Alisertib Plus Ibrutinib and Rituximab Shows Preclinical Efficacy in MCL Models

Combining the Aurora A kinase ATP-site inhibitor alisertib (MLN8237) with ibrutinib (Imbruvica) plus rituximab (Rituxan) displayed significantly stronger tumor growth inhibition compared with doublet therapies in mantle cell lymphoma (MCL), according to findings from a preclinical study published in Cancers.

Findings from an MCL mouse model demonstrated that the triplet therapy led to tumor growth inhibition of approximately 82% compared with approximately 76% for ibrutinib plus rituximab and approximately 62% for alisertib plus ibrutinib. Ibrutinib monotherapy demonstrated tumor growth inhibition of approximately 50% and the rate for alisertib monotherapy was approximately 13%. All treatment approaches were found to be well tolerated and no significant differences to body weight were reported throughout the study.

Kaplan-Meier survival estimates showed a significant difference between the vehicle control compared with mice who received the doublet and those who received the triplet (P < .0001). There was also a significant survival difference between the ibrutinib- vs combination-treated mice (P < .0001). No significant differences between the combinations (P ≥ .589) vs ibrutinib or the control group were reported.

“Previously, we showed in a pre-clinical model and in a clinical trial that combining alisertib with vincristine plus rituximab is synthetically lethal in aggressiveB-cell non-Hodgkin lymphoma, including MCL,” the study authors wrote. “Here, we replaced vincristine with ibrutinib and evaluated the effects of alisertib plus ibrutinib, ibrutinib plus rituximab, and alisertib plus ibrutinib plus rituximab, respectively, in a Granta-519 MCL cell-derived xenograft mouse model.”

To conduct their study, investigators obtained MCL cell lines consisting of Granta-519 cells from Deutsche Sammlung von Mikroorganismen and Zellkulturen GmbH as well as JeKo-1 cells from ATCC mycoplasma which had tested negative. Cell lines were maintained in an RPMI 1640 medium with L-glutamine IX supplemented with 10% fetal bovine serum at 37 degrees Celsius in a humidified atmosphere containing 5% CO2.

Study authors cultured Granta-519 and JeKo-1 cells in 96-well culture plates seeded at 10,000 per well. Cells were allowed to grow for 1 day then the treatments were designed with increasing concentrations of the agents for 4 days. The equipotent ratio of alisertib in combination with ibrutinib was calculated to determine the combination index.

The mice received a subcutaneous injection of 1 x 107 cells of Granta-519 cells into the right hind flank and were divided into experimental and control arms. Mice in the experimental arm were further divided into 4 groups of 12 mice each. When tumors reached a volume of 100 mm3, groups received the following therapies: saline (control group), ibrutinib, ibrutinib plus alisertib, ibrutinib plus rituximab, and ibrutinib plus alisertib and rituximab. Ibrutinib was given at a dose of 10 mg/kg orally once daily for 3 weeks, alisertib was administered at 30 mg/kg orally once daily for 3 weeks, and rituximab was given at 10 mg/kg intravenously once a week for 4 weeks.

Additional findings from the study showed that Granta-519 and JeKo-1 MCL cell lines treated with alisertib displayed IC50 values of 23.7 nM and 3.01 nM, respectively. Comparatively, ibrutinib treatment led to an IC50 of 14.35 µM and 0.56 µM, respectively. When ibrutinib was present at 0.95 µM, the alisertib cytotoxicity curve shifted to the left when the IC50 was lowered to 12.5 nM in Granta-519 cells, demonstrating the synergy between the agents.

“In contrast, in the presence of 0.56 µM of ibrutinib, the IC50 of alisertib in JeKo-1 cells was 4.2 nM vs 3.01 nM in the absence of ibrutinib. However, combination index analysis suggests that ibrutinib and alisertib were synergistic on both Granta-519 and JeKo-1 cells,” the study authors added.

Furthermore, data from an apoptosis analysis by cell cycle analysis showed that alisertib plus ibrutinib progressively increased apoptosis over ibrutinib alone at 48 hours, 72 hours, 96 hours, and 7 days in Granta-519 cells. Additionally, 1 week of alisertib treatment led to a significant decrease in phosphorylated RSK1/2/3 and Hck (P = .01) and a modest reduction in p38 phosphorylation (P = .068). No p53 phosphorylation sites were affected by alisertib.

“Our study suggests that alisertib has multidimensional effects, suppressing numerous proteins in the cell proliferative signaling pathways; alisertib and ibrutinib are synergistic, treating ibrutinib-insensitive MCL,” the study authors wrote. “A B-cell receptor-related gene signature may be a predictive biomarker for the therapeutic response to alisertib and ibrutinib combination.”

Reference

1. Subramani B, Conway PJ, Al-Khinji A, Zhang K, Pandey R, Mahadevan D. A novel triplet of alisertib plus ibrutinib plus rituximab is active in mantle cell lymphoma. Cancers (Basel). 2024;16(24):4257. doi:10.3390/cancers16244257