Cancer cells can communicate with the tumor microenvironment and contribute to tumor progression. However, the effects of drug-resistant tumor cells on angiogenesis are unclear. Current anti-angiogenic strategies also have limitations and it would be useful to develop novel targets and treatment strategies. Here, our study showed that the conditioned medium and exosomes from 5-FU-resistant colon cancer cells promoted angiogenesis, and we observed that exosomal dipeptidyl peptidase IV (DPP4) was a potent inducer of this angiogenesis. DPP4-enriched exosomes increased periostin (POSTN) expression in human umbilical vein endothelial cells via Twist1 nuclear translocation or activating Smad signaling pathway, while silencing or inhibition of DPP4 neutralized those effects. The in vivo and clinical data indicated that high DPP4 expression was related to tumor progression. These findings indicate that DPP4 may be a target for inhibiting angiogenesis in 5-FU-resistant colon cancer. Furthermore, exosomal DPP4 concentrations may be a useful prognostic marker for colon cancer.