Sulforaphane as an adjunctive to everolimus counteracts everolimus resistance in renal cancer cell lines.
BACKGROUND
The mechanistic target of rapamycin (mTOR) inhibitors, everolimus and temsirolimus, have widened therapeutic options to treat renal cell carcinoma (RCC). However, chronic treatment with these inhibitors often induces resistance, leading to therapeutic failure.


OBJECTIVE
The natural compound, sulforaphane (SFN), was added to an everolimus based regime in vitro in the hopes of preventing resistance development.


METHODS
A panel of RCC cell lines (A498, Caki-1, KTCTL-26) was treated with everolimus or SFN or with an everolimus-SFN-combination, either short- (24h) or long-term (8 weeks), and cell growth, proliferation, apoptosis, and cell cycle phases were measured. The cell cycle regulating proteins cdk1, cdk2, cyclin A, cyclin B, akt and raptor (both total and activated) were also evaluated.


RESULTS
Short-term incubation with everolimus (1nM) or SFN (5µM) significantly reduced RCC cell growth. Additive effects on tumor growth and proliferation were evoked by the SFN-everolimus combination. Long-term everolimus-incubation led to resistance development in Caki-1 cells, evidenced by elevated growth and proliferation, associated with an increased percentage of G2/M (non-synchronized cell model) or S-phase (synchronized cell model) cells. Molecular analysis revealed up-regulation of the cdk1-cyclin B and cdk2-cyclin A axis, along with elevated phosphorylation of the mTOR sub-member, raptor. In contrast, resistance development was not observed with the long-term combination of SFN-everolimus. The combination suppressed Caki-1 growth and proliferation, and was associated with an increase in G0/G1-phase cells, diminished cdk1 and akt (both total and activated), cyclin B and raptor expression.


CONCLUSIONS
Adding SFN to an everolimus based RCC treatment regimen in vitro delayed resistance development observed with chronic everolimus monotherapy. Ongoing in vivo studies are necessary to verify the in vitro data.
Department of Urology, Goethe-University, Interdisciplinary Science Building, Building 25A, Room 404, Theodor-Stern-Kai 7, Frankfurt am Main, D-60590, Germany.
Eva
E
Juengel
Department of Urology, Goethe-University, Interdisciplinary Science Building, Building 25A, Room 404, Theodor-Stern-Kai 7, Frankfurt am Main, D-60590, Germany.
Stephanie
S
Euler
Department of Urology, Goethe-University, Interdisciplinary Science Building, Building 25A, Room 404, Theodor-Stern-Kai 7, Frankfurt am Main, D-60590, Germany.
Sebastian
S
Maxeiner
Department of Urology, Goethe-University, Interdisciplinary Science Building, Building 25A, Room 404, Theodor-Stern-Kai 7, Frankfurt am Main, D-60590, Germany.
Jochen
J
Rutz
Department of Urology, Goethe-University, Interdisciplinary Science Building, Building 25A, Room 404, Theodor-Stern-Kai 7, Frankfurt am Main, D-60590, Germany.
Saira
S
Justin
Department of Urology, Goethe-University, Interdisciplinary Science Building, Building 25A, Room 404, Theodor-Stern-Kai 7, Frankfurt am Main, D-60590, Germany.
Frederik
F
Roos
Department of Urology, Goethe-University, Interdisciplinary Science Building, Building 25A, Room 404, Theodor-Stern-Kai 7, Frankfurt am Main, D-60590, Germany.
Wael
W
Khoder
Department of Vascular and Endovascular Surgery, Goethe-University, Frankfurt am Main, Germany.
Karen
K
Nelson
Department of Urology, Goethe-University, Interdisciplinary Science Building, Building 25A, Room 404, Theodor-Stern-Kai 7, Frankfurt am Main, D-60590, Germany; Department of General and Visceral Surgery, Goethe-University, Frankfurt am Main, Germany.
Wolf O
WO
Bechstein
Department of Urology, Goethe-University, Interdisciplinary Science Building, Building 25A, Room 404, Theodor-Stern-Kai 7, Frankfurt am Main, D-60590, Germany. Electronic address: blaheta@em.uni-frankfurt.de.
Roman A
RA
Blaheta
D016588:Anticarcinogenic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D002292:Carcinoma, Renal Cell / Q000188:drug therapy*; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor / Q000187:drug effects*; D049109:Cell Proliferation / Q000187:drug effects; D019008:Drug Resistance, Neoplasm / Q000187:drug effects*; D000068338:Everolimus / Q000494:pharmacology*; D006801:Humans; D017879:Isothiocyanates / Q000494:pharmacology*; D020123:Sirolimus / Q000031:analogs & derivatives* / Q000494:pharmacology
D016588:Anticarcinogenic Agents; D017879:Isothiocyanates; C401859:temsirolimus; D000068338:Everolimus; D020123:Sirolimus
Cell growth; Everolimus; Renal cell carcinoma; Resistance; Sulforaphane