In Vivo Drug-Response in Patients with Leukemic Non-Hodgkin’s Lymphomas Is Predicted by In Vitro Chemosensitivity Testing and Gene Expression Profiling.

Only a few approaches are available to address the mechanisms of cell death in vivo which are induced by anticancer treatment in patients with malignancies. In this study in vitro chemosensitivity testing of primary peripheral blood leukemic cells of five patients suffering from different leukemic Non-Hodgkin’s lymphomas (atypical CLL, typical CLL, Immunocytoma, Mantle Cell Lymphoma, Prolymphocytic Leukemia (PLL)) was combined with the analysis of the in vivo rate of apoptosis by flow-cytometry (Annexin V and depolarisation of mitochondrial membrane potential (MMP) by JC-1). Furthermore, changes in expression patterns of apoptosis related proteins during chemotherapeutic treatment were detected by Western Blot. Gene expression profiling (HG-U133A, Affymetrix, Santa Clara, CA) was employed to identify common marker genes of in vivo drug response. In vitro chemosensitivity was tested using the cytotoxic agents which the patients were scheduled to receive and was strongly correlated with effective reduction of leukemic lymphoma cells in patients resulting in complete remissions in all five cases. Due to the rapid clearance of apoptotic tumor cells in vivo neither the analysis of the in vivo rate of apoptosis and depolarisation of MMP nor the assessment of expression of regulators of apoptosis showed concordant results concerning the drug response. However, assessment of gene expression during therapy could identify a set of 30 genes to significant discriminate between samples from patients before treatment compared to samples from the same patients after receiving cytotoxic therapy. Among these 30 genes we found a high proportion of genes associated with apoptotic cell death and cell proliferation signalling including complement lysis inhibitor (clusterin, CLU, SP40), beta-catenin interacting protein (ICAT), peroxisome proliferator activated receptor alpha (PPARα), TNF alpha converting enzyme (ADAM 17 / TACE), homeo box A3 (HOXA), inositol polyphosphate 5 phophatase (PPI 5 PIV, SHIP1), FK 506 binding protein (FKBP 38) and inhibitor of p53 induced apoptosis alpha (NME 6). Clusterin is able to mediate apoptosis via p53 and increases drug-induced cell death when overexpressed as detected in our treated samples. The downregulation of NME 6 during chemotherapeutic treatment may enhance this effect. These results indicate that in vitro chemosensitivity testing and gene expression profiling can successfully be utilised to predict in vivo drug response in patients with leukemic NHL’s and can be used to explore new pathway models of drug-induced cell death in vivo which are independent of different lymphoma subtypes and different treatment regimens.