Pharmacogenomic modeling of bortezomib resistance in B cell malignancies

Abstract

Proteasome inhibitors are a class of drugs that have been largely successful in the treatment of cancer patients, particularly those with the plasma cell malignancy, multiple myeloma. The most successful of these drugs, bortezomib (Bz), has paved the way for the development of next-generation proteasome inhibitors. Although Bz has significantly contributed to improved outcomes in myeloma patients, acquired resistance to Bz is imminent. Furthermore, a portion of patients never initially respond to the drug. Therefore, the goal of these studies was to further characterize Bz resistance with the aim to better predict secondary therapies that may be used successfully with Bz to recapture drug sensitivity.In the first study, we describe the creation of an <italic>in vitro</italic> malignant mouse plasma cell system from which we create isogenic pairs of Bz-sensitive and -resistant cell lines. We further characterize the transcriptional responses of these cell line pairs to identify both conserved and unique expression signatures. Using the expression signatures that are unique to each pair of cell lines, we identify secondary therapies that may be useful for treatment of the Bz-refractory cell line using an <italic>in silico</italic> database called Connectivity Map (CMAP). This analysis predicted a unique response to histone deacetylase inhibitors, a class of drugs that are currently being tested for efficacy in myeloma, in only one mouse cell line pair. Indeed, we find that the predicted Bz-resistant cell line has increased sensitivity to this class of drugs (including the drug panobinostat). When these cells were transferred back into syngeneic recipient mice, panobinostat treatment could successfully extend the life of Bz-resistant animals suggesting that the Bz-resistant phenotype may select also for increased sensitivity to other drugs that may be identified through <italic>in silico</italic> approaches. In the second study, we follow up these observations by investigating other CMAP prediction patterns, such as those that are conserved across all cell line pairs. A second prediction of one class of these CMAP-predicted drugs using high-throughput drug screening of the cell lines revealed that a combination of these approaches may be highly successful for accurate prediction of secondary therapies. Based on these predictions, we further investigate the efficacy of topoisomerase inhibitors in combination with Bz for the treatment of Bz-resistant cell lines.In the third study, we provide further immunophenotypic characterization of the Bz-sensitive and -resistant mouse cell lines revealing not only cell surface markers that are associated with "acquired" and "innate" Bz resistance but perhaps a mechanism of resistance. Although Bz-sensitive mouse cells display a classic myeloma phenotype, homing to the bone marrow <italic>in vivo</italic> and expressing classic plasma cell markers, Bz-resistant mouse cells present as extramedullary disease and express a more B cell-like immunophenotype. We identify that differences in migration may be linked to the differential expression of the bone marrow homing protein, CXCR4. Lower expression of this gene in a Bz human clinical trial was also associated with inferior survival. Immunophenotypic characterization of these cell populations further revealed that forced differentiation of the Bz-resistant population could restore Bz-sensitivity.The final study investigates the acquisition of Bz-resistance in a B cell malignancy, Burkitt lymphoma, that is currently undergoing Bz clinical trials. In this particular malignancy, a DNA mutator, AID, is known to be expressed that may contribute to other types of drug resistance. Here, we identify that this is unlikely a mechanism for developing resistance to Bz. Furthermore, we provide evidence that AID activity is reduced in Bz-resistant clones and, in fact, that high AID expression may be selectively eliminated during Bz selection.