Study design
This monocentric, open label dose-escalation phase I clinical trial enrolled 21 patients with NSCLC, in the National Cancer Center in Beijing, China. All participants were informed about the study and potential risks and required to provide written informed consent prior to undergoing study-related procedures.
A traditional 3 + 3 dose escalation design was implemented [
12]. Successive cohorts of patients (3 patients/cohort) were each started on a fixed dose of the vaccine Hu-rhEGF-rP64k/Mont. Patients were assigned sequentially to one of four dose-escalation cohorts. The protocol specified 0.6 mg (group A) of the vaccine on days 4, 18, 32, 46 and 76 (5 vaccine dose), for the first cohort. Successive cohorts were given doses of 1.2 mg (group B), 1.8 mg (group C), 2.4 mg (group D) the same dose intervals. The dose groups were determined on the basis of the vaccine’s safety information available from previous clinical studies [
8,
13,
14]. Each vaccine dose was administered at 1 injection site, deltoid region (group A), 2 injection sites, deltoids regions (group B), 3 injection sites, deltoids regions and gluteus region (group C) and 4 injection sites, deltoids and gluteus regions (group D).
A DLT was defined as any Grade 3 or 4 adverse event (AE) using the CTCAE 4.03 that was possibly drug-related. CTCAE 4.03 Grade 3 is a severe AE and Grade 4 is a life-threatening or disabling AE. Such events interfere with activities of daily living and include: skin toxicity, diarrhea or antidiarrheal therapy, vomiting at same grade for > 4 days despite aggressive antiemetic therapy, central nervous system, lung or renal toxicity or elevation of liver transaminases or bilirubin lasting more than 1 week. The number of patients who experienced dose-limiting toxicities (DLTs) was assessed at each dose level. If no DLTs were observed for 4 weeks after administration of the last dose of the vaccine, a new cohort of 3 patients was enrolled at the next planned dose level. If DLTs were observed in 1 patient in the cohort, another 3 patients were treated with the same dose level.
The maximum-tolerated dose (MTD) was defined as 1 dose level below the dose in which DLTs were observed in > 33% of the participants. That is, if DLTs were observed in at least 2 of 3 participants, the MTD was determined to be the dose administered to the previous cohort. Similarly, in a cohort of 6 participants, 3 of 6 participants would have to experience DLTs to determine the MTD.
Toxicities were graded using the Common Terminology Criteria for Adverse Events Version 4.03 (CTCAE 4.03) [
15]. Health status assessments, including physical exams (vital signs, height, weight, ECOG score, skin of injection site, et al.), complete blood chemistry, electrocardiography was performed at baseline and post vaccination. Chest X-ray, computed tomography (CT) scan, and abdominal ultrasound were conducted at 46 days ±1 week and at 106 days ±1 week to assess clinical response according the Response Evaluation Criteria in Solid Tumors, version 1.1 (RECIST1.1). The objective response rate includes complete response (CR), partial response (PR), stable disease (SD), and progressive disease (PD). Disease control rate (DCR) was defined as CR + PR + SD.
Blood samples were collected at baseline (pre-treatment, day 1) and on days 18, 32, 46, 76 and 106 days post-dose for serum EGF concentration and anti-EGF antibody titers detection. Anti-EGF antibody titers were measured through a validated enzyme linked immunosorbent assay (ELISA), described in previous studies [
8,
10] and EGF concentration in serum was measured with a commercial ELISA (Quantikine EGF; R&D Systems Inc., Minneapolis, MN). The detection was conducted by the Beijing Eastern Biotech, Co., Ltd.
The protocol and informed consent documents were reviewed and approved by the hospital human subjects review board and the study was performed in accordance with the Declaration of Helsinki.
Patient eligibility
Inclusion criteria
Adult patients, regardless of gender, aged from 18 to 70 years with histologically or cytologically confirmed NSCLC at stages IIIB or IV and Eastern Cooperative Oncology Group (ECOG) performance status (PS) of 0 to 2 were eligible for the study. All patients received 4 to 6 cycles of platinum-based first-line chemotherapy, from 4 to 8 weeks, and were evaluated as SD or PR after the first line chemotherapy.
Patients were required to have at least 1 measurable lesion (lesions were defined by Response Evaluation Criteria in Solid Tumors (RECIST, 1.1) [
16]. If patients have ever received radiotherapy, the disease in the field of radiotherapy cannot be the target lesions and additional requirements were adequate organ function and CBC results and life expectancy of at least 3 months”. Women participants should have negative pregnancy test results. All the participants should take contraception measures during the study period. Participants should sign informed consent before the study.
Exclusion criteria
Exclusion criteria included patients receiving immunosuppressant drugs 1 month before study, or receiving other immunotherapy 3 months before the study and patients receiving corticosteroid within 1 month before the research (including oral, intramuscular or intravenous injection). Inhaled corticosteroid for respiratory insufficiency or for local use were not excluded).
Patients with uncontrolled epilepsy seizure; central nervous system dysfunction or cognitive ability lose due to psychosis; CNS metastasis (confirmed or suspicious); chronic alcohol or drug abuse within 6 months before the research; pregnancy or lactation or those who refused to take contraception measures during the study were excluded. Patients with a history of severe allergy or allergic constitution; splenectomy; autoimmune disease or secondary immunodeficiency (such as AIDS, et al.); secondary malignancy within 5 years (except for basal cell carcinoma of the skin or intraepithelial carcinoma of the cervix) were also excluded.
Treatment schedule
The therapeutic cancer vaccine is composed of hu-rhEGF conjugated to the carrier protein, r-P64K, and emulsified with the adjuvant Montanide ISA 51 VG, just prior to administration to patients. Hu-rhEGF-rP64k conjugate injection solution (batch number: 121005, 121006, 121203) and Montanide ISA 51 VG (batch number: 501101, 501201) were mixed in equal parts (0.8 mL of rhEGF/rP64k conjugate + 0.8 mL of Montanide ISA 51 VG). Hu-rhEGF-rP64k conjugate injection solution and the adjuvant were produced by the Center of Molecular Immunology (Havana, Cuba). CTX (Cyclophosphamide) injection solution (batch number: 04111001) was provided by BIOTECH PHARMA Co., Ltd.
Four to six weeks after finishing first-line chemotherapy, patients were assigned to the group of treatment with the vaccine. All patients received a low-dose of cyclophosphamide intravenously (200 mg/m2) on day 1. The vaccine was administered intramuscular at different dose levels: 0.6 mg (group A), 1.2 mg (group B), 1.8 mg (group C), and 2.4 mg (group D) on days 4, 18, 32, 46 and 76 (5 vaccine doses was administered for each patients. Monthly vaccination was administered according to the best benefits of patients and their wishes, and until the disease progressed or intolerable toxicity occurred.