Molecular Signatures of Tumor-Initiating Cells Unveil Wnt Pathway As a Therapeutic Target in Mantle Cell Lymphoma

Introduction

Mantle cell lymphoma (MCL) is an aggressive and incurable form of non-Hodgkin’s lymphoma. Despite initial responses to intense chemotherapy, up to 50% of cases of MCL relapse, often in a chemoresistant form. We hypothesized that the recently identified MCL-initiating cells (MCL-ICs) are the main reason for relapse and chemoresistance of MCL.

Methods

We isolated MCL-ICs from primary MCL cells on the basis of a defined marker expression pattern; CD34-CD3-CD45+CD19-. The MCL-ICs, MCL-non-ICs, and peripheral blood lymphocytes from healthy donors were analyzed for gene expression using the Arraystar platform. The differences in mRNA levels of genes of interest were confirmed by quantitative RT-PCR. The prominent differentially expressed transcripts were analyzed using the Ingenuity Platform. Primary MCL cells were co-culture with mesenchymal stem cells to assess the effects of chemotherapeutic agents such as vincristine, doxorubicin and the newly approved Burton tyrosine kinase inhibitor ibrutinib, and Wnt signaling inhibitors.

Results

Approximately 1% of primary MCL cells are MCL-ICs and they can be maintained in co-culture with mesenchymal stem cells. Comparison of gene expression profiles of MCL-ICs and MCL-non-ICs revealed activation of stem cell-specific pathways in MCL-ICs by expression of Wnt, Notch, and Hedgehog and enhanced expression of Nanog, Oct4, KLF4, ADH1, MT1b and ABCC3. Gene expression microarray data and RT-PCR data suggested predominant activation of the Wnt/Frizzled pathway. Indeed, MCL-ICs were particularly sensitive to Wnt pathway inhibitors. Targeting Wnt-dependent β-catenin‒TCF4 interaction with CCT036477, iCRT14, or PKF118-310 preferentially eliminated the MCL-ICs, reduced the expression of stem cell transcription factors (Myc, Nanog, Oct4, Klf4), and sensitized MCL cells to vincristine, doxorubicin, and ibrutinib. Interestingly, while vincristine, doxorubicin or ibrutinib did kill MCL cells, they did not reduce the percentage of MCL-ICs in treated co-culture.

Conclusion

MCL-ICs are present in primary MCL isolates and they show gene expression pattern of chemoresistant, stem cell-like cells with predominant activation of Wnt signaling. In order to produce durable remissions in MCL patients, treatment strategies should be directed to target MCL-ICs.