Angiotensin-converting enzyme 2 identifies immuno-hot tumors suggesting angiotensin-(1–7) as a sensitizer for chemotherapy and immunotherapy in breast cancer

Fig. 1. ACE2 predicts an inflamed TME in BC. A Expression levels of 122 immunomodulators between the high and low ACE2 groups in BC. B Differences in the levels of TIICs calculated using five algorithms between the high and low ACE2 groups. C Differences in Tumor Purity, ESTIMATE Score, Immune Score, and Stromal Score estimated by the ESTIMATE method between the high and low ACE2 groups. Significance was calculated with Student’s t-test. *P < 0.05, ****P < 0.0001. D Differences in the gene markers of the common TIICs between the high and low ACE2 groups. E Differences in the various steps of the cancer immunity cycle between the high and low ACE2 groups. Significance was calculated with Student’s t-test. ns-P > 0.05, *P < 0.05, ****P < 0.0001. F Correlation between ACE2 and common inhibitory immune checkpoints. Significance was calculated with Pearson correlation analysis. The color reveals the Pearson correlation coefficient

Fig. 2. Correlation between ACE2 and the immune phenotype in BC. A Differences in expression levels of immune-related targets the high and low ACE2 groups in BC. Significance was calculated with Student’s t-test. ****P < 0.0001. B Differences in T cell inflamed scores between the high and low ACE2 groups. The T cell inflamed score is positively related to the response to cancer immunotherapy. Significance was calculated with Student’s t-test. ****P < 0.0001. C, D Mutational landscape in the high and low ACE2 groups. E Differences in TMB levels between the high and low ACE2 groups. Significance was calculated with Student’s t-test. ****P < 0.0001. F Differences in deficiency rates of MMR proteins between the high and low ACE2 groups. Significance was calculated with Pearson’s Chi-squared test. G Differences in IPS scores between the high and low ACE2 groups. Significance was calculated with Student’s t-test. ****P < 0.0001

Fig. 3. ACE2 predicts the therapeutic options in BC. A Correlations between ACE2 and clinic-pathological features in BC. Significance was calculated with Pearson’s Chi-squared test or Student’s t-test. ns-P > 0.05, ***P < 0.001, ****P < 0.0001. B Correlations between ACE2 and the enrichment scores of several therapeutic signatures. C Correlation between ACE2 and the drug-target genes extracted from the Drugbank database. D Differences in IC50 of common anti-cancer drugs between the high and low ACE2 groups. Significance was calculated with Student’s t-test. ****P < 0.0001

Fig. 4. Roles of ACE2 in predicting clinical and immune phenotypes in the recruited TMA cohort. A Schematic protocol of validation on the TMA cohort. B Representative images revealing ACE2 expression in tumor and para-tumor tissues using anti-ACE2 staining. Magnification, 200 × . C Expression levels of ACE2 in tumor and para-tumor tissues. Significance was calculated with Mann Whitney test for ER and PR status and Student’s t-test for HER2 and TNBC status. *P < 0.05. D Representative images revealing ACE2 expression in BC tissues with different molecular features. Magnification, 200 × . E Expression levels of ACE2 in various molecular subtypes. Significance was calculated with Student’s t-test. *P < 0.05. F Representative images revealing CD8+ T cell infiltration and PD-L1 expression in the high and low ACE2 groups. Magnification, 200 × . G Differences in CD8+ T cell infiltration between the high and low ACE2 groups. Significance was calculated with Student’s t-test. *P < 0.05. H Differences in PD-L1 expression between the high and low ACE2 groups. Significance was calculated with Student’s t-test. *P < 0.05. I Representative images uncovering CD8.+ T cell infiltration and ACE2…

Fig. 5. Ang-1–7 sensitizes the response to chemotherapy and immunotherapy. A Proliferative potential of MDA-MB-231 cells was checked using CCK-8 assay after ACE2 overexpression. B The transcriptional level of PD-L1 was checked using qPCR after ACE2 overexpression. C The protein level of PD-L1 was checked using Western blotting after ACE2 overexpression. D Schematic protocol of the combinated therapy in immunocompetent BLAB/c mice. E Tumor volume was monitored every three days. Significance was calculated with One-way ANOVA with Tukey’s multiple comparisons test. *P < 0.05, ***P < 0.001. F The tumor weight of isolated tumors from these six groups, mice were sacrificed on day 25 after treatment. Significance was calculated with One-way ANOVA with Tukey’s multiple comparisons test. ***P < 0.001, ****P < 0.0001. G Relative infiltration level of CD3+ and CD8+ T cells in control and Ang-1–7 treated tumors. Magnification, 200 × . H Schematic diagram of time point of plasma sample collection. I Comparison of plasma Ang-1–7 levels pre-and post-chemotherapy. Significance was calculated with Student’s t-test. ns-P > 0.05. J Correlation between the response to neoadjuvant chemotherapy and plasma…