: orthostatic hypotension (800 µg/day) and syncope (1,200 µg/day). Grade
3and 4 events are summarized in Table 4.
There were 2 deaths during Cycle 1 due to progressive dis- ease. No deaths occurred during Cycle 2.
Pharmacodynamics
Twenty-nine patients received at least 5 days of PT-100 and, thus were included in this analysis. At PT-100 daily doses of 200 µg, 400 µg, 800 µg (Days 2–8), 800 µg (Days 5–11), and 1,200 µg, 2/6, 3/6, 4/7, 2/6, and 1/4 patients experienced at least a one-day improvement in Grade 4 neutropenia com- pared to Cycle 1. All 4 of the 7 patients who received PT-100 at the 800 µg/day on the Days 2–8 schedule also demonstrated at least a 2-day improvement in Grade 4 neutropenia in Cycle 2 versus Cycle 1 (Table 5), and 2 of these patients had a 4- and 5-day improvement. Only 1 of 6 patients receiving PT-100 on the 800 µg/day Days 5–11 schedule showed at least a 2-day improvement in Grade 4 neutropenia. The mean time to Grade
4neutropenia in these 2 cohorts ranged from 9.7 to 11.0 days, and the mean time to recovery ranged from 5.0 to 11.0 days.
Cytokine response
Cytokine levels were summarized on Days 5, 8, 11, and 18. In general, IL-6, G-CSF, and IL-8 levels were increased in both Cycles 1 and 2. In patients with at least a 2-day improvement in Grade 4 neutropenia in Cycle 2, the upregulation in cytokines generally tracked the decrease and subsequent increase in abso- lute neutrophil count (Figure 2). The mean time to peak levels of G-CSF, IL-6, and IL-8 occurred on Day 11 in both cycles in the 3 dosing cohorts. All samples for IL-11 remained below the limit of quantitation.
A comparison of increases in cytokines (G-CSF, IL-6, IL-8) from baseline (Cycle 1, Day 1) between Cycles 1 and 2 was conducted both for patients who had at least a one-day improve- ment in Grade 4 neutropenia and those who did not. In Cycle 2, cytokines values post-PT-100 initiation were included in this comparison. Two patients (one treated with 800 µg Days 5– 11 and one at 1,200 µg days 2–8) did not have complete cy- tokine panels; neither patient showed an improvement in Grade
4neutropenia in Cycle 2. The results, presented as maximum fold-increase in cytokines from baseline, are shown in Table 6.
556 J. Nemunaitis et al.
Table 3. Adverse events reported in ≥5 percent of PT-100 patientsa,b
N (% of patients)
PT-100 Dose
MedDRA Term Cycle 1 control (n = 34) 200 µg (n = 6) 400 µg (n = 6) 800 µgc (n = 17) 1,200 µg (n = 5) All PT-100 (n = 34)
Weakness 9 (26.5) 3 (50.0) 1 (16.7) 6 (35.3) 0 10 (29.4)
Anemia 5 (14.7) 1 (16.7) 0 6 (35.3) 1 (20.0) 8 (23.5)
Dizziness 5 (14.7) 2 (33.3) 1 (16.7) 4 (23.5) 1 (20.0) 8 (23.5)
Edema peripheral 1 (2.9) 0 0 6 (35.3) 2 (40.0) 8 (23.5)
Fatigue 11 (32.4) 0 2 (33.3) 3 (17.6) 2 (40.0) 7 (20.6)
Pyrexia 9 (26.5) 0 0 5 (29.4) 2 (40.0) 7 (20.6)
Hypotension 3 (8.8) 0 2 (33.3) 4 (23.5) 0 6 (17.6)
Nausea 20 (58.8) 1 (16.7) 2 (33.3) 2 (11.8) 0 5 (14.7)
Headache 9 (26.5) 3 (50.0) 1 (16.7) 1 (5.9) 0 5 (14.7)
Vomiting 8 (23.5) 1 (16.7) 1 (16.7) 3 (17.6) 0 5 (14.7)
Neutropenia 7 (20.6) 0 1 (16.7) 2 (11.8) 2 (40.0) 5 (14.7)
Edema/fluid retention 0 0 0 5 (29.4) 0 5 (14.7)
Hypovolaemia 0 0 3 (50.0) 2 (11.8) 0 5 (14.7)
Febrile neutropenia 5 (14.7) 1 (16.7) 0 3 (17.6) 0 4 (11.8)
Pain 2 (5.9) 0 0 4 (23.5) 0 4 (11.8)
Anorexia 2 (5.9) 1 (16.7) 0 3 (17.6) 0 4 (11.8)
Syncope 1 (2.9) 0 1 (16.7) 2 (11.8) 1 (20.0) 4 (11.8)
Constipation 10 (29.4) 0 1 (16.7) 1 (5.9) 1 (20.0) 3 (8.8)
Diarrhoea 10 (29.4) 0 2 (33.3) 1 (5.9) 0 3 (8.8)
Leukopenia 2 (5.9) 0 0 3 (17.6) 0 3 (8.8)
Rhinorrhoea 2 (5.9) 1 (16.7) 1 (16.7) 1 (5.9) 0 3 (8.8)
Pain in limb 1 (2.9) 1 (16.7) 0 2 (11.8) 0 3 (8.8)
Rigors 1 (2.9) 1 (16.7) 0 2 (11.8) 0 3 (8.8)
Pruritis 0 0 0 3 (17.6) 0 3 (8.8)
Vaginosis fungal 0 1 (16.7) 0 2 (11.8) 0 3 (8.8)
Alopecia 16 (47.1) 1 (16.7) 0 1 (5.9) 0 2 (5.9)
Arthralgia 5 (14.7) 0 0 2 (11.8) 0 2 (5.9)
Back pain 4 (11.8) 1 (16.7) 0 0 1 (20.0) 2 (5.9)
Appetite decreased 3 (8.8) 0 0 2 (11.8) 0 2 (5.9)
Dehydration 2 (5.9) 0 0 2 (11.8) 0 2 (5.9)
Dyspnea 2 (5.9) 0 0 1 (5.9) 1 (20.0) 2 (5.9)
Dysgeusia 2 (5.9) 0 1 (16.7) 1 (5.9) 0 2 (5.9)
Stomatitis 2 (5.9) 1 (16.7) 1 (16.7) 0 0 2 (5.9)
Neck pain 1 (2.9) 0 0 1 (5.9) 1 (20.0) 2 (5.9)
Orthostatic hypotension 1 (2.9) 0 0 2 (11.8) 0 2 (5.9)
Sweating increased 1 (2.9) 0 0 2 (11.8) 0 2 (5.9)
Chest pain 0 0 1 (16.7) 1 (5.9) 0 2 (5.9)
Deep venous thrombosis 0 0 0 2 (11.8) 0 2 (5.9)
Face edema 0 0 0 1 (5.9) 1 (5.9) 2 (5.9)
Hoarseness 0 0 2 (33.3) 0 0 2 (5.9)
Pneumonia 0 0 0 1 (5.9) 0 2 (5.9)
Vision blurred 0 0 0 2 (11.8) 0 2 (5.9)
aCycle 2, Day 1 adverse events for 3 patients are counted in Cycle 1 since PT-100 was not administered until Cycle 2, Day 2.
bTotals may not add up within a body system if a patient experienced more than one adverse event (AE) within a body system.
cPT-100 800 µg/day groups are combined; there was no difference in AEs between dosing schedules.
Pharmacokinetics
Following oral administration of PT-100, dose-related in- creases in plasma concentration were observed (Table 7). Mean Cmax values after the first PT-100 dose were 1.35, 2.96, 6.48, and 9.15 ng/mL for doses of 100 µg, 200 µg, 400 µg (Days 2–8), and 600 µg, respectively. Corresponding mean AUC(0- 12) values were 9.09, 17.8, 33.0, and 54.1 ng-hr/ml. Sim- ilar mean Cmax and AUC(0-12) values were noted for the 800 µg/day dose group (400 µg administered twice daily on Days 5–11) as with the Days 2–8 dosing schedule. Af-
ter 13 doses of PT-100, increases in both Cmax and AUC(0- 12) were observed. Mean Cmax values were 1.95, 4.67, 7.38, and 13.1ng/mL for the 100 µg, 200 µg, 400 µg (Days 2–8), and 600 µg dose groups, respectively. Corresponding mean AUC(0-12) values were 15.0, 32.2, 50.0, and 87.2 ng-hr/ml. The mean Cmax and AUC(0-12) for the 800 µg/day dose group (400 µg administered twice daily on Days 5–11) were higher (10.4 and 79.9 ng-hr/ml, respectively) than that of the Days 2–8 schedule.
A comparison of mean Cmax and AUC(0-12) between responders (patients who experienced at least a one-day
Cytokine-Inducing Small Molecule 557
Table 4. Grade 3 or 4 AEs reported in ≥2 patients by dose
N (%) of patients
Cycle 1
Grade 3 control (n = 34) 200 µg (n = 6) 400 µg (n = 6) 800 µga (n = 17) 1,200 µg (n = 5) All PT-100 (n = 34)
Neutropenia 7 (20.6) 0 1 (16.7) 2 (11.8) 1 (20.0) 4 (11.8)
Febrile neutropenia 2 (5.9) 1 (16.7) 0 3 (17.6) 0 3 (8.8)
Leukopenia 2 (5.9) 0 0 3 (17.6) 0 3 (8.8)
Infection 1 (2.9) 0 1 (16.7) 2 (11.8) 0 3 (8.8)
Hypotension 0 0 1 (16.7) 1 (5.9) 1 (20.0) 3 (8.8)
Syncope 0 0 1 (16.7) 1 (5.9) 1 (20.0) 3 (8.8)
Anemia 1 (2.9) 0 0 1 (5.9) 1 (20.0) 2 (5.9)
Pyrexia/increased temp. 0 0 0 1 (5.9) 1 (20.0) 2 (5.9)
Weakness 0 0 0 2 (11.8) 0 2 (5.9)
Dehydration 1 (2.9) 0 0 1 (5.9) 0 1 (2.9)
Nausea 1 (2.9) 0 0 1 (5.9) 0 1 (2.9) Grade 4
Neutropenia 4 (11.8) 0 0 0 2 (40.0) 2 (5.9) a PT-100 800 µg/day groups are combined; there was no difference in AEs between dosing schedules.
Table 5. Number of patients with a 2+ day improvement in Grade 3+ neutropenia in Cycle 2 versus Cycle 1a
Dose groups (total dose/day)
200 µg (n = 6) 400 µg (n = 6) 800 µg (n = 7) 800 µg (n = 6) 1200 µg (n = 4)
Grade 3 Neutropenia 1 0 0 1 0
Grade 4 Neutropenia 1 1 3 1 1 a Patients who received at least 5 days of PT-100.
Table 6. Fold increase from baseline in G-CSF, IL-6, and IL-8a
Patient #b
(responsec ) Dose/Schedule G-CSF IL-6 IL-8
C1 C2 C1 C2 C1 C2
5502(n) 800 µg D2-8 25.5 8.2 18.6 17.0 5.4 6.6
5504 (n) 800 µg D2-8 9.3 2.4 57.0 24.9 20.4 6.7
5507 (n) 800 µg D2-8 4.4 4.4 2.4 3.7 4.9 1.5
5503(r) 800 µg D2-8 12.6 9.5 122.5 57.8 20.2 15.4
5509(r) 800 µg D2-8 2.5 5.2 15.9 11.3 1.9 2.1
5510(r) 800 µg D2-8 13.2 5.1 10.4 68.3 5.2 6.2
5511(r) 800 µg D2-8 1.6 2.4 1.4 3.5 1.7 2.1
1603 (n) 800 µg D5-11 1.6 2.7 6.8 12.0 1.2 1.2
5610 (n) 800 µg D5-11 0 3.5 16.0 909.0 3.7 28.8
6608 (n) 800 µg D5-11 10.1 8.4 14.8 28.9 1.9 2.2
5601 (r) 800 µg D5-11 11.3 63.6 39.2 251.4 27.4 23.4
5609 (r) 800 µg D5-11 10.2 1.9 12.3 4.8 10.3 5.7
1706(n) 1,200 µg D2-8 32.9 64.0 3.0 2.5 1.8 2.4
1708 (n) 1,200 µg D2-8 6.4 2.2 33.0 6.6 38.8 18.5
1707(r) 1,200 µg D2-8 3.3 6.2 1.8 2.3 4.2 5.5
aBaseline is Cycle 1 Day 1. Post-PT-100 peak cytokine levels are presented for Cycle 2.
bTwo patients (6607 and 1702) did not have a complete cytokine panel and are not included.
cResponders (r) are defined as patients with ≥one-day improvement in Grade 4 neutropenia. Nonresponders (n) are defined as patients with <1-day improvement in Grade 4 neutropenia.
558 J. Nemunaitis et al.
Figure 2. Comparison of ANC and plasma cytokine levels for a subject receiving 800 µg PT-100 BID from Days 2–8 in Cycle 2 of chemotherapy. Time-dependent changes in ANC (■) and G-CSF levels (■) were compared for Cycles 1 and 2 of chemotherapy. The horizontal line indicates the ANC level beneath which a subject would be considered neutropenic. Changes in IL-6 (▲) and IL-8 (♦) levels are shown for Cycles 1 and 2. (▲) indicates time points of Talabostat administrations.
improvement in Grade 4 neutropenia in Cycle 2 versus Cy- cle 1) and nonresponders (those who had no change or worsened between Cycles 1 and 2) was performed. The most data were available for the PT-100 dose cohort of 800 µg/day (Days 2–8).
At this dose, mean Cmax and AUC(0-12) after 7 days of dos- ing were substantially higher in responders than non-responders (Table 8). In the 400 µg/day group, mean plasma levels and AUC(0-12) were higher in nonresponders.
Table 7. Summary of pharmacokinetic parameters by dose group
Dose groups (total dose/day)
200 µg 400 µg 800 µg 800 µg 1,200 µg
Day: 2 8 2 8 2 8 5 11 2 8
N= Parameter
6
6
6
6
9 7 Total Plasma
7
5
5
3
Cmax, ng/mL 1.35 1.95 2.96 4.67 6.48 7.38 7.15 10.4 9.15 13.1
Tmaxa , h 2.00 3.00 1.04 2.00 2.00 1.00 1.25 1.08 2.00 2.00
AUC (0-12), ng · h/mL 9.09 15.0 17.8 32.2 33.0 50.0 39.2 79.9 54.1 87.2
a Expressed as median.
N = number of patients with samples.
Cytokine-Inducing Small Molecule 559
Table 8. Summary of mean Cmax and AUC(0-12) by dose group
Plasma levels: Mean (SD)
PT-100 total
daily dosea n/rc Cmax—Nonrespondersc Cmax—Responders AUC—Nonresponders AUC—Responders
Day 2 or 5 Day 8 or 11 Day 2 or 5 Day 8 or 11 Day 2 or 5 Day 8 or 11 Day 2 or 5 Day 8 or 11
200 µg 4/2 1.34 (0.45) 1.94 (1.67) 1.38 (0.12) 1.98 (0.91) 9.20 (2.71) 15.78 (9.12) 8.68 (0.10) 13.29 (1.89)
400 µg 3/3 3.41 (0.91) 5.10 (0.50) 2.52 (0.30) 4.23 (2.20) 22.16 (5.53) 37.72 (4.15) 13.54 (0.99) 26.61 (11.99)
800 µg 3/4 6.61 (2.71) 5.34 (1.14) 6.33 (0.55) 8.91 (3.99) 31.84 (8.27) 39.04 (6.48) 34.46 (4.06) 58.27 (23.06)
800 µgb 3/2 8.41 (7.27) 11.41 (3.71) 4.01 (1.63) 6.21 (0.00) 47.33 (37.49) 90.85 (31.99) 19.13 (2.96) 36.27 (0.00)
1,200 µg 2/1 8.94 (3.05) 14.50 (4.67) 10.00 (0.00) 10.30 (0.00) 52.42 (17.38) 87.57 (7.05) 60.75 (0.00) 86.12 (0.00)
aPT-100 administered on Days 2–8 except for regimen marked byb .
bPT-100 administered on Days 5–11.
cn/r = (# nonresponders)/(# responders). Responders are defined as patients with ≥one-day improvement in Grade 4 neutropenia. Nonresponders are defined as patients with <one-day improvement in Grade 4 neutropenia.
DISCUSSION
This is the first study to evaluate the safety of PT-100 and its effects on neutrophil recovery and up-regulation of key hematopoietic cytokines in patients receiving myelosuppressive chemotherapy. PT-100 total daily doses ranging from 200 µg to 1,200 µg (divided twice daily) generally were well tolerated. An MTD was not reached in this study. Adverse events reported con- siderably more frequently in patients receiving PT-100 in Cycle 2 compared with Cycle 1 were oedema, hypotension (including orthostatic), hypovolemia, syncope, and pruritis. Oedema, hy- potension, hypovolemia and dizziness have been previously re- ported in healthy subjects and patients receiving PT-100 in other studies; these events are consistent with a cytokine-mediated ef- fect. Interestingly, typical chemotherapy-related toxicity such as nausea, vomiting, alopecia, fatigue, constipation, and diar- rhea all appeared substantially lest frequently in patients who received PT-100 in Cycle 2 compared with Cycle 1.
This study was not statistically powered to adequately evalu- ate the effects of PT-100 on hematopoiesis or cytokine upregu- lation. However, through doses up to and including 800 µg/day, the data suggested a reduction in days of Grade 4 neutropenia when PT-100 was administered on Days 2–8. The mean time to Grade 4 neutropenia in both 800 µg/day cohorts ranged from 9.7 to 11.0 days, and the mean time to recovery ranged from
5to 11 days. Delaying the initiation of treatment with PT-100 800 µg/day to 5 days following chemotherapy showed only half as many patients with improved neutrophil recovery compared to Cycle 1. Too few patients were treated at the 1,200 µg/day PT-100 dose to allow for any conclusion as to the effects of this dose on neutrophil recovery.
In general, IL-6, G-CSF, and IL-8 levels were increased from baseline in both Cycles 1 and 2. In patients demonstrating at least a one-day improvement in Grade 4 neutropenia, the elevations were more pronounced in Cycle 2, and generally corresponded to the decrease and subsequent increase in ANC. The mean time to peak levels of G-CSF, IL-6, and IL-8 was on Study Day 11 in both cycles suggesting that the timing of the increase in cytokines was related to the onset of Grade 4 neutropenia.
ThepharmacokineticsofPT-100werelinearandproportional from single doses of 100 µg to 600 µg after the first dose. At the highest dose, the Cmax was 9.15 ng/mL and AUC(0-12) was 54.1 ng-hr/mL. Both parameters were higher after mul- tiple dosing of PT-100 for 7 days. The relationship between the parameters and dose continued to be linear. In the PT-100 800 µg/day dose group (Days 2–8), Cmax and AUC were sub- stantially higher in patients who showed at least a one-day im- provement in Grade 4 neutropenia.
As with most small studies, substantial variability was ob- served in the pharmacodynamic and pharmacokinetic data in patients. This variability may be due to differences in absorp- tion or the timing of PT-100 dosing with respect to meals. Nausea and vomiting were reported in 15 percent of patients receiving PT-100 in Cycle 2, and these events may have also impacted the drug’s bioavailability and potential activity.
PT-100 is a novel, orally-available agent that has shown both hematopoietic stimulation and anti-tumor effects in preclinical models. It was well-tolerated clinically at doses up to 1,200 µg/day in patients receiving myelosuppressive chemotherapy. A trend toward enhancing neutrophil recovery and reduced chemotherapy-related was noted in this small study, with the best effects seen at PT-100 800 µg/day admin- istered Days 2–8. In some patients with accelerated neutrophil recovery, enhanced cytokine upregulation was observed in Cycle 2. Further studies to evaluate both the antitumor effects of PT-100, its effects on reduction of chemotherapy-related toxicity, and its potential enhancement of immune-mediated antitumor effect when used in combination with rituximab or trastuzumab are warranted.
REFERENCES
1.Coutts, S.J.; Kelly, T.A.; Snow, R.J.; Kennedy, C.A.; Barton, R.W.; et al. Structure-activity relationships of boronic acid inhibitors of dipeptidyl peptidase IV. 1. Variation of the P2 position of Xaa – boroPro dipeptides. J. Med. Chem. 1996, 39, 2087–2094.
2.Cheng, J.D.; Valianou, M.; Canutescu, A.A.; Jaffe, E.K.; Lee, H.O.; et al. Abrogation of fibroblast activation protein enzymatic activity attenuates tumor growth. Mol. Cancer Ther. 2005, 4, 351–360.
560 J. Nemunaitis et al.
3.Jones, B.; Adams, S.; Miller, G.T.; Jesson, M.I.; Watanabe, T.; et al. Hematopoietic stimulation by a dipeptidyl peptidase inhibitor reveals a novel regulatory mechanism and therapeutic treatment for blood cell deficiencies. Blood 2003, 102, 1641–1648.
4.Coutts, S.J.; Kelly, T.A.; Snow, R.J.; Kennedy, C.A.; Barton, R.W.; et al. Structure-activity relationships of boronic acid inhibitors of dipeptidyl peptidase IV. 1. Variation of the P2 position of Xaa- boroPro dipeptides. J. Med. Chem. 1996, 39, 2087–2094.
5.Flentke, G.R.; Munoz, E.; Huber, B.T.; Plaut, A.G.; Kettner, C.A.; et al. Inhibition of dipeptidyl aminopeptidase IV (DP-IV) by Xaa- boroPro dipeptides and use of these inhibitors to examine the role of DP-IV in T-cell function. Proc. Natl. Acad. Sci. USA 1991, 88, 1556–1559.
6.von Bonin, A.; Huhn, J.; Fleischer, B. Dipeptidyl-peptidase IV/CD26 on T cells: analysis of an alternative T-cell activation pathway. Im- munol. Rev. 1998 161, 43–53.
7.Adams, S.; Miller, G.T.; Jesson, M.I.; Watanabe, T.; Jones, B.; et al. PT-100, a small molecule dipeptidyl peptidase inhibitor, has potent antitumor effects and augments antibody-mediated cytotoxicity via a novel immune mechanism. Cancer Res. 64, 5471–5480.
8.Garin-Chesa, P.; Old, L.J.; Rettig, W.J. Cell surface glycoproteins of reactive stromal fibroblasts as a potential antibody target in human epithelial cancers. Proc. Natl. Acad. Sci. USA 1990, 87, 7235– 7239.
9.Park, J.E.; Lenter, M.C.; Zimmerman, R.N.; Garin-Chesa, P.; Old, L.J.; et al. Fibroblast activation protein, a dual-specificity serine protease expressed in reactive human tumor stromal fibroblasts. J. Biol. Chem. 1999, 274, 36505–36512.
10.Jones, B.; Adams, S.; Miller, G.; Jesson, M.I.; Watanabe, T.; et al. Hematopoeitic stimulation by a dipeptidyl peptidase in- hibitor reveals a novel regulatory mechanism and therapeutic treatment for blood cell deficiencies. Blood 2003, 102, 1641– 1648.
11.Aoyama, A.; Chen, W.-T. A 170-kDa membrane-bound protease is associated with the expression of invasiveness by human ma- lignant melanoma cells. Proc. Natl. Acad. Sci. 1990, 87, 8296– 8300.
12.Kelly, T.; Kechelava, S.; Rozypal, T.; West, K.; Korourian, S. Seprase, a membrane-bound protease is overexpressed by inva- sive ductal carcinoma cells of human breast cancers. Mod. Pathol. 1998, 11, 855–863.
13.Adams, S.; Miller, G.T.; Jesson, M.I.; Watanabe, T.; Jones, B.; et al. PT-100, a small molecule dipeptidyl peptidase inhibitor, has po- tent antitumor effects and augments antibody-mediated cytotoxic- ity via a novel immune mechanism. Cancer Res. 2004, 64, 5471– 5480.
14.Jones, B.; Adams, S.; Miller, G.; Jessen, M.; Wallner, B. Data on file. Point Therapeutics, Inc., Boston, MA.
Cytokine-Inducing Small Molecule 561