Efficiency of the induction protocol is crucial for the generation of insulin-producing cells (IPCs) from human dental pulp stem cells (hDPSCs). Here, we established the integrative induction protocol by merging genetic manipulation technique with our previous published 3-step induction protocol aiming to enhance the pancreatic progenitor commitment and production yield. We found that the over-expression of PDX1 following with 3-step induction protocol were able to generate the 3-dimensional (3D) colony structure of pancreatic progenitors (PPs) with the beneficial trends of pancreatic endo-derm commitment and production yield, while other protocols using the prolong maintenance of PDX1-overexpressed hDPSCs and the PDX1 overexpression after definitive endoderm induction were unable to generate and sustain the 3D structure of the colonies. Further Notch signaling manipulation by DAPT treatment showed lesser degree of positive effects on progenitor commitment and production yield. Although the generated PPs from the integrative protocol expressed pancreatic mRNA markers along with pro-insulin and insulin proteins, they still contained the defective glucose-responsive C-peptide secretion. Only basal secreted C-peptide level was observed. In summary, the integrative induction protocol potentially enhanced the PP generation with high colony production yield and could serve as an efficient platform for further hDPSC-derived IPC production and maturation. (C) 2020 Elsevier Inc. All rights reserved.