Combining microenvironment normalization strategies to improve cancer immunotherapy

Abstract

Advances in immunotherapy have revolutionized the treatment of multiple cancers. Unfortunately, tumors usually have impaired blood perfusion, which limits the delivery of therapeutics and cytotoxic immune cells to tumors and also results in hypoxia-a hallmark of the abnormal tumor microenvironment (TME)-that causes immunosuppression. We proposed that normalization of TME using antiangiogenic drugs and/or mechanotherapeutics can overcome these challenges. Recently, immunotherapy with checkpoint blockers was shown to effectively induce vascular normalization in some types of cancer. Although these therapeutic approaches have been used in combination in preclinical and clinical studies, their combined effects on TME are not fully understood. To identify strategies for improved immunotherapy, we have developed a mathematical framework that incorporates complex interactions among various types of cancer cells, immune cells, stroma, angiogenic molecules, and the vasculature. Model predictions were compared with the data from five previously reported experimental studies. We found that low doses of antiangiogenic treatment improve immunotherapy when the two treatments are administered sequentially, but that high doses are less efficacious because of excessive vessel pruning and hypoxia. Stroma normalization can further increase the efficacy of immunotherapy, and the benefit is additive when combined with vascular normalization. We conclude that vessel functionality dictates the efficacy of immunotherapy, and thus increased tumor perfusion should be investigated as a predictive biomarker of response to immunotherapy.

Keywords: anti-angiogenic therapy; immunotherapy; mechanotherapeutics; normalization; vascular function.

Fig. 1.

Schematic of the interactions among model components. The model accounts for various cell populations (orange boxes) and tumor angiogenic factors (blue boxes). TME component: Increases in functional vascular density and tumor perfusion enhance tumor oxygenation. Higher oxygen levels accelerate the proliferation rates of CCs and CD4⁺ T cells and the activity of immune cells and polarize TAMs from an immune inhibitory M2-like phenotype toward an immune stimulatory M1-like phenotype. Along with the immunostimulatory action of M1-like TAMs, the model accounts for their tumoricidal effect on CCs. According to previous studies, CD4⁺ T cells stimulate CD8⁺ T cells, and an increased immune response leads to more efficient killing of all types of cancer cells. Increased proliferation of all cancer cell types results in increased oxygen consumption, inactivation of immune cells, and decreased vessel diameters due to compression-induced hypoxia. Hypoxia favors proliferation of CSCs and ICCs and increases VEGF and CXCL12 levels. In addition, targeting of stroma components through CXCL12/CXCR4 signaling alleviates solid stresses, which are associated with vascular dysfunction. Tumor vasculature component: This initiates angiogenesis through the proliferation and migration of endothelial cells that form the vessels. Angiogenesis is enhanced by high levels of Ang2, which destabilizes vessels, and inhibited by Ang1 and PDGF-B, which recruit pericytes and stabilize vessels. In addition, knockout of CD4⁺ T cells results in overexpression of VEGF, which is correlated with higher numbers of M2-like TAMs. On the other hand, a decrease in M2-like TAMs results in higher numbers of effector immune cells (CD8⁺ T cells and NK cells). Increased numbers of CD4⁺ and CD8⁺ T cells enhances production rates of IFNγ, which is associated with decreased vessel wall pore size and permeability, leading to vascular normalization. Vascular normalization improves the functionality of the vascular network, leading to an increase in functional vascular density, which enhances cancer cell proliferation.

Fig. 2.

Comparison of model predictions with experimental data from Chauhan et al. (26) for E0771 (A) and MCa-M3C (B) breast tumors. (C) Comparison of model predictions with experimental data from Chen et al. (24) in MCa-M3C breast tumors. The x-axis shows the various treatment groups included in the experimental studies: TMA-ARB, tumor-selective angiotensin receptor blocker; anti–PD-1, PD-1- blocker; anti–CTLA-4, CTLA4- blocker; and AMD3100, a CXCR4 inhibitor.

Fig. 3.

Comparison of model predictions with experimental data reported by Huang et al. (36) (A), Zheng et al. (19) (B), and Shigeta et al. (49) (C). The x-axis shows the various treatment groups included in the experimental studies: DC101, an anti-VEGF antibody; anti–PD-1, a PD-1 blocker; and anti–CTLA-4, a CTLA-4 blocker.

Fig. 4.

Effect of different doses of anti-VEGF treatment combined with different values of the source term of CD8⁺ T cells to model immunotherapy for sequential administration. Shown are phase diagrams for the effect of combinatorial treatment on functional vascular density (A), tumor oxygenation (B), VEGF level (C), CD4⁺ T cells (D), effector immune cells (NK and CD8⁺ T cells) (E) , M1-like (F) and M2-like (G) TAMs, cancer cell population (H), and tumor volume (I). Values of model parameters presented in the figure were calculated at the location equidistant from the tumor center and periphery. On the x-axis, a value of 1 corresponds to the baseline value of source term of CD8⁺ T cells (SI Appendix, Table S1).

Fig. 5.

Temporal distribution of the values of model parameters: hypoxia fraction (A), ratio of CD8⁺ T cells to Tregs (B), ratio of M1-like to M2-like TAMs (C), cancer cell population (D), solid stress (E) and functional vascular density (F), calculated at the center of the tumor for untreated tumors and tumors receiving immunotherapy alone or combined with a normalization treatment.

Fig. 6.

Spatial distribution of the values of model parameters: solid stress (A), functionalvascular density (B), hypoxia fraction (C), ratio of CD8⁺ T cells to Tregs (D), ratio of M1-like to M2-like TAMs (E), cancer cell population (F) for untreated tumors and tumors receiving immunotherapy alone or combined with a normalization treatment.

Fig. 7.

Schematic of proposed mechanism of action of normalization strategies to improve immunotherapy. (Adapted with permission from ref. .) Combined administration of immunotherapy with stroma and/or vascular normalization restores vascular functionality and alleviates solid stress, leading to improved tumor perfusion and oxygenation, skewing TAM polarization away from the immunosuppressive M2-like phenotype to the M1-like phenotype, stimulating immunity (e.g., CD4⁺ and CD8⁺ T cells) and thus increasing cancer cell killing.