DataSheet1_Dihydroartemisinin remodels macrophage into an M1 phenotype via ferroptosis-mediated DNA damage.docx

Lung cancer recruits tumor-associated macrophages (TAMs) massively, whose predominantly pro-tumor M2 phenotype leads to immunosuppression. Dihydroartemisinin (DHA) has been proven to remodel TAM into an anti-tumor M1 phenotype at certain concentrations in the present study, which was hypothesized to facilitate anti-lung cancer immunotherapy. However, how DHA remodels the TAM phenotype has not yet been uncovered. Our previous work revealed that DHA could trigger ferroptosis in lung cancer cells, which may also be observed in TAM thereupon. Sequentially, in the current study, DHA was found to remodel TAM into the M1 phenotype in vitro and in vivo. Simultaneously, DHA was observed to trigger ferroptosis in TAM and cause the DNA damage response and NF-κB activation. Conversely, the DHA-induced DNA damage response and NF-κB activation in TAM were attenuated after the inhibition of ferroptosis in TAM using an inhibitor of ferroptosis. Importantly, a ferroptosis inhibitor could also abolish the DHA-induced phenotypic remodeling of TAM toward the M1 phenotype. In a nutshell, this work demonstrates that DHA-triggered ferroptosis of TAM results in DNA damage, which could activate downstream NF-κB to remodel TAM into an M1 phenotype, providing a novel strategy for anti-lung cancer immunotherapy. This study offers a novel strategy and theoretical basis for the use of traditional Chinese medicine monomers to regulate the anti-tumor immune response, as well as a new therapeutic target for TAM phenotype remodeling.

肺癌可大量招募肿瘤相关巨噬细胞（tumor-associated macrophages, TAMs），其以促瘤的M2表型为主，进而引发免疫抑制。本研究证实，双氢青蒿素（Dihydroartemisinin, DHA）在特定浓度下可将TAM重极化为抗瘤的M1表型，据此推测其可助力抗肺癌免疫治疗。然而，DHA重塑TAM表型的具体机制尚未阐明。我们此前的研究发现，DHA可诱导肺癌细胞发生铁死亡（ferroptosis），该现象同样可能在TAM中出现。随后，本研究通过实验证实，DHA在体外及体内均可将TAM重极化为M1表型。同时，本研究观察到DHA可诱导TAM发生铁死亡，并引发DNA损伤应答（DNA damage response）与核因子κB（NF-κB）活化。反之，若使用铁死亡抑制剂抑制TAM中的铁死亡过程，则DHA诱导的TAM DNA损伤应答与NF-κB活化均会被削弱。值得注意的是，铁死亡抑制剂同样可阻断DHA诱导的TAM向M1表型的重塑。简言之，本研究证实，DHA诱导的TAM铁死亡可引发DNA损伤，进而激活下游NF-κB通路，最终将TAM重极化为M1表型，为抗肺癌免疫治疗提供了全新策略。本研究为利用中药单体调控抗瘤免疫应答提供了新思路与理论基础，同时也为TAM表型重塑提供了全新的治疗靶点。