Engineering Immune-Modulating Nanoparticles to Promote Macrophage-T cell Synergy for Cancer Immunotherapy = 항암 면역치료를 위한 대식세포-T세포 시너지 촉진형 면역 조절 나노입자 설계

국문 초록 (Abstract)

T세포와 대식세포는 항종양 면역에서 중심적인 역할을 수행하며, 상호 협력적인 작용을 통해 종양 제거 효율을 높일 수 있다. 대식세포는 식세포작용을 통해 종양세포를 직접 제거하고, 이후 종양과 관련된 항원을 제시함으로써 T세포를 활성화시킬 수 있다. 활성화된 T세포는 종양 세포를 공격할 뿐만 아니라, 종양 관련 대식세포를 항종양성 M1 표현형으로 재분극시켜 상호 강화적인 면역 순환을 형성한다. 그러나 종양세포에 발현된 PD-L1과 CD47과 같은 면역관문 단백질은 이러한 면역세포 간의 협력적 작용을 억제하여 항종양 면역 반응을 저해한다. 또한 대식세포 또는 T세포 중 하나만을 표적으로 하는 단일 면역관문억제제는 암 환자에서 제한적인 치료 효능을 보인다. 이러한 문제를 해결하기 위해, 본 연구에서는 대식세포와 T세포 모두를 대상으로 하는 이중 면역관문억제제를 종양으로 효율적으로 전달할 수 있는 나노입자를 개발하였다. 본 연구에서 제작한 PD-1 단백질 발현 대식세포막 유래 나노입자(PMMNP)는 종양세포의 CD47과 PD-L1을 각각 차단하디 위해 표면에 SIRPα와 PD-1을 함께 제시할 수 있다. PMMNP는 대식세포의 식세포 및 항원제시 능력을 향상시키고, T세포 활성을 촉진하며, 대식세포를 항종양성 표현형으로 재분극시켰다. 또한 동종 종양 동물모델에서 PMMNP는 단일 면역관문억제제를 탑재한 나노입자나 비특이적 항체 전달에 비해 우수한 항종양 치료효과를 나타냈다. 이러한 결과는 PMMNP가 T세포와 대식세포간의 항종양 면역 상호작용을 증진시키는 유망한 생체모사형 나노플랫폼으로서 차세대 항암 면역치료 전략으로 활용될 수 있음을 시사한다.

다국어 초록 (Multilingual Abstract)

T cells and macrophages function as central effectors of antitumor immunity and are capable of cooperative interactions that enhance tumor clearance. Macrophages can directly phagocytose tumor cells and subsequently process and present tumor-associated antigens to activate T cells. Once activated, T cells can destroy tumor cells and simultaneously reprogram tumor-associated macrophages toward an M1-like, antitumoral phenotype, thereby establishing a reinforcing immune circuit. However, the expression of immune checkpoint molecules such as PD-L1 and CD47 on tumor cells suppresses this reciprocal interaction between the two cell types. Although monotherapies targeting a single checkpoint pathway have shown limited success, their efficacy remains suboptimal in clinical settings. To overcome these limitations, we developed PD-1-transfected macrophage membrane-derived nanoparticles (PMMNPs) that deliver dual immune checkpoint inhibitors to tumors for both macrophages and T cells. The resulting nanoparticles are capable of homing to tumor sites and simultaneously blocking the CD47–SIRPα and PD-1–PD-L1 axes through the presentation of SIRPα and PD-1 on their surface. This dual blockade enhances macrophage phagocytic and antigen-presenting activity, promotes T cell activation, and induces the repolarization of macrophages toward a robust antitumoral phenotype. In syngeneic tumor-bearing mice, PMMNP treatment resulted in markedly improved therapeutic efficacy compared with nanoparticles carrying single inhibitors or non-targeted antibody delivery. Collectively, these findings highlight a biomimetic nanoplatform that strengthens the crosstalk between macrophages and T cells, offering a promising strategy for potentiating antitumor immunity.

목차 (Table of Contents)

• Abstract i
• Table of Contents iii
• List of Tables vii
• List of Figures viii
• Chapter 1. Research background and objective 1
• 1.1 Research Background 1
• 1.1.1. Immune system as a homeostatic regulator 1
• 1.1.2. Cancer 2
• 1.1.3. Immune checkpoint inhibitor 6
• 1.1.4. Therapeutic nanoparticles 11
• 1.1.5. Strategies for tumor targeted delivery 14
• 1.1.6. Nanoparticle-based immune checkpoint inhibitor delivery 16
• 1.2. Research Objective 18
• Chapter 2. Experimental Section 20
• 2.1. Cell culture 20
• 2.2. Mice 22
• 2.3. Preparation of PD-1-transfected macrophage membranes 23
• 2.4. Preparation of PMMNPs 25
• 2.5. Characterization of PMMNPs 26
• 2.6. Evaluation of in vivo biodistribution of PMMNPs 29
• 2.7. Assessment of in vitro and in vivo adhesion of PMMNPs on cancer cells 30
• 2.8. Evaluation of blood pharmacokinetics of PMMNPs in vivo 31
• 2.9. Evaluation of macrophage phagocytosis of cancer cells in vitro 32
• 2.10 Evaluation of T cell cytotoxicity to cancer cells in vitro 33
• 2.11. Evaluation of macrophage-mediated antigen presentation to T cells, T cell proliferation, and T cell activation in vitro 35
• 2.12. In vitro effects of nanoparticles on macrophage phenotye 36
• 2.13. In vivo anti-tumor efficacy of PMMNPs 37
• 2.14. Tumor-infiltrating lymphocyte (TIL) analysis 39
• 2.15. Statistical analyses 40
• Chapter 3. Preparation and characterization of PMMNPs 41
• 3.1. Preparation of PMMNPs 41
• 3.2. Optimization of PMMNP composition 41
• 3.3. Characterization of PMMNPs 43
• 3.4. Evaluation of in vitro cytotoxicity and in vivo toxicity of PMMNPs 49
• Chapter 4. Interactions with tumor cells and tumor-targeted delivery of PMMNPs 51
• 4.1. Interactions between tumor cells and PMMNPs 51
• 4.2. Tumor-targeted delivery of PMMNPs 55
• Chapter 5. In vitro effects of dual immune checkpoint inhibition by PMMNPs on macrophages and T cells 56
• 5.1. Enhancement of macrophage phagocytosis via SIRPα-mediated CD47 blockade by PMMNPs 56
• 5.2. Enhancement of T cell cytotoxicity via PD-1-mediated PD-L1 blockade by PMMNPs 61
• 5.3. The effects of PMMNPs on macrophage-mediated antigen presentation to T cells, subsequent T cell activation, and macrophage phenotype 64
• Chapter 6. In vivo antitumoral efficacy of PMMNPs with immunomodulation of tumor microenvironment 69
• 6.1. In vivo antitumoral efficacy of PMMNPs 69
• 6.2. Reprogramming of the tumor microenvironment 75
• Chapter 7. Discussion and conclusion 76
• 7.1. Discussion 76
• 7.2. Future direction 86
• 7.3. Conclusion 89
• References 90
• 국문초록 (Abstract in Korean) 99