Molecular Mechanisms of Patupilone Resistance

Abstract

Patupilone is an epothilone in advanced clinical development that has shown promising efficacy in heavily pretreated patients. This study aimed at characterizing the mechanisms of patupilone activity in resistant patients. To this end, we generated patupilone-resistant cells using two cellular models, the first characterized by high chemosensitivity and low class III B-tubulin (TUBB3) expression (A2780), and the second by low chemosensitivity and high TUBB3 expression (OVCAR-3). The obtained cell lines were named EPO3 and OVCAR-EPO, respectively. The same selection procedure was done in A2780 cells to generate a paclitaxel-resistant cell line (TAX50). Factors of resistance are expected to increase in the drugresistant cell lines, whereas factors of drug sensitivity will be down-regulated. Using this approach, we found up-regulation of TUBB3 in TAX50, but not EPO3, cells, showing that TUBB3 mediates the resistance to paclitaxel but not to patupilone. Moreover, TUBB3 was a factor of patupilone sensitivity because OVCAR-EPO cells exhibited a dramatic reduction of TUBB3 and a concomitant sensitization to hypoxia and cisplatin-based chemotherapy. To identify the mechanisms underlying patupilone resistance, tubulin genes were sequenced, thereby revealing that a prominent mechanism of drug resistance is represented by point mutations in class I B-tubulin. Overall, these results suggest that paclitaxel and patupilone have nonoverlapping mechanisms of resistance, thus allowing the use of patupilone for those patients relapsing after paclitaxel-based chemotherapy. Furthermore, patupilone represents a promising first-line option for the treatment of high-risk ovarian cancer patients, who exhibit high TUBB3 levels and poor response to standard paclitaxelplatin chemotherapy.

Gender Influences the Class III and V β-Tubulin Ability to Predict Poor Outcome in Colorectal Cancer

“…Oxaliplatin and SN-38 were purchased from Tocris and diluted in dimethyl sulfoxide. Growth inhibition effects were carried out as previously described (11), and a comprehensive table showing the IC 50 values for all the used cell lines is shown in Supplementary Table SI. All other chemicals were purchased from Sigma-Aldrich if not otherwise specified.…”

“…TUBB3 primary antibody was from Covance (Tuj1 clone), instead TUBB6 was developed in house as previously described and validated (11). AR (441 clone) was from Santa Cruz.…”

“…Characterization of the anti-TUBB6 antibody used in this study has been reported previously (11). To get information on the role of these antigens as predictive biomarkers, we carried out a retrospective analysis in paraffin-embedded samples from 180 colorectal cancer patients.…”

Class III beta-tubulin, drug resistance and therapeutic approaches in cancers

“…In one of the studies, class III beta-tubulin gene silencing desensitized effects of both taxanes as well as other drugs, in particular cisplatin. In another study, epothilones were found to be extremely active in ovarian cancer cell models and other gynecological cancers but not for lung cancer, even when both sets of cancers exhibited high class III beta-tubulin content (Gan et al, 2007;Mozzetti et al, 2008;Carrara et al, 2012). To further gain insight into this contradiction, our group recently did a comprehensive review of 59 translational studies assessing class III beta-tubulin in many different types of cancers.…”

IL-17A exacerbates cisplatin-based resistance of OVCA via upregulating the expression of ABCG2 and MDR1 through Gli1-mediated Hh signaling

“…One of major challenges of successful treatment in OVCA is overcoming multi-drug resistance (MDR) [18][19][20]. DDP is known to bind DNA to form cisplatin-DNA adducts and therefore could inhibit DNA replication or transcription of cancer cells [21,22]. As proved in previous reports, DDP-resistance occurs and develops through a plethora of molecular mechanisms, including enhancing cytotoxic agents extrusion by energy-dependent pumps such as ATP-binding cassette (ABC) transporters [23].…”