Pericyte phenotype switching alleviates immunosuppression and sensitizes vascularized tumors to immunotherapy in preclinical models
Zhi-Jie Li, Bo He, Alice Domenichini, Jiulia Satiaputra, Kira H. Wood, Devina D. Lakhiani, Abate A. Bashaw, Lisa M. Nilsson, Ji Li, Edward R. Bastow, Anna Johansson-Percival, Elena Denisenko, Alistair R.R. Forrest, Suraj Sakaram, Rafael Carretero, Günter J. Hämmerling, Jonas A. Nilsson, Gabriel Y.F. Lee, Ruth Ganss
Zhi-Jie Li, Bo He, Alice Domenichini, Jiulia Satiaputra, Kira H. Wood, Devina D. Lakhiani, Abate A. Bashaw, Lisa M. Nilsson, Ji Li, Edward R. Bastow, Anna Johansson-Percival, Elena Denisenko, Alistair R.R. Forrest, Suraj Sakaram, Rafael Carretero, Günter J. Hämmerling, Jonas A. Nilsson, Gabriel Y.F. Lee, Ruth Ganss
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Research Article Oncology

Pericyte phenotype switching alleviates immunosuppression and sensitizes vascularized tumors to immunotherapy in preclinical models

Abstract

T cell–based immunotherapies are a promising therapeutic approach for multiple malignancies, but their efficacy is limited by tumor hypoxia arising from dysfunctional blood vessels. Here, we report that cell-intrinsic properties of a single vascular component, namely the pericyte, contribute to the control of tumor oxygenation, macrophage polarization, vessel inflammation, and T cell infiltration. Switching pericyte phenotype from a synthetic to a differentiated state reverses immune suppression and sensitizes tumors to adoptive T cell therapy, leading to regression of melanoma in mice. In melanoma patients, improved survival is correlated with enhanced pericyte maturity. Importantly, pericyte plasticity is regulated by signaling pathways converging on Rho kinase activity, with pericyte maturity being inducible by selective low-dose therapeutics that suppress pericyte MEK, AKT, or notch signaling. We also show that low-dose targeted anticancer therapy can durably change the tumor microenvironment without inducing adaptive resistance, creating a highly translatable pathway for redosing anticancer targeted therapies in combination with immunotherapy to improve outcome.

Authors

Zhi-Jie Li, Bo He, Alice Domenichini, Jiulia Satiaputra, Kira H. Wood, Devina D. Lakhiani, Abate A. Bashaw, Lisa M. Nilsson, Ji Li, Edward R. Bastow, Anna Johansson-Percival, Elena Denisenko, Alistair R.R. Forrest, Suraj Sakaram, Rafael Carretero, Günter J. Hämmerling, Jonas A. Nilsson, Gabriel Y.F. Lee, Ruth Ganss

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Figure 1

RGS5 expression levels regulate pericyte phenotype in vitro.

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RGS5 expression levels regulate pericyte phenotype in vitro. (A) Relativ...
(A) Relative RGS5 mRNA expression in 10T1/2, RGS5myc overexpressing and Rgs5 knockdown cell lines (RGS5shRNA1/3). n = 3 replica. Data are represented as mean ± SEM. *P < 0.01; **P < 0.001; ***P < 0.0001, 1-way ANOVA. (B) Microscopic images depicting 10T1/2, RGS5 overexpressing, and Rgs5 knockdown cells. Scale bar: 50 μm. (C) Cell proliferation (48 hours) in parental and transfectant 10T1/2 cells. n = 5 replica. Data are represented as mean ± SEM. ***P < 0.0001, 1-way ANOVA. (D) FACS blots showing PI histograms with color-coded cell cycle phases. Green, G0/G1 phase; yellow, S phase; blue, G2/M phase. Quantitative analysis of cell-cycle progression. n = 3 experiments. Data are represented as mean ± SEM. *P = 0.02; **P = 0.001; ***P = 0.0001, 1-way ANOVA. (E) Western blot (WB) of phosphorylated/total FOXO3a and phosphorylated/total p27KIP proteins. Red box highlights results in RGS5 overexpression cells. Duplicates are shown for each marker. Relative phosphorylated and total FOXO3a expression were normalized to tubulin; relative phosphorylated p27KIP was normalized to total p27KIP expression and quantified. n = 3 experiments (2 replica each). Data are represented as mean ± SEM. tFOXOa: ****P < 0.0001; pFOXO3a: *P = 0.028; p-p27: *P = 0.020, **P = 0.006, 1-way ANOVA. (F) WB of contractile (CNN1, ACTG2) and synthetic (KLF4, CNX43) markers in correlation to Rho kinase activity (p-MLC). Red box highlights results in RGS5 overexpression cells. Duplicates are shown for each marker, and relative protein expression normalized to tubulin was quantified. n = 3 experiments (2 replica each). Data are represented as mean ± SEM. CNN1: ****P < 0.0001, **P = 0.011; ACTG2: ***P = 0.0001, **P = 0.012, *P = 0.047; KLF4: **P = 0.003; CNX43: ***P = 0.003; p-MLC: ****P < 0.0001, *P = 0.017, 1-way ANOVA. (G) WB of ROCK1 and ROCK2 proteins. Duplicates are shown for each marker. Relative protein expression normalized to tubulin was quantified. n = 3 experiments (2 replica each). Data are represented as mean ± SEM. **P = 0.005, Student’s t test. (H) Contractile and synthetic pericyte markers in relation to RGS5 high (Rgs5hi) or low (Rgs5low) expression.