Towards Immunotherapy-Induced Normalization of the Tumor Microenvironment

Abstract

Immunotherapies modulate the function of immune cells to eradicate cancer cells through various mechanisms. These therapies are successful across a spectrum of cancers, but they are curative only in a subset of patients. Indeed, a major obstacle to the success of immunotherapies is the immunosuppressive nature of the tumor microenvironment (TME), comprising the stromal component and immune infiltrate of tumors. Importantly, the TME in most solid cancers is characterized by sparsely perfused blood vessels resulting from so-called pathological angiogenesis. In brief, dysregulated development of new vessels results in leaky tumor blood vessels that inefficiently deliver oxygen and other nutrients. Moreover, the occurrence of dysregulated fibrosis around the lesion, known as pathological desmoplasia, further compresses tumor blood vessels and impairs blood flow. TME normalization is a clinically tested treatment strategy to reverse these tumor blood vessel abnormalities resulting in stimulated antitumor immunity and enhanced immunotherapy efficacy. TME normalization includes vascular normalization to reduce vessel leakiness and reprogramming of cancer-associated fibroblast to decompress vessels. How immunotherapies themselves normalize the TME is poorly understood. In this review, we summarize current concepts and progress in TME normalization. Then, we review observations of immunotherapy-induced TME normalization and discuss the considerations for combining vascular normalizing and immunotherapies. If TME could be more completely normalized, immunotherapies could be more effective in more patients.

Keywords: angiogenesis; hypoxia; immune cell infiltrate; immune checkpoints; immunotherapy; tumor microenvironment; vascular normalization.

FIGURE 1

Cancer cells contribute to angiogenesis, desmoplasia, and immunosuppression thereby reducing blood flow and immune cell infiltration. (A) A simplified schematic of leaky tumor blood vessels and relevant cells. Cancer cells secrete angiogenic factors, including vascular endothelial growth factor (VEGF), that reduce the expression of molecules, such as intercellular adhesion molecule 1 (ICAM-1), which facilitate the interaction of endothelial cells with themselves and perivascular cells in normally functioning vessels. Plasma and blood-borne molecules (arrows) flow out of pores in the vessel wall thereby reducing flow. (B) A simplified schematic of compressed tumor blood vessels and relevant cells. Cancer cells activate cancer-associated fibroblasts through various signals including transforming growth factor (TGF) β. As a result, more fibrosis is produced and maintained thereby transferring compressive physical force onto blood vessels. Blood flow is reduced. (C) A simplified schematic of immunosuppression and relevant cells. As in (A) and (B), cancer cells secrete VEGF and TGFβ among other factors that affect angiogenesis and fibrosis. VEGF recruits regulatory T cells and shifts tumor-associated macrophages (TAMs) towards M2-like immunosuppressive phenotypes. VEGF also blocks the recruitment and transmigration of CD8⁺ T cells. TGFβ signaling leads to increased fibrosis that physically impedes the migration of CD8⁺ T cells to cancer cells and blocks the vascular transmigration of these cells through endothelin one signaling through the endothelin receptor type (B)

FIGURE 2

Vascular normalization and reprogramming of cancer-associated fibroblasts shift the microenvironment towards antitumor immunity. (A) A schematic of a magnified, cross-sectional view of a single blood vessel in an untreated tumor. A pinch point in the blood vessel (red tube) and the lack of a consistent endothelial cell layer fortified with pericytes restricts blood flow (gray arrows). The tumor is replete with immunosuppressive cancer-associated fibroblasts (CAFs), fibrosis, and regulatory T cells (CD4⁺CD25⁺FOXP3⁺) while lacking CD8⁺ and other subsets of CD4⁺ T cells. (B) A schematic of a magnified, cross-sectional view of a single blood vessel in a tumor treated with vascular normalizing therapy. As the balance of pro- and anti-angiogenic factors shifts towards the latter, endothelial cells are aligned, and blood vessels are fortified with pericytes yet remain compressed by mechanical stress. Perfusion increases especially in tumors with low levels of mechanical stress. Immune cells such as CD8⁺ T cells more efficiently traffic to tumors and transmigrate across the vessel wall. There are fewer immunosuppressive cells because of reduced angiogenic and hypoxia signaling. Vascular normalization by immune checkpoint inhibitors could rely on the accumulation of activated eosinophils. (C) A schematic of a magnified, cross-sectional view of a single blood vessel in tumor treated with CAF reprogramming therapy. As CAFs shift to quiescent fibroblasts, they produce and maintain lower levels of fibrosis. Mechanical stress is alleviated and vessels are decompressed. Perfusion increases. Immune cells such as CD8⁺ T cells flow through tumors and migrate the interstitial space because of less immunosuppressive CAF and hypoxia signaling. Also, there is less physical restriction of migration by components of fibrosis, such as collagen. (D) A schematic of a magnified, cross-sectional view of a single blood vessel in tumor treated with both vascular normalizing and CAF reprogramming therapy. Given the reduced signaling and physical barriers, immune cells such as CD8⁺ T cells efficiently traffic to tumors, negotiate transport through the vessel wall, and penetrate to clusters of cancer cells.

FIGURE 3

Vascular normalization through immunotherapies. Immunotherapies stimulate various immune cells to act on endothelial and mural cells resulting in vascular normalization. Several immunotherapies normalize vessels through distinct mechanisms. (A) The effects of immune checkpoint inhibitors (ICIs) are the most well-studied. ICIs activate CD8⁺ T cells, which secrete IFNγ. These cells can interact with activated eosinophils through IFNγ to induce M1-like TAM phenotypes, which reduces VEGF signaling and induces VCAM-1 expression. As a result, more CD8⁺ T cells and activated eosinophils adhere to and transmigrate across the endothelium. The later secretes chemokines (i.e., CCL5, CXCL9 and CXCL10), which increase trafficking of CD8⁺ T cells and eosinophils to tumors. For this reason, activated eosinophil accumulation precedes and is required for increased CD8⁺ T cell homing to tumors. This process is a potential feedback loop of vascular normalization and antitumor immunity. DLL1-Notch signaling promotes CD8⁺ T cell activation and IFNγ production thereby reinforcing this positive feedback loop for long-term vascular normalization. (B) STING agonists cause an increase in antiangiogenic factors, which results in increased pericyte coverage through VCAM-1 expression, which also facilitates the infiltration of T cells. (C) CpG-ODN directly act on TAMs promoting an M1-like phenotype, which induces the upregulation of ICAM-1 and VCAM-1 expression. In some contexts, depletion of regulatory T cells has similar effects on ICAM-1 and VCAM-1 expression. (D) Oncolytic vaccines reduce vascular density and increase VCAM-1 expression through unelucidated mechanisms.

FIGURE 4

Vascular fortification by immune checkpoint inhibition promotes vascular normalization by subsequent antiangiogenic therapy in hepatocellular carcinoma. (A) Immune checkpoint inhibition monotherapy normalizes tumor blood vessels in various murine models of cancer by increasing the interaction between endothelial cells and with perivascular cells in a CD8⁺ T cell dependent manner. (B) Following immune checkpoint inhibition with antiangiogenic therapy, such as the small-molecule tyrosine kinase inhibitor sorafenib, increases the density of tumor blood vessels fortified by pericytes and antitumor efficacy in a CD8⁺ T cell dependent manner. In contrast, sorafenib monotherapy induces vascular regression and does not induce an antitumor effect. Thus, vascular-fortifying ICI can shift the effect of subsequent antiangiogenic therapy from vessel destruction to normalization.