Background: Cancer stem cells (CSCs) are a subpopulation of cancer cells known for their self-renewal capacity, tumorigenicity, and resistance to treatment. Toll-like receptor 3 (TLR3) plays a complex role in cancer, exhibiting both pro-apoptotic and pro-tumorigenic effects. This study investigates the pro-tumorigenic role of TLR3, specifically its impact on CSCs in head and neck cancer. Methods: We have investigated Detroit 562, FaDu and SQ20B cell lines, the latter being stably transfected with a plasmid containing inducible shRNA for TLR3, by cultivating them to form tumor spheres in order to study CSCs. Results: Our findings demonstrate that TLR3 activation promotes stemness in head and neck cancer cell lines. This is evidenced by increased tumor sphere formation, promotion of epithelial-to-mesenchymal transition (EMT), upregulated stemness gene expression, and elevated aldehyde dehydrogenase (ALDH) activity. Conditional TLR3 knockdown abolished tumor sphere formation, confirming its important role. Furthermore, TLR3 activation triggers the secretion of damage-associated molecular patterns (DAMPs) into the tumor microenvironment, leading to increased cancer cell migration. This was inhibited by DAMP inhibitors. In patient tissue samples, we observed co-localization of TLR3 with stemness markers CD133 and ALDH1, as well as with heat shock protein 70 (HSP70) and receptor for advanced glycation end products (RAGE). We then explored potential CSC-targeted therapies, initially combining the apoptosis inducer poly (I:C) with DAMP inhibitors and γ-irradiation. While this combination proved effective in adherent cells, it failed to eliminate tumor spheres. Nevertheless, we discovered that proton radiotherapy, particularly when combined with aspirin (HMGB1 inhibitor) and poly (I:C), effectively eliminates CSCs. Conclusions: This novel combination holds promise for the development of new therapeutic strategies for head and neck cancers, particularly given the promising results of proton therapy in treating this disease.
背景：癌症干细胞（CSCs）是具有自我更新能力、致瘤性和治疗抵抗性的癌细胞亚群。Toll样受体3（TLR3）在癌症中扮演复杂角色，既能促进细胞凋亡，也能增强肿瘤发生。本研究探讨TLR3的促肿瘤作用，特别是其对头颈癌中CSCs的影响。方法：通过培养Detroit 562、FaDu及SQ20B细胞系（后者稳定转染含TLR3诱导型shRNA的质粒）形成肿瘤球以研究CSCs。结果：研究发现TLR3激活能促进头颈癌细胞系的干性特征，具体表现为肿瘤球形成增加、上皮-间质转化（EMT）增强、干性基因表达上调以及乙醛脱氢酶（ALDH）活性升高。条件性敲低TLR3可完全抑制肿瘤球形成，证实其关键作用。此外，TLR3激活会触发损伤相关分子模式（DAMPs）向肿瘤微环境分泌，从而增强癌细胞迁移，该现象可被DAMP抑制剂阻断。在患者组织样本中，我们观察到TLR3与干性标志物CD133、ALDH1以及热休克蛋白70（HSP70）、晚期糖基化终末产物受体（RAGE）存在共定位。随后我们探索了靶向CSCs的潜在疗法，初步将凋亡诱导剂聚肌胞苷酸（poly (I:C)）与DAMP抑制剂及γ射线照射联用。该组合虽对贴壁细胞有效，但未能清除肿瘤球。然而，我们发现质子放疗（特别是与阿司匹林（HMGB1抑制剂）及poly (I:C)联用时）能有效清除CSCs。结论：这种创新性联合疗法为头颈癌新治疗策略的开发带来希望，尤其考虑到质子疗法在该疾病治疗中已展现的良好前景。
肿瘤(癌症)患者之家