Fig. 7: MIF promotes cancer cell growth in vitro and in vivo through its nuclease activity.

a, b Colony formation of parental, MIF-KO2, KO2-WT, and KO2-E22A MDA-MB-231 cells cultured for 14 days. Representative images from three experiments were shown in a and quantification of colony numbers was shown in b (mean ± SEM). Statistical significance was determined by one-way ANOVA Dunnett’s multiple comparisons test. c–e Growth of parental, MIF-KO2, KO2-WT, and KO2-E22A MDA-MB-231 tumors in mice. Tumor volume, image, and weight were shown in c, d and e, respectively (mean ± SEM, n = 5 mice). ****P < 0.0001 parental vs MIF-KO2, ****P < 0.0001 MIF-KO2 vs KO2-WT, **P = 0.0013, KO2-WT vs KO2-E22A by two-way ANOVA Tukey’s multiple comparisons (c) and statistical significance in e was determined by two-tailed Student’s t test. f–h Growth of parental, WT MIF-, and E22A-MIF-overexpressed (OE) MDA-MB-231 tumors in mice. Tumor volume, image, and weight were shown in f, g, and h, respectively (mean ± SEM, n = 5 mice per group). ****P < 0.0001 by two-way ANOVA Tukey’s multiple comparisons (f) and one-way ANOVA Dunnett’s multiple comparisons test (h). i Immunoblot analysis of WT and E22A MIF protein levels in overexpressed tumors. j–l Analysis of MIF mRNA levels in human breast tumors and normal breast tissues. Data were retrieved from the TCGA dataset and presented as mean ± SEM. n = 112 normal breasts, n = 1041 primary tumors, n = 7 metastatic tumors (j); n = 112 normal breast, n = 422 Luminal A, n = 194 Luminal B, n = 67 Her2 + , n = 142 basal-like (k); n = 112 normal breast, n = 133 Stage 1, n = 445 Stage 2, n = 175 Stage 3, n = 15 Stage 4 (l). **P < 0.01, ****P < 0.0001, versus normal breast, by one-way ANOVA Dunnett’s multiple comparisons test. m Kaplan–Meier survival analysis for patients with breast cancer. Patients were divided by median expression levels of MIF mRNA. Data were retrieved from the GEO dataset (GSE1456). Statistical significance was determined by log-rank test. n MIF genetic alteration analysis in TCGA PanCancer Atlas studies (cBioPortal database). o A proposed model of MIF in the regulation of DNA proofreading and tumor growth. PARP1 interacts with MIF and recruits MIF to the DNA replication sites, where PARP1 detects DNA damage including 3’ flap structures and MIF excises unpaired flaps to facilitate DNA synthesis executed by polymerases in cancer cells, leading to cancer cell proliferation. Inhibition of MIF suppresses DNA replication and cancer cell growth.