Treatment with the HIV protease inhibitor nelfinavir triggers the unfolded protein response and may overcome proteasome inhibitor resistance of multiple myeloma in combination with bortezomib: a phase I trial (SAKK 65/08)

Abstract

Downregulation of the unfolded protein response mediates proteasome inhibitor resistance in multiple myeloma. The Human Immunodeficieny Virus protease inhibitor nelfinavir activates the unfolded protein response in vitro. We determined dose-limiting toxicity and recommended dose for phase II of nelfinavir in combination with the proteasome inhibitor bortezomib. Twelve patients with advanced hematologic malignancies were treated with nelfinavir (2500-5000 mg/day p.o., days 1-14, 3+3 dose escalation) and bortezomib (1.3 mg/m(2), days 1, 4, 8, 11; 21-day cycles). A run in phase with nelfinavir monotherapy allowed pharmakokinetic/pharmakodynamic assessment of nelfinavir in the presence or absence of concomittant bortezomib. End points included dose-limiting toxicity, activation of the unfolded protein response, proteasome activity, toxicity and response to trial treatment. Nelfinavir 2×2500 mg was the recommended phase II dose identified. Nelfinavir alone significantly up-regulated expression of proteins related to the unfolded protein response in peripheral blood mononuclear cells and inhibited proteasome activity. Of 10 evaluable patients in the dose escalation cohort, 3 achieved a partial response, 4 stable disease for 2 cycles or more, while 3 had progressive disease as best response. In an exploratory extension cohort with 6 relapsed, bortezomib-refractory, lenalidomide-resistant myeloma patients treated at the recommended phase II dose, 3 reached a partial response, 2 a minor response, and one progressive disease. The combination of nelfinavir with bortezomib is safe and shows promising activity in advanced, bortezomib-refractory multiple myeloma. Induction of the unfolded protein response by nelfinavir may overcome the biological features of proteasome inhibitor resistance. (clinicaltrials.gov identifier: 01164709).

Copyright© Ferrata Storti Foundation.

Figures

Figure 1.

Graphical overview of dosing of nelfinavir (dose levels DL0, DL1, DL2) and bortezomib together with the time points at which samples for pharmakokinetic/pharmakodynamic (PK/PD) analysis were collected (PK1-5). Nelfinavir dosing in week -1 was only performed in cycle 1.

Figure 2.

Overview of the best treatment response of all evaluable patients with multiple myeloma (MM) that have received trial treatment. The changes in paraprotein levels compared to baseline while on trial are displayed relative to baseline. Individual patients are represented by UPN numbers, patients of the phase I dose escalation cohort (UPN 1–12) in light gray, patients of the extension cohort (UPN 13–19) in dark gray. The streaked lines pattern denotes the only indivudual MM patient in the trial (UPN 08, dose escalation cohort) that was not bortezomib refractory by IMWG criteria before receiving bortezomib+nelfinavir therapy.

Figure 3.

Expression of unfolded protein response (UPR)-related proteins and proteasome activity in peripheral blood mononuclear cells (PBMC) from treated patients PBMC were collected at the indicated time points PK1 (base-line predose), PK2 (nelfinavir 4 h postdose), PK3 (nelfinavir trough + bortezomib pre dose), PK4 (nelfinavir+bortezomib 4 h post dose), PK5 (nelfinavir trough, 24 h post bortezomib). Equal amounts of cellular protein from PBMC was resolved by 12.5% SDS PAGE. (A) exemplary changes in expression of UPR-related protein and proteasome activity in PBMC from one patient (UPN04). Upper left: protein representing the UPR and its related apoptotic machinery (BIP, CHOP, PDI, PARP), as well as pAKT, was assessed from PBMC collected at the time points PK1–PK5, and visualized by western blot, GAPDH served as a control. Upper right: Quantitative comparison of proteasome activity (seperately for the β2-type and β1/β5-type subunits) at the different time points PK1–PK5 was achieved with activity based proteasome probes in the same PBMC samples (see Methods). (B) Mean quantitative changes in expression of UPR-related proteins and proteasome activity in PBMC for all patients Botom left: effect of nelfinavir monotherapy on expression of UPR related protein in PBMC. The relative change in expression of the UPR-associated proteins was assessed for all patients by western blot as (as shown for UPN04 above), and quantified by flourescence scanning comparing the time points PK1 (baseline) versus PK2 (post-nelfinavir); n= 11 patients, one technical failure). The mean relative difference from a base-line signal and standard deviation of expression of the proteins indicated is displayed for PK1 versus PK2. Bottom right: changes in proteasome activity in PBMC for the entire dose escalation cohort are shown after nelfinavir monotherapy (PK1 vs. PK3) and after treatment with nelfinavir in combination with bortezomib (PK1 vs. PK4). The relative changes in proteasome activity in PBMC were quantified using affinity-based probes and flourescence scanning. Mean changes from 11 patients and standard deviation are presented.

Figure 4.

Mean nelfinavir plasma concentrations (μM) for the 4 pharmacokinetic sampling points and the 3 dosing cohorts (1250, 1850 and 2500 mg bid). Significance levels are given for the difference between mean NFV peak concentration (PK2) and either mean NFV through concnetration (PK3) or NFV peak concentration in combination with BTZ (PK4).