Identification of a nucleoside analog active against adenosine kinase–expressing plasma cell malignancies
Utthara Nayar, … , Kenneth M. Kaye, Ethel Cesarman
Utthara Nayar, … , Kenneth M. Kaye, Ethel Cesarman
Published May 15, 2017
Citation Information: J Clin Invest. 2017;127(6):2066-2080. https://doi.org/10.1172/JCI83936.
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Research Article Oncology

Identification of a nucleoside analog active against adenosine kinase–expressing plasma cell malignancies

Abstract

Primary effusion lymphoma (PEL) is a largely incurable malignancy of B cell origin with plasmacytic differentiation. Here, we report the identification of a highly effective inhibitor of PEL. This compound, 6-ethylthioinosine (6-ETI), is a nucleoside analog with toxicity to PEL in vitro and in vivo, but not to other lymphoma cell lines tested. We developed and performed resistome analysis, an unbiased approach based on RNA sequencing of resistant subclones, to discover the molecular mechanisms of sensitivity. We found different adenosine kinase–inactivating (ADK-inactivating) alterations in all resistant clones and determined that ADK is required to phosphorylate and activate 6-ETI. Further, we observed that 6-ETI induces ATP depletion and cell death accompanied by S phase arrest and DNA damage only in ADK-expressing cells. Immunohistochemistry for ADK served as a biomarker approach to identify 6-ETI–sensitive tumors, which we documented for other lymphoid malignancies with plasmacytic features. Notably, multiple myeloma (MM) expresses high levels of ADK, and 6-ETI was toxic to MM cell lines and primary specimens and had a robust antitumor effect in a disseminated MM mouse model. Several nucleoside analogs are effective in treating leukemias and T cell lymphomas, and 6-ETI may fill this niche for the treatment of PEL, plasmablastic lymphoma, MM, and other ADK-expressing cancers.

Authors

Utthara Nayar, Jouliana Sadek, Jonathan Reichel, Denise Hernandez-Hopkins, Gunkut Akar, Peter J. Barelli, Michelle A. Sahai, Hufeng Zhou, Jennifer Totonchy, David Jayabalan, Ruben Niesvizky, Ilaria Guasparri, Duane Hassane, Yifang Liu, Shizuko Sei, Robert H. Shoemaker, J. David Warren, Olivier Elemento, Kenneth M. Kaye, Ethel Cesarman

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Figure 1

Selection of 6-ETI.

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Selection of 6-ETI. (A) BC3 (KSHV+) and LCL9001 (KSHV–) cells were treat...
(A) BC3 (KSHV+) and LCL9001 (KSHV–) cells were treated for 24 hours with selected doses of NSC39368 (6-ETI), followed by nuclear extraction and EMSA to determine binding to an NF-κB response element. Control lanes include no compound, competition with cold probe, untreated, and DMSO-treated samples. Experiment was performed 3 separate times. A representative blot is shown. (B) A panel of KSHV+ (black bars) and KSHV– (gray bars) cell lines was plated at 1 × 105 cells/ml in RPMI with 20% FBS with increasing concentrations of 6-ETI, followed by analysis for ATP content by CellTiter-Glo assay at 48 hours. LC50s were determined by online EC50 software. Results are mean ± SEM of at least 3 independent experiments. (C) BC3 and IBL1 cells plated at 1 × 105 cells/ml in RPMI with 20% FBS at a range of concentrations of 6-ETI were examined for cell death by trypan blue staining–aided cell counts at 48 hours after treatment. Results plotted are mean ± SEM of at least 3 independent experiments. (D) BC3 and IBL1 cells plated at 2 × 105 cells/ml in RPMI with 20% FBS were treated with DMSO or 500 nM or 5 μM 6-ETI for 48 hours and analyzed by flow cytometry for cell death after staining with annexin V and 7-aminoactinomycin D. Results are the average of at least 2 independent experiments (mean ± SEM).