PARP-1 improves leukemia outcomes by inducing parthanatos during chemotherapy.

Previous chemotherapy research has focused almost exclusively on apoptosis. Here, a standard frontline drug combination of cytarabine and idarubicin induces distinct features of caspase-independent, poly(ADP-ribose) polymerase 1 (PARP-1)-mediated programmed cell death "parthanatos" in acute myeloid leukemia (AML) cell lines (n = 3/10 tested), peripheral blood mononuclear cells from healthy human donors (n = 10/10 tested), and primary cell samples from patients with AML (n = 18/39 tested, French-American-British subtypes M4 and M5). A 3-fold improvement in survival rates is observed in the parthanatos-positive versus -negative patient groups (hazard ratio [HR] = 0.28-0.37, p = 0.002-0.046). Manipulation of PARP-1 activity in parthanatos-competent cells reveals higher drug sensitivity in cells that have basal PARP-1 levels as compared with those subjected to PARP-1 overexpression or suppression. The same trends are observed in RNA expression databases and support the conclusion that PARP-1 can have optimal levels for favorable chemotherapeutic responses.

RNA Targeting in Acute Myeloid Leukemia.

Nucleosides and their analogues constitute an essential family of anticancer drugs. DNA has been the presumptive target of the front-line prodrug for acute myeloid leukemia (AML), cytarabine (ara-C), since the 1980s. Here, the biomolecular targeting of ara-C was evaluated in primary white blood cells using the ara-C mimic "AzC" and azide-alkyne "click" reactions. Fluorescent staining and microscopy revealed that metabolic incorporation of AzC into primary white blood cells was unexpectedly enhanced by the DNA polymerase inhibitor aphidicholine. According to RNaseH digestion and pull-down-and-release experiments, AzC was incorporated into short RNA fragments bound to DNA in peripheral blood monocytes (PBMCs) collected from all six healthy human donors tested. Samples from 22 AML patients (French-American-British classes M4 and M5) exhibited much more heterogeneity, with 27% incorporating AzC into RNA and 55% into DNA. The overall survival of AML patients whose samples incorporated AzC into RNA was approximately 3-fold higher as compared to that of the DNA cohort (p ≤ 0.056, χ2 = 3.65). These results suggest that the RNA primers of DNA synthesis are clinically favorable targets of ara-C, and that variable incorporation of nucleoside drugs into DNA versus RNA may enable future patient stratification into treatment-specific subgroups.

Superresolution imaging of individual replication forks reveals unexpected prodrug resistance mechanism.

Many drugs require extensive metabolism en route to their targets. High-resolution visualization of prodrug metabolism should therefore utilize analogs containing a small modification that does not interfere with its metabolism or mode of action. In addition to serving as mechanistic probes, such analogs provide candidates for theranostics when applied in both therapeutic and diagnostic modalities. Here a traceable mimic of the widely used anticancer prodrug cytarabine (ara-C) was generated by converting a single hydroxyl group to azide, giving "AzC." This compound exhibited the same biological profile as ara-C in cell cultures and zebrafish larvae. Using azide-alkyne "click" reactions, we uncovered an apparent contradiction: drug-resistant cells incorporated relatively large quantities of AzC into their genomes and entered S-phase arrest, whereas drug-sensitive cells incorporated only small quantities of AzC. Fluorescence microscopy was used to elucidate structural features associated with drug resistance by characterizing the architectures of stalled DNA replication foci containing AzC, EdU, γH2AX, and proliferating cell nuclear antigen (PCNA). Three-color superresolution imaging revealed replication foci containing one, two, or three partially resolved replication forks. Upon removing AzC from the media, resumption of DNA synthesis and completion of the cell cycle occurred before complete removal of AzC from genomes in vitro and in vivo. These results revealed an important mechanism for the low toxicity of ara-C toward normal tissues and drug-resistant cancer cells, where its efficient incorporation into DNA gives rise to highly stable, stalled replication forks that limit further incorporation of the drug, yet allow for the resumption of DNA synthesis and cellular division following treatment.

The novel microtubule targeting agent BAL101553 in combination with radiotherapy in treatment-refractory tumor models.

BACKGROUND AND PURPOSE: Resistance to microtubule targeting agents (MTA) represents a major drawback in successful cancer therapy with MTAs. Here we investigated the combined treatment modality of the novel MTA BAL101553 in combination with radiotherapy in paclitaxel and epothilone-resistant tumor models. MATERIAL AND METHODS: Multiple regimens of BAL101553, or its active moiety BAL27862 for in vitro experiments, were probed in combination with radiotherapy in P-glycoprotein-overexpressing, human colon adenocarcinoma cells (SW480) and in tubulin-mutated human NSCLC cells (A549EpoB40) and tumors thereof. RESULTS: BAL27862 reduced the proliferative activity of SW480 and A549EpoB40 tumor cells with similar potency as in A549 wildtype cells. Combined treatment of BAL27862 with ionizing radiation in vitro resulted in an additive reduction of clonogenicity. Moreover, treatment of paclitaxel- and epothilone-resistant tumors with fractionated irradiation and different regimens of BAL101553 (a single i.v. bolus vs. oral daily) suppressed tumor growth and resulted in an extended additive tumor growth delay with strong reduction of tumor proliferation and mean tumor vessel density. CONCLUSIONS: BAL101553 is a promising alternative in taxane- and epothilone-refractory tumors as part of a combined treatment modality with ionizing radiation. Its potent antitumor effect is not only tumor cell-directed but also targets the tumor microenvironment.