Institute of Biomedicine, University of Turku, Turku, Finland.
School of Pharmacy, University of Eastern Finland, Kuopio, Finland.
Eur J Pharm Sci. 2023 Nov 1;190:106571. doi: 10.1016/j.ejps.2023.106571. Epub 2023 Aug 29.
Tumour associated macrophages (TAMs) are important players in breast tumour progression and metastasis. Clinical and preclinical evidence suggests a role for zoledronate (ZOL) in breast cancer metastasis prevention. Further, zoledronate is able to induce inflammatory activation of monocytes and macrophages, which can be favourable in cancer treatments. The inherent bone tropism of zoledronate limits its availability in soft tissues and tumours. In this study we utilised an orthotopic murine breast cancer model to evaluate the possibility to use liposomes (EMP-LIP) to target zoledronate to tumours to modify TAM activation.
Triple-negative breast cancer 4T1 cells were inoculated in the 4th mammary fat pad of female Balb/c mice. Animals were divided according to the treatment: vehicle, ZOL, EMP-LIP and liposome encapsulated zoledronate (ZOL-LIP). Treatment was done intravenously (with tumour resection) and intraperitoneally (without tumour resection). Tumour growth was followed by bioluminescence in vivo imaging (IVIS) and calliper measurements. Tumour-infiltrating macrophages were assessed by immunohistochemical and immunofluorescence staining. Protein and RNA expression levels of inflammatory transcription factors and cytokines were measured by Western Blotting and Taqman RT-qPCR.
Liposome encapsulated zoledronate (ZOL-LIP) treatment suppressed migration of 4T1 cell in vitro. Tumour growth and expression of the angiogenic marker CD34 were reduced upon both ZOL and ZOL-LIP treatment in vivo. Long-term ZOL-LIP treatment resulted in shift towards M1-type macrophage polarization, increased CD4 T cell infiltration and activation of NF-κB indicating changes in intratumoural inflammation, whereas ZOL treatment showed similar but non-significant trends. Moreover, ZOL-LIP had a lower bisphosphonate accumulation in bone compared to free ZOL.
Results show that the decreased bisphosphonate accumulation in bone promotes the systemic anti-tumour effect of ZOL-LIP by increasing inflammatory response in TNBC tumours via M1-type macrophage activation.
肿瘤相关巨噬细胞（TAMs）是乳腺癌肿瘤进展和转移的重要参与者。临床前和临床证据表明唑来膦酸（ZOL）在预防乳腺癌转移方面具有作用。此外，唑来膦酸能够诱导单核细胞和巨噬细胞的炎症激活，这在癌症治疗中可能是有利的。唑来膦酸固有的骨亲嗜性限制了其在软组织和肿瘤中的可用性。在这项研究中，我们利用了一种原位乳腺癌小鼠模型，来评估使用脂质体（EMP-LIP）将唑来膦酸靶向肿瘤以改变 TAM 激活的可能性。
将三阴性乳腺癌 4T1 细胞接种于雌性 Balb/c 小鼠的第 4 对乳腺脂肪垫中。根据治疗方法将动物分为以下几组：载体、ZOL、EMP-LIP 和包封唑来膦酸的脂质体（ZOL-LIP）。治疗通过静脉内（带肿瘤切除）和腹腔内（不带肿瘤切除）途径进行。通过体内生物发光成像（IVIS）和卡尺测量来监测肿瘤生长。通过免疫组织化学和免疫荧光染色评估肿瘤浸润巨噬细胞。通过 Western Blotting 和 Taqman RT-qPCR 测量炎症转录因子和细胞因子的蛋白和 RNA 表达水平。
脂质体包封的唑来膦酸（ZOL-LIP）处理抑制了 4T1 细胞的体外迁移。体内 ZOL 和 ZOL-LIP 处理均降低了肿瘤生长和血管生成标志物 CD34 的表达。长期 ZOL-LIP 处理导致 M1 型巨噬细胞极化的转变，增加了 CD4 T 细胞浸润和 NF-κB 的激活，表明肿瘤内炎症发生了变化，而 ZOL 处理则显示出类似但无统计学意义的趋势。此外，ZOL-LIP 在骨骼中的双膦酸盐积累低于游离 ZOL。
结果表明，ZOL-LIP 在骨骼中的双膦酸盐积累减少通过 M1 型巨噬细胞激活增加 TNBC 肿瘤中的炎症反应，从而促进了 ZOL-LIP 的全身性抗肿瘤作用。
