Cancer stem cells (CSCs) are the tumor cell subpopulations that can self-renew, differentiate, initiate and maintain tumor growth. CSCs are frequently drug-resistant, resulting in tumor recurrence, metastasis, and angiogenesis. Herein, using in vitro oral squamous cell carcinoma (OSCC) CSCs and in vivo xenograft mice model, we have systematically studied the apoptotic potentiality of quinacrine-gold hybrid nanoparticle (QAuNP) and its underlying mechanism after NIR irradiation. QAuNP + NIR caused DNA damage and induced apoptosis in SCC-9-CSCs by deregulating mitochondrial membrane potential (ΔΨm) and activation of ROS. Upregulation of CASPASE-3 and DR-5/DR-4 and reduction of heat shock protein (HSP-70) up to 5-fold were also noticed upon the treatment. The increased expression of DR-5 and CASPASE-3 and decreased expression of HSP-70, CD-44 and Ki-67 were also noted in the xenograft mice treated with QAuNP + NIR + TRAIL. Thus, data suggest that the combined treatment enhances apoptosis in OSCC-CSCs by modulating HSP-70 in the DISC.