缺氧是塑造免疫环境的主要因素，已经开发了多种癌症模型来模拟缺氧肿瘤，但迄今为止，它们仍存在一些局限性，如缺乏可重复性、生物物理学刺激不足、免疫细胞浸润有限和氧气（O2）控制差，导致非病理生理的肿瘤反应。因此，开发更好的癌症模型非常重要，这些模型可以模拟肿瘤细胞外基质的关键特征，重建肿瘤相关的缺氧环境，同时允许细胞浸润和癌症-免疫细胞相互作用。在这里，我们使用透明质酸（HA）工程化缺氧诱导冷冻凝胶（HICs），制造三维微组织并模拟缺氧肿瘤微环境。具体来说，设计了肿瘤细胞负载的HICs，以局部耗尽氧气并诱导持久缺氧。HICs促进了缺氧响应基因表达和表型的改变，代谢适应无氧糖酵解，以及化疗抗药性。此外，HIC支持的肿瘤模型抑制了树突状细胞（DC），显示出浆细胞样DC（pDC）亚群表型变化和缺氧条件下常规DC（cDC）反应的受损。最后，基于HIC的黑色素瘤模型导致了CD8+T细胞抑制，与促炎细胞因子分泌下调、免疫调节因子表达增加以及T细胞脱颗粒和细胞毒性能力下降相关。总体而言，这些数据表明HICs可以用作模拟类似实体的肿瘤微环境的工具，并具有在低氧条件下加深我们对癌症免疫细胞关系的理解以及开发更有效疗法的潜力。© 2023 The Authors.

Hypoxia is a major factor shaping the immune landscape, and several cancer models have been developed to emulate hypoxic tumors. However, to date, they still have several limitations, such as the lack of reproducibility, inadequate biophysical cues, limited immune cell infiltration, and poor oxygen (O2) control, leading to non-pathophysiological tumor responses. Therefore, it is essential to develop better cancer models that mimic key features of the tumor extracellular matrix and recreate tumor-associated hypoxia while allowing cell infiltration and cancer-immune cell interactions. Herein, hypoxia-inducing cryogels (HICs) have been engineered using hyaluronic acid (HA) to fabricate three-dimensional microtissues and model a hypoxic tumor microenvironment. Specifically, tumor cell-laden HICs have been designed to deplete O2 locally and induce long-standing hypoxia. HICs promoted changes in hypoxia-responsive gene expression and phenotype, a metabolic adaptation to anaerobic glycolysis, and chemotherapy resistance. Additionally, HIC-supported tumor models induced dendritic cell (DC) inhibition, revealing a phenotypic change in the plasmacytoid DC (pDC) subset and an impaired conventional DC (cDC) response in hypoxia. Lastly, our HIC-based melanoma model induced CD8+ T cell inhibition, a condition associated with the downregulation of pro-inflammatory cytokine secretion, increased expression of immunomodulatory factors, and decreased degranulation and cytotoxic capacity of T cells. Overall, these data suggest that HICs can be used as a tool to model solid-like tumor microenvironments and has great potential to deepen our understanding of cancer-immune cell relationship in low O2 conditions and may pave the way for developing more effective therapies.© 2023 The Authors.