Abstract
Abstract
The activated B-cell cell-of-origin subtype of diffuse large B-cell lymphoma (ABC-DLBCL) requires NF-kB pathway activation to maintain the malignant phenotype. NF-kB activation is downstream of B-cell receptor (BCR) stimulation and can become constitutively turned on in ABC-DLBCL through mutations in multiple different BCR signaling intermediates. Drugs targeting upstream signaling proteins such as Bruton’s Tyrosine Kinase (BTK, ibrutinib) or protein kinase C (PKC, AEB071) have shown promising results in other lymphomas driven by BCR activation and are under evaluation in ABC-DLBCL. Many ABC-DBCL cases, however, have mutations in mediators that are downstream from these targets, particularly coiled-coil domain mutations in CARD11 that stabilize NF-kB activation and A20 loss-of-function alterations that reduce protein turnover of oncogenic NF-kB intermediates. In this study we explore a potential role for PIM kinase inhibition in ABC-DLBCL and find the clinical pan-PIM inhibitor LGH447 has promising activity in particular against cells carrying these downstream mutations. We find some ABC cells were highly sensitive to LGH447 with IC50 < 0.4 µM (OCI-Ly3 and OCI-Ly10), while some others were completely insensitive with IC50 > 10.0 µM (TMD8 and HBL-1). Strikingly, all ABC lines sensitive to LGH447 carry mutations in either CARD11, TNFAIP3 (encoding A20), or both, while insensitive typically lines lack such lesions. Insensitive lines including TMD8 and HBL-1 instead have upstream mutations in CD79B and are highly sensitive to the upstream inhibitors ibrutinib and AEB071. The PIM1-3 kinases inhibited by LGH447 have multiple targets mediating cell growth and survival, including several that activate cap-dependent protein translation activation. We find LGH447 is toxic to sensitive cells due to lost translational activation. Western blots show reduced phosphorylation of ribosomal protein S6 and 4EBP1, indicating loss of mTORC1 activity. In addition, LGH447, in a manner similar to the potent direct cap-dependent translation inhibitor silvestrol, causes knockdown of key translationally regulated oncoproteins, including c-MYC, MCL1, and Cyclin D3. We also directly monitored protein synthesis through O-Propargyl-puromycin (OP-PURO) incorporation and found a direct effect of LGH447 that was similar to silvestrol, although requiring higher concentrations. PIM’s effects on activation of protein translation therefore are required in LGH447-sensitive ABC-DLBCL cells but dispensable in insensitive cells. In conclusion, pan-PIM kinase inhibition provides a strong potential therapeutic opportunity in a subset of ABC-DLBCL. Cases that bypass upstream signaling to turn on NF-kB activation more directly also bypass pathways with redundant activation of cap-dependent translation, making them dependent on the therapeutically targetable PIM kinases to carry out this critical process.
Disclosures
No relevant conflicts of interest to declare.