PIM-1–specific mAb suppresses human and mouse tumor growth by decreasing PIM-1 levels, reducing Akt phosphorylation, and activating apoptosis
Xiu Feng Hu, … , Nancy S. Magnuson, Pei Xiang Xing
Xiu Feng Hu, … , Nancy S. Magnuson, Pei Xiang Xing
Published January 19, 2009
Citation Information: J Clin Invest. 2009;119(2):362-375. https://doi.org/10.1172/JCI33216.
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Research Article Oncology

PIM-1–specific mAb suppresses human and mouse tumor growth by decreasing PIM-1 levels, reducing Akt phosphorylation, and activating apoptosis

Abstract

Provirus integration site for Moloney murine leukemia virus (PIM1) is a proto-oncogene that encodes a serine/threonine kinase with multiple cellular functions. Overexpression of PIM-1 plays a critical role in progression of prostatic and hematopoietic malignancies. Here we describe the generation of a mAb specific for GST–PIM-1, which reacted strongly with most human and mouse cancer tissues and cell lines of prostate, breast, and colon origin but only weakly (if at all) with normal tissues. The mAb binds to PIM-1 in the cytosol and nucleus as well as to PIM-1 on the surface of human and murine cancer cells. Treatment of human and mouse prostate cancer cell lines with the PIM-1–specific mAb resulted in disruption of PIM-1/Hsp90 complexes, decreased PIM-1 and Hsp90 levels, reduced Akt phosphorylation at Ser473, reduced phosphorylation of Bad at Ser112 and Ser136, and increased cleavage of caspase-9, an indicator of activation of the mitochondrial cell death pathway. The mAb induced cancer cell apoptosis and synergistically enhanced antitumor activity when used in combination with cisplatin and epirubicin. In tumor models, the PIM-1–specific mAb substantially inhibited growth of the human prostate cancer cell line DU145 in SCID mice and the mouse prostate cancer cell TRAMP-C1 in C57BL/6 mice. These findings are important because they provide what we believe to be the first in vivo evidence that treatment of prostate cancer may be possible by targeting PIM-1 using an Ab-based therapy.

Authors

Xiu Feng Hu, Jie Li, Scott Vandervalk, Zeping Wang, Nancy S. Magnuson, Pei Xiang Xing

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Figure 7

The effect of siRNA-mediated PIM1 knockdown on PIM-1 expression, cell growth, and phosphorylation of Akt and Bad.

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The effect of siRNA-mediated PIM1 knockdown on PIM-1 expression, cell gr...
(A–C) Flow cytometry analysis shows reduced FITC-conjugated P9 binding to cell surface PIM-1 in DU145 (A), PC3 (B) and TRAMP-C1 (C) cells, respectively, after 72 hours of transfection with PIM1 siRNA (dotted line) and control siRNA (solid line). FITC-conjugated mIgM was used as negative control in the PIM1 siRNA transfected cells (gray area). (D) Trypan blue exclusion assay shows that transfection with PIM1 siRNA for 72 hours induced 42%, 43%, and 39% of cell death in TRAMP-C1, PC3, and DU145 cells, respectively, compared with the cells transfected with controls siRNA. Columns show means of viable cell counts of triplicate experiments; bars represent ± SD. (E) Western blot analysis of the DU145 cells following PIM1 siRNA transfection shows decreased levels of 44-, 33-, and 37-kDa molecule of PIM-1; reduced phosphorylation of Akt at Ser473, without changing total Akt levels, and promoted dephosphorylation of Bad at Ser112 and Ser136 compared with total levels of Bad; and induced cleavage of caspase-9 at Asp33 following 66 hours (lane 2) and 78 hours (lane 3) transfection using PIM1 siRNA compared with control siRNA (lane 1).