Two fundamental parameters of the highly dynamic, ultrathin lamellipodia of migrating fibroblasts have been determined-its thickness in living cells (176 +/- 14 nm), by standing-wave fluorescence microscopy, and its F-actin density (1580 +/- 613 microm of F-actin/microm(3)), via image-based photometry. In combination with data from previous studies, we have computed the density of growing actin filament ends at the lamellipodium margin (241 +/- 100/microm) and the maximum force (1.86 +/- 0.83 nN/microm) and pressure (10.5 +/- 4.8 kPa) obtainable via actin assembly. We have used cell deformability measurements (. J. Cell Sci. 44:187-200;. Proc. Natl. Acad. Sci. USA. 79:5327-5331) and an estimate of the force required to stall the polymerization of a single filament (. Proc. Natl. Acad. Sci. USA. 78:5613-5617;. Biophys. J. 65:316-324) to argue that actin assembly alone could drive lamellipodial extension directly.

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PARP inhibitors have recently been approved as monotherapies for the treatment of recurrent ovarian cancer and metastatic BRCA-associated breast cancer, and ongoing studies are exploring additional indications and combinations with other agents. PARP inhibitors trap PARP onto damaged chromatin when combined with temozolomide and methyl methanesulfonate, but the clinical relevance of these findings remains unknown. PARP trapping has thus far been undetectable in cancer cells treated with PARP inhibitors alone. Here, we evaluate the contribution of PARP trapping to the tolerability and efficacy of PARP inhibitors in the monotherapy setting. We developed a novel implementation of the proximity ligation assay to detect chromatin-trapped PARP1 at single-cell resolution with higher sensitivity and throughput than previously reported methods. We further demonstrate that the PARP inhibitor-induced trapping appears to drive single-agent cytotoxicity in healthy human bone marrow, indicating that the toxicity of trapped PARP complexes is not restricted to cancer cells with homologous recombination deficiency. Finally, we show that PARP inhibitors with dramatically different trapping potencies exhibit comparable tumor growth inhibition at MTDs in a xenograft model of BRCA1-mutant triple-negative breast cancer. These results are consistent with emerging clinical data and suggest that the inverse relationship between trapping potency and tolerability may limit the potential therapeutic advantage of potent trapping activity. Implications: PARP trapping contributes to single-agent cytotoxicity of PARP inhibitors in both cancer cells and healthy bone marrow, and the therapeutic advantage of potent trapping activity appears to be limited.

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The ability of a cancer cell to avoid apoptosis is crucial to tumorigenesis and can also contribute to chemoresistance. The Bcl-2 family of prosurvival proteins (Bcl-2, Bcl-X L , Bcl-w, Mcl-1, and A1) plays a key role in these processes. We previously reported the discovery of ABT-263 (navitoclax), a potent small-molecule inhibitor of Bcl-2, Bcl-X L , and Bcl-w. While navitoclax exhibits single-agent activity in tumors dependent on Bcl-2 or Bcl-X L for survival, the expression of Mcl-1 has been shown to confer resistance to navitoclax, most notably in solid tumors. Thus, therapeutic agents that can downregulate or neutralize Mcl-1 are predicted to synergize potently with navitoclax. Here, we report the activity of navitoclax in combination with 19 clinically relevant agents across a panel of 46 human solid tumor cell lines. Navitoclax broadly enhanced the activity of multiple therapeutic agents in vitro and enhanced efficacy of both docetaxel and erlotinib in xenograft models. The ability of navitoclax to synergize with docetaxel or erlotinib corresponded to an altered sensitivity of the mitochondria toward navitoclax, which was associated with the downmodulation of Mcl-1 and/or upregulation of Bim. These data provide a rationale to interrogate these combinations clinically.