Marked Activity of Irinotecan and Rapamycin Combination toward Colon Cancer Cells In vivo and In vitro Is Mediated through Cooperative Modulation of the Mammalian Target of Rapamycin/Hypoxia-Inducible Factor-1α Axis

Abstract

Purpose: Despite recent progress, colon cancer is often resistant to combination chemotherapy, highlighting the need for development of novel therapeutic approaches. An attractive target is hypoxia-inducible factor-1a (HIF-1a), a key transcription factor with a pivotal role in tumor cell metabolism. One potential class of therapeutic agents targeting HIF-1a are mammalian target of rapamycin inhibitors such as rapamycin. A second class are topoisomerase I inhibitors, such as irinotecan, which are able to inhibit the accumulation of HIF-1a. We here investigated whether combination of rapamycin and irinotecan was active in human colon cancer models. Experimental Design: Human metastatic tumors were xenografted in nude mice and treated with low doses of irinotecan alone, rapamycin alone, or combination of both drugs. The cellular effects of irinotecan and rapamycin were further characterized for HT-29 and HCT-116 colon cancer cells in vitro. Results: In contrast to single-agent therapy, xenografted tumors treated with combination of irinotecan and rapamycin showed potent inhibition of the mammalian target of rapamycin/ HIF-1a axis, which was accompanied by a dramatic reduction in tumor volume. In vitro experiments showed that exposure to low concentrations of the two drugs resulted in massive HT-29 cell death under hypoxic, but not normoxic, conditions, in full agreement with a cytotoxic effect mediated through HIF-1a rather than through induction of genotoxic lesions. HCT-116 cells were less sensitive to the combined treatment due to constitutive activation of phosphatidylinositol 3-kinase/Akt and Ras/mitogen-activated protein kinase pathways. Conclusion: These results identify HIF-1a as a promising target and provide a rationale for clinical trials of low-dose irinotecan and rapamycin combination toward metastatic colon cancer.

In Vivo Topoisomerase I Inhibition Attenuates the Expression of Hypoxia-Inducible Factor 1α Target Genes and Decreases Tumor Angiogenesis

“…Reduced expression of these genes may result in decreased ATP production by the glycolytic pathway, which is needed for tumor growth during hypoxia (32,33). This in vivo signature is in agreement with our previous observations showing that downregulation of HIF1A by siRNA in colon cancer cells induces antiproliferative effects in hypoxia conditions, validating HIF1A as a functionally important target (21).…”

“…These results are also consistent with those reported for topotecan, another topoisomerase I inhibitor that did not affect HIF1A mRNA accumulation or protein half-life, but rather acted on the translational level (54). In addition, we have recently shown that inhibition of HIF1A protein accumulation by irinotecan does not rely on inhibition of the mammalian target of rapamycin (mTOR) pathway (21).…”

“…Reverse transcription of 2 μg RNA was performed using AMV reverse transcriptase (Finnzymes, Ozyme, Saint Quentin, France) and oligo(dT) primers. Validated QuantiTect ® Primer Assays (Qiagen, Courtaboeuf, France) were used for amplification as previously described (21). All quantifications were performed in duplicate and normalized to two independent endogenous reference genes, hydroxymethylbilane synthase (HMBS) and keratin 19 (KRT19).…”

Tirapazamine Sensitizes Hepatocellular Carcinoma Cells to Topoisomerase I Inhibitors via Cooperative Modulation of Hypoxia-Inducible Factor-1α

“…E and F, effects of CPT-11 and/or tirapazamine on expression levels of HIF-1a in Bel-7402 cellderived tumors were determined by H&E staining and immunohistochemical and immunofluorescence analysis. a protective effect against DNA damage induced by topoisomerase inhibitors (32,33). As shown in Supplementary Fig.…”

Rapamycin-topotecan combination exhibited enhanced antitumor efficacy compared to topotecan used alone in cisplatin-resistant ovarian cancer cells

“…Rapamycin is a mTOR inhibitor that shows potent antitumor activity in a variety of malignances including ovarian cancers in preclinical and clinical studies [8][9][10]. Notably, rapamycin and its derivatives have been demonstrated to enhance the efficacy of cytotoxic chemotherapeutic drugs including topotecan, oxaliplatin, and 5-fluorouracil (5-FU) against various types of cancers such as endometrial, gastric and colon cancer [8,[11][12][13][14]. Given that mTOR signaling pathway was shown to be hyperactivated in platinum-resistant ovarian carcinoma cells [15], we investigated the efficacy of combination of topotecan with rapamycin against cisplatin-resistant ovarian carcinoma in the present study.…”