A Phase 1b/2 trial of mapatumumab in patients with relapsed/refractory non-Hodgkin's lymphoma

Background: We conducted a multicentre Phase 1b/2 trial to evaluate the safety and efficacy of mapatumumab, a fully human agonistic monoclonal antibody to the tumour necrosis factor-related apoptosis-inducing ligand receptor 1 (TRAIL-R1) in patients with relapsed non-Hodgkin's lymphoma (NHL). Methods: Forty patients with relapsed or refractory NHL were treated with either 3 or 10 mg kg−1 mapatumumab every 21 days. In the absence of disease progression or prohibitive toxicity, patients received a maximum of six doses. Results: Mapatumumab was well tolerated, with no patients experiencing drug-related hepatic or other dose-limiting toxicity. Three patients with follicular lymphoma (FL) experienced clinical responses, including two with a complete response and one with a partial response. Immunohistochemistry staining of the TRAIL-R1 suggested that strong staining in tumour specimens did not appear to be a requirement for mapatumumab activity in FL. Conclusions: Mapatumumab is safe and has promising clinical activity in patients with FL.

monoclonal antibody specific and agonistic to the tumour necrosis factor-related apoptosis-inducing ligand receptor 1 (TRAIL-R1, DR4). The TRAIL-R1 is expressed more frequently on the surface of tumour cells than on the surface of normal cells (Zang et al, 2001;Koornstra et al, 2003). Mapatumumab, like the native ligand TRAIL, mediates apoptosis by binding to TRAIL-R1, leading to activation of the caspase cascade, cleavage of key intracellular signalling components and DNA and subsequent cell death (Pukac et al, 2005). Mapatumumab induces apoptosis in a wide range of haematologic and solid tumour cell lines and xenograft models (Snell et al, 1997;Georgakis et al, 2005;Pukac et al, 2005;Menoret et al, 2006). In addition to a direct induction of apoptosis, mapatumumab may also induce antibody-dependent cellular cytotoxicity. Furthermore, in vivo experiments using disseminated B cell lymphoma models showed that mapatumumab in combination with rituximab increased animal survival compared with control, rituximab alone or mapatumumab alone (Maddipatla et al, 2007).
Mapatumumab was evaluated first as a single agent and subsequently in combination with chemotherapeutic agents in a series of Phases 1, 1b and 2 clinical trials (Tolcher et al, 2007;Greco et al, 2008;Hotte et al, 2008;Mom et al, 2009;Trarbach et al, 2010). In general, it is well tolerated both as a single agent and in combination with chemotherapy. No disease response was observed in the other single-agent trials. This clinical trial was designed to evaluate single-agent mapatumumab in non-Hodgkin's lymphoma (NHL). The doses and design were based on data available from the initial and on-going Phase 1 trials of mapatumumab (Tolcher et al, 2007;Hotte et al, 2008).

Patients
Eligibility criteria included histologically confirmed NHL with measurable disease X1.5 cm in the longest transverse diameter by computed tomography scan. Patients were previously treated with at least one therapeutic regimen and had relapsed or progressed, or failed to achieve an objective response after their last therapeutic regimen. Adequate haematologic, bone marrow, hepatic and renal functions, and an Eastern Cooperative Oncology Group performance status 0 -2 were required. Patients were excluded if they had been treated with a monoclonal antibody or radioimmunotherapy within 8 weeks or had persistent clinical evidence of toxicity, were eligible for stem cell transplantation (SCT), had undergone autologous SCT within 16 weeks or had ever undergone allogeneic SCT. The IRB-approved, written informed consent was obtained from all patients.

Study design
This multicentre, open-label study was designed to evaluate the safety and efficacy of mapatumumab in patients with relapsed or refractory NHL. At the time this study was initiated, safety data were available from on-going Phase 1 trials for doses up to 3 mg kg À1 every 28 days. This trial was designed to allow for treatment of a small number of NHL patients with 3 mg kg À1 every 21 days before treating a larger number of patients with 10 mg kg À1 every 21 days. Specifically, safety data from eight patients treated with 3 mg kg À1 mapatumumab were reviewed before enrolment of the 10 mg kg À1 cohort. Mapatumumab was administered intravenously (i.v.) over 2 h in a total volume of 250 ml normal saline. No dose reduction was allowed. Premedication with diphenhydramine and acetaminophen was allowed and usually given. Patients were treated on day 1 of each 21( ± 2)-day cycle for up to six cycles in the absence of disease progression or dose-limiting toxicity. Patients who demonstrated a tumour response after six cycles of treatment could receive permission to continue treatment. Disease assessment was performed at baseline, and after cycles three and six, and then every 3-month intervals until disease progression. Disease response was evaluated using response criteria according to the International Working Group Recommendations for NHL (Cheson et al, 1999). Adverse events (AEs) and clinical laboratory test results were graded based on the NCI-CTCAE (Version 3.0).
Plasma mapatumumab concentrations were measured before dosing, immediately post-dosing and 4 -6 h post-dosing for the first mapatumumab administration. Additional sampling was performed before dosing, immediately post-dosing and 24 h post-dosing after the second administration, and before dosing and day 17 (day of disease assessment) for the third and sixth administration. Anti-mapatumumab antibodies were analysed using an enzyme-linked immunosorbent assay. Serum samples were obtained at baseline, prior to dosing in each treatment cycle, at cycle 1, day 17 and 28 -30 days after the last dose of mapatumumab. Formalin-fixed, paraffin-embedded tissue blocks or 4 -6 mm tissue sections from biopsies collected at baseline, post-dose or from archival specimens were requested from all patients and collected as available. The immunohistochemistry (IHC) staining method utilised standardised reagents according to a protocol supplied by DAKO (Carpinteria, CA, USA) for this prototype pharmDx test intended for investigative use (Humphreys and Halpern, 2008).

Statistical considerations
Progression-free survival was calculated from baseline to disease progression (or death) using the Kaplan -Meier method. To assess potential relationships between AEs and plasma mapatumumab concentrations, dose normalised plasma mapatumumab concentrations at selected time points were compared among patients who experienced serious adverse event (SAEs) and those who did not, only if nX3 for each subgroup, using two-sided unpaired t-tests at a significance level of a ¼ 0.05.

RESULTS
Among the 40 patients treated in this study, 8 received 3 mg kg À1 mapatumumab and 32 received 10 mg kg À1 mapatumumab. Patient demographics and baseline disease characteristics are summarised in Table 1. The majority of patients (26 out of 40, 68%) completed at least 3 cycles and 12 patients received the full 6 cycles. Two patients receiving 10 mg kg À1 demonstrated a tumour response after six cycles and received permission to continue mapatumumab until disease progression. One patient received 12 cycles, discontinued the study for an unrelated medical condition and, at the time of discontinuation, had an increase in tumour size not yet reaching disease progression. The other patient received 27 cycles before disease progression. Three patients with follicular lymphoma (FL) (Grade 1 or 2) achieved major responses: one complete response (CR) in the 3 mg kg À1 cohort, and one partial response (PR) and one CR in the 10 mg kg À1 cohort. The median time to response (CR or PR) was 3.9 months (range: 1.8 -4.1 months). The patient in the 3-mg kg À1 cohort achieved a PR after completing six cycles of treatment; her tumour continued to shrink, and she achieved a CR at 11.4 months. The mechanism of this continued response is unknown, but based on the plasma halflife of mapatumumab, the subject would have been exposed to mapatumumab for several months after discontinuing drug. This patient had disease progression at 26 months. The two patients in the 10 mg kg À1 cohort who achieved disease response after six cycles received IRB permission to continue on mapatumumab. The patient with the PR received a total of 12 cycles and was taken off study to allow for treatment of bilateral urethral stones. This patient had progressing disease at the time of discontinuation at 8.5 months with a 45.9% increase in tumour. The second patient received 27 cycles and converted from a PR to a CR at 9.3 months. The patient exhibited progressive disease after 21 months. A total of six patients experienced tumour shrinkage following mapatumumab therapy ( Figure 1A). The median PFS for all patients in the study was 2 months ( Figure 1B). For the subset of 17 FL patients, the median PFS was 6 months (95% CI: 2.1 -21 months).  Table 2. Most of these events were Grade 1 or 2 in severity. The most common AEs considered at least possibly related to mapatumumab included nausea, fatigue, diarrhoea, anorexia and pyrexia. These events were Grade 1 or 2 with the exception of a single event of Grade 3 pyrexia. Eleven patients experienced 29 SAEs. Three SAEs reported in the 10 mg kg À1 cohort were considered either possibly, probably or definitely related to treatment with mapatumumab. One patient experienced peripheral, facial and angioneurotic oedema, and discontinued treatment. The other two SAEs, pyrexia and herpes zoster occurred in a single patient each. Laboratory data showed no evidence of significant hepatotoxicity or renal toxicity. One patient developed Grade 3 GGT levels. One developed Grade 2 hyperbilirubinemia. Two patients developed hyperamylasemia (Grade 3 or 4). No patient developed anti-mapatumumab antibodies.
The mean serum mapatumumab concentration -time profiles are illustrated in Figure 2. The observed plasma mapatumumab concentrations tended to be higher than those predicted from the Phase 1 trials of mapatumumab in solid tumours and is due to a slower clearance of mapatumumab in these study patients (Tolcher et al, 2007;Hotte et al, 2008). The terminal elimination half-life (t 1/2,z ) was B26 days in NHL patients compared with 12 -24 days seen in patients with solid tumours (Tolcher et al, 2007;Hotte et al, 2008).
Twenty-four patients provided a total of 29 evaluable tissue specimens: 18 archival specimens, 7 specimens before treatment and 4 post-treatment. Of these, TRAIL-R1 was detected in 16 specimens from 14 patients, with at least 10% of tumour cells staining. Tumours with an aggressive phenotype demonstrated the strongest TRAIL-R1 staining. The TRAIL-R1-specific staining was not detected or was weak in the archival or baseline specimens from two of the three patients with FL who responded to mapatumumab; biopsy material from the third patient was not evaluable.

DISCUSSION
This is the first trial to demonstrate disease response from singleagent mapatumumab in lymphoma. The results introduce three important questions: (1) what dose(s) of mapatumumab should be evaluated in future studies of mapatumumab in NHL; (2) whether staining for TRAIL-R1 by IHC can help identify patients most likely to benefit from mapatumumab and (3) whether mapatumumab should be further evaluated as a single agent for NHL or in combination with other therapeutics that target relapsed or refractory NHL.