Combined chemotherapy is recommended strategy as a first-line treatment method in patients with cervical cancer. Ganetespib (STA-9090) is a second-generation heat shock protein 90 (Hsp90) inhibitor that blocks the ATPase function of Hsp90 and inhibits the proper folding of oncogenic client proteins. Venetoclax (ABT-199) is an orally bioavailable Bcl-2 (B-cell lymphoma 2) inhibitor that stimulates apoptotic signaling pathways in cancer cells. This study evaluated the anticancer effects of STA-9090 combined with Venetoclax in the human cervical cancer cell line (HeLa). The human cervical cancer cells were treated with STA-9090, Venetoclax, and Sta-9090 plus Venetoclax for 48 h, and cell viability was measured using the XTT assay. The alteration of the Hsp90 protein expression level and the chaperone activity of HSP90 were detected by ELISA and luciferase aggregation assay, respectively. For the apoptotic process, qRT-PCR was applied to study Bcl-2-associated X protein (Bax), B-cell lymphoma 2 (Bcl-2), Bcl-2-like protein 1 (Bcl-xL ), Cytochrome c (Cyt-c), Caspase3 (Cas-3), and Caspase7 (Cas-7) expression levels after drug treatments. Also, a colorimetric Cas-3 activity assay was performed to detect the induction of the apoptosis process. Our results demonstrated that 8 nM of STA-9090 combined with 4 µM of Venetoclax synergistically inhibited cervical cancer cell proliferation more than STA-9090 or Venetoclax alone after 48 h of treatment. STA-9090 and Venetoclax combination decreased the protein expression level of Hsp90 and significantly inhibited chaperone activity of Hsp90. This combination stimulated apoptosis in cervical cancer cells by down-regulating of anti-apoptotic markers while inducing pro-apoptotic markers. Also, the STA-9090-Venetoclax combination increased Cas-3 activity in Hela cells. Collectively, these findings pointed out that the STA-9090-Venetoclax combination exhibited more activity than the individual drugs to stimulate toxicity and apoptosis in cervical cancer cells based on HSP90 inhibition.