Afatinib

Treatment group A (conventional chemotherapy intervention): on the 1st and 8th days, patients were given 100 mg/m² gemcitabine hydrochloride mixed by intravenous drip (Qilu Pharmaceutical Co., Ltd., H20113285) and 250 ml sodium chloride injection (Harbin triple drug Industrial Co., Ltd., approved by Chinese medicine H20184091), 30 mg/m² cisplatin (Qilu Pharmaceutical Co., Ltd., approved by Chinese medicine, H37021362) was intravenously infused on the second, third, and fourth days, once a day, for 3 months. Treatment group B (targeted therapy intervention with afatinib based on the treatment of group A): patients were given warm water orally with afatinib (Boehringer Ingelheim Pharma GmbH & Co., Germany, National Medicine Zhunzi J20170028), 3 times a day, continuous treatment 3 months.

The human lung adenocarcinoma cell line, PC-9, established from an untreated patient as previously described [10 (link)], was donated by K. Hayata. PC-9 cells, which were cultured in RPMI-1640 medium with 10% fetal bovine serum, penicillin (100 U/mL), and streptomycin (100 µg/mL) in a 5% CO₂ incubator at 37 °C. The cells were passaged for less than 4 months before renewal from frozen stocks. The cell lines were authenticated using short tandem repeat analysis at the Japanese Collection of Research Bioresources Cell Bank (Promega, Madison, WI) and tested for Mycoplasma using a MycoAlert Mycoplasma detection kit (Lonza, Basel, Switzerland). Afatinib and xentuzumab were from Boehringer-Ingelheim (Ingelheim am Rhein, Germany); other inhibitors and chemicals were obtained from Selleck Chemicals (Houston, TX, USA) and Sigma-Aldrich (St. Louis, MO, USA), respectively.

Thirty-one Japanese patients with EGFR mutation-positive NSCLC (15 women and 16 men) who were hospitalized from October 2014 through December 2020 were consecutively enrolled in this study. The grade for diarrhea was determined based on CTCAE version 4.0. Three patients (2 women and 1 man) were excluded because of withdrawal due to CTCAE grade 3 diarrhea just after beginning and before blood sampling for afatinib pharmacokinetics. Patient characteristics at the start of afatinib therapy are listed in Table 1. The study protocol was approved by the Ethics Committee of Akita University School of Medicine (approval no. 790), and all patients gave written informed consent. This study was performed in accordance with the guidelines of the Declaration of Helsinki.
An initial dose of 30 or 40 mg afatinib (Giotrif; Boehringer Ingelheim, Tokyo, Japan) was orally administered once daily at a designated time (11:00 a.m.). On day 15 after beginning afatinib therapy, whole blood samples were collected just prior to (C₀, 24 h after the 14th administration) and at 1, 2, 4, 6, 8, 12, and 24 h after the 15th administration of afatinib. Plasma was isolated by centrifugation at 1900× g for 15 min and was stored at −80 °C until analysis. For the 15 days prior to plasma sampling, nurses managed the administration of afatinib for hospitalized patients.