― 191 ―attempted to expand lymphocytes in medium containing IL-2 for a short period of time. These patient-derived cells were analyzed at the single cell level using multicolor flow cytometry or single cell sequencing. In addition, using a model in which patient-derived cancer cells were transplanted into immunodeficient mice (EML4-ALK-positive lung cancer xenograft model), we analyzed residual tumors and tumors that regrown despite continued treatment. In addition, a genome-wide CRISPR screen was performed using independently established ALK fusion-positive lung cancer cells to identify factors involved in treatment resistance. From the CRISPR screen, in addition to previously reported genes such as BAX or NF2, we newly identified that knockout of ERRFI1 significantly increased the number of residual cells after ALK inhibitor treatment (drug-tolerant persister cells; DTPs). Detailed analysis of the mechanism by which ERRFI1 loss increases DTPs revealed that deletion or downregulation of ERRFI1 enables cells to respond to very low concentrations of EGF ligand, activates EGFR, and allows cells to survive in the presence of ALK inhibitors, and that this resistance can be overcome by the combination of anti-EGFR antibodies. In addition, using an in vivo model of ALK-positive lung cancer, we found that certain immune cells appear to secrete a ligand for Axl, the expression of which is induced by EMT induction and other mechanisms, leading to therapeutic resistance. These immune cells appeared to accumulate in the tumor microenvironment after ALK-TKI treatment. On the other hand, using the syngeneic mouse model, we have established to create immunological memory, which is established complete remission (CR) of tumor after immune checkpoint inhibitor therapy or surgical removal of residual tumor cells after the short period of treatment. Detailed analysis of the immunological memory model, mainly from the T cell repertoire analysis, specific TCR sequence positive CD8 T cell clones expanded and were resident in the body as memory T cells. These specific TCR repertoire harboring cells were also observed in the metastasized colorectal tumor tissues surgically resected in the different timing and drug treatments. Similar analysis has also been performed using the syngeneic mouse lung cancer model with mouse EGFR-del19 mutation and untreated and kinase inhibitor treated pleural effusion cells from EGFR-positive lung cancer patients. To date, many molecularly targeted anticancer drugs and multiple anticancer immunotherapies have been developed and have shown high therapeutic efficacy, but have not yet been able to cure cancer. In many cases, the tumor has relapsed even though they have shown sustained and marked therapeutic efficacy for several years. We believe that this research partially addresses this issue and finds a combined treatment strategy that has the potential to completely eliminate residual tumor resistance, but further studies are needed to clarify the characteristics of treatment-resistant tumor and identify the therapeutic target to achieve complete cure.はじめに肺がんの各種薬物療法は目覚ましい進歩を遂げてきたものの，獲得耐性が今なお大きな問題として立ちはだかり，今なお，がんを薬物療法によって完治を達成するのは困難である．薬剤耐性機構の研究と耐性克服法の探索研究が世界中で進められてきたが，新たな耐性克服法に対してさえがんは耐性を獲得してしまうことが問題である．近年は，耐性がんの芽となる治療残存腫瘍の微小環境を理解し，耐性を生みにくいまたは耐性出現を遅らせることのできるような治療標的の探索研究が行われつつある．本研究では，治療残存腫瘍を，がん細胞だけでなく，間質細胞や免疫細胞にも着目して研究・解
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