A 61-year-old non-smoking Chinese male visited our hospital in August 2017, because of a persistent cough for over one month. He had hypertension for two years, taking lercanidipine tablets for blood pressure control, without any medical history of diabetes, pulmonary tuberculosis, or drug allergy. Positron emission tomography (PET) and computed tomography (CT) examinations revealed the presence of a lesion in the right upper lobe ( Figure 1A ) and multiple lymph nodes metastases at the right hilum of the lung, right mediastinum, the right-side clavicle, and pleural effusion in the right side. The percutaneous biopsy and immunohistochemistry (IHC) analysis revealed a poorly differentiated adenocarcinoma (ADC,  Figure 1K ) with positive expression of Napsin A, thyroid transcription factor-1 (TTF-1), and negative CK5/6. The EGFR L858R mutation was detected ( Table 1 ) by the capture-based hybrid next-generation sequencing (NGS, GeneseeqPrime ) targeting 425 cancer-related genes ( Supplementary Table S1 ). He was primarily diagnosed with a stage IV ADC of the right lung (T2N3M1a) and received gefitinib as the first-line treatment. Partial response (PR) was achieved after one month ( Figure 1B ) and progression-free survival (PFS) was 27 months for the first-line gefitinib treatment. During the treatment of gefitinib, the patient experienced grade 2 rash, which was symptomatically treated.
In December 2019, multiple metastatic lesions were found in both lungs and rapidly progressed. As shown in  Figures 1C, D , the two CT images taken with an interval of 19 days (2019.12.06 and 2019.12.25) showed over two times size increase on multiple metastatic sites. As the primary tumor remained stable, a re-biopsy of the metastatic lesion was performed. However, the pathological examinations suggested a histology change to squamous cell carcinoma (SCC,  Figure 1L ) with positive expression of CK5/6 and P63 but negative for NapsinA and TTF-1. The SCC antigen and CYFRA21-1 levels in serum were significantly increased compared to the baseline tests ( Figure 1J ). He firstly received one cycle of paclitaxel (175 mg/m2), carboplatin (AUC5) plus endostar (15 mg/m2). Then, the IHC test showed a positive PD-L1 expression with a tumor proportion score (TPS) of over 50% ( Figure 1M ). The L858R mutation of EGFR was maintained in the metastatic SCC and companied by the amplification of MET, CD274, and PDCD1LG2 ( Table 1 ). Thus, a combination of paclitaxel (175 mg/m2), carboplatin (AUC5) plus pembrolizumab (200 mg) was administrated. The follow-up examination after one cycle of combined treatment showed that the rapidly progressed metastatic lesions had been under control ( Figure 1E ). Then, partial response was achieved after an additional 5 cycles of this combined therapy and another four cycles of paclitaxel (300 mg) plus pembrolizumab (200 mg) were performed as maintained treatment. The disease remained stable until August 2020 and the patient tolerated well with current treatment.
Due to the progression of the primary tumor in August 2020, a second biopsy for the primary site was performed ( Figure 1F ), whose pathologic histology was still ADC ( Figure 1N ) with dramatic decreases in the levels of SCC antigen and CYFRA21-1 in serum ( Figure 1J ). Compared to the mutational profile of the metastatic SCC lesion, the progressed ADC maintained the oncogenic EGFR (L858R) mutation. However, the amplification of MET, CD274, and PDCD1LG2 was not detected ( Table 1 ). Whereas no genetic alterations were detected in the circulating tumor DNA by targeted NGS which suggested a very low level of tumor mutational burden. After a total of 3600cGy radiotherapy targeting the progressed primary lesion (12 times in 4 weeks), the patient continued pembrolizumab (200 mg) treatment until the March of 2021.
Multiple metastatic sites were found by repeat PET-CT including kidney, liver, and chest wall ( Figure 1G ) with a dramatic increase in carcinoembryonic antigen (CEA) level ( Figure 1J ). The metastatic chest wall lesion was pathologically identified as ADC ( Figure 1O ) with the same genetic alterations as the progressed ADC ( Table 1 ). Due to the clearance of the gene amplification of CD274 and PDCD1LG2, the IHC test of PD-L1 was negative for the chest wall lesion ( Figure 1P ). A combination treatment of pemetrexed (500 mg/m2), carboplatin (AUC5), and bevacizumab (7.5 mg/kg) led to a stable disease for five months until August 2021. With the progression on the metastatic chest wall lesion ( Figure 1H ), afatinib (40 mg/d) and anlotinib (10 mg/d) were administrated to the patient, but anolotinib was discontinued after one month because of severe side effects, including hemoptysis and tooth loss. Afatinib monotherapy relieved chest pain and led to a PR of the metastatic chest wall lesion as shown by the CT scan in December 2021 ( Figure 1I ). Until the last follow-up in January 2022, the PFS upon afatinib treatment reached five months and the treatment is ongoing.