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Protein kinase D (PKD) is a member of a novel family of serine/threonine kinases that regulate fundamental cellular processes. PKD is implicated in the pathogenesis of several diseases, including cancer. Progress in understanding the biological functions and therapeutic potential of PKD has been hampered by the lack of specific inhibitors. The benzoxoloazepinolone CID755673 was recently identified as the first potent and selective PKD inhibitor. The study of structure-activity relationships (SAR) of this lead compound led to further improvements in PKD1 potency. We describe herein the synthesis and biological evaluation of novel benzothienothiazepinone analogues. We achieved a 10-fold increase in the in vitro PKD1 inhibitory potency for the second generation lead kb-NB142-70 and accomplished a transition to an almost equally potent novel pyrimidine scaffold, while maintaining excellent target selectivity. These promising results will guide the design of pharmacological tools to dissect PKD function and pave the way for the development of potential anticancer agents. © 2010 American Chemical Society.

Purpose: To evaluate the effectiveness of mitigation of acute ionizing radiation damage by mitochondrion-targeted small molecules. Methods and Materials: We evaluated the ability of nitroxide-linked alkene peptide isostere JP4-039, the nitric oxide synthase inhibitor-linked alkene peptide esostere MCF201-89, and the p53/mdm2/mdm4 protein complex inhibitor BEB55 to mitigate radiation effects by clonogenic survival curves with the murine hematopoietic progenitor cell line 32D cl 3 and the human bone marrow stromal (KM101) and pulmonary epithelial (IB3) cell lines. The p53-dependent mechanism of action was tested with p53+/+ and p53-/- murine bone marrow stromal cell lines. C57BL/6 NHsd female mice were injected i.p. with JP4-039, MCF201-89, or BEB55 individually or in combination, after receiving 9.5 Gy total body irradiation (TBI). Results: Each drug, JP4-039, MCF201-89, or BEB55, individually or as a mixture of all three compounds increased the survival of 32D cl 3 (p = 0.0021, p = 0.0011, p = 0.0038, and p = 0.0073, respectively) and IB3 cells (p = 0.0193, p = 0.0452, p = 0.0017, and p = 0.0019, respectively) significantly relative to that of control irradiated cells. KM101 cells were protected by individual drugs (p = 0.0007, p = 0.0235, p = 0.0044, respectively). JP4-039 and MCF201-89 increased irradiation survival of both p53+/+ (p = 0.0396 and p = 0.0071, respectively) and p53 -/- cells (p = 0.0007 and p = 0.0188, respectively), while BEB55 was ineffective with p53-/- cells. Drugs administered individually or as a mixtures of all three after TBI significantly increased mouse survival (p = 0.0234, 0.0009, 0.0052, and 0.0167, respectively). Conclusion: Mitochondrial targeting of small molecule radiation mitigators decreases irradiation-induced cell death in vitro and prolongs survival of lethally irradiated mice. © 2011 Elsevier Inc.

Fanconi anemia (FA) is an inherited disorder characterizedby defective DNA repair and cellular sensitivity to DNAcrosslinking agents. Clinically, FA is associated with highrisk for marrow failure, leukemia and head and necksquamous cell carcinoma (HNSCC). Radiosensitivity in FApatients compromises the use of total-body irradiation forhematopoietic stem cell transplantation and radiationtherapy for HNSCC. A radioprotector for the surroundingtissue would therefore be very valuable during radiotherapyfor HNSCC. Clonogenic radiation survival curves weredetermined for pre- or postirradiation treatment with theparent nitroxide Tempol or JP4-039 in cells of four FApatient-derived cell lines and two transgene-correctedsubclonal lines. FancG-/- (PD326) and FancD2-/- (PD20F)patient lines were more sensitive to the DNA crosslinkingagent mitomycin C (MMC) than their transgene-restoredsubclonal cell lines (both P , 0.0001). FancD2-/- cells weremore radiosensitive than the transgene restored subclonalcell line (n2.0 6 0.7 and 4.7 6 2.2, respectively, P 0.03).In contrast, FancG-/- cells were radioresistant relative to thetransgene-restored subclonal cell line (n 9.4 6 1.5 and 2.26 05, respectively, P0.001). DNA strand breaks measuredby the comet assay correlated with radiosensitivity. Celllines from a Fanc-C and Fanc-A patients showed radiosensitivitysimilar to that of Fanc-D2-/- cells. A fluorophoretaggedJP4-039 (BODIPY-FL) analog targeted the mitochondriaof the cell lines. Preirradiation or postirradiationtreatment with JP4-039 at a lower concentration thanTempol significantly increased the radioresistance andstabilized the antioxidant stores of all cell lines. Tempolincreased the toxicity of MMC in FancD2-/- cells. These dataprovide support for the potential clinical use of JP4-039 fornormal tissue radioprotection during chemoradiotherapy inFA patients. © 2011 by Radiation Research Society.

Chemical equations presented. JP4-039 is a novel nitroxide conjugate capable of crossing lipid bilayer membranes and scavenging reactive oxygen species (ROS). An efficient and scalable one-pot hydrozirconation- transmetalation-imine addition methodology has been developed for its asymmetric preparation. Furthermore, this versatile methodology allows for the synthesis of cyclopropyl and fluorinated analogs of the parent lead structure. © 2011 American Chemical Society.

Protein kinase D (PKD) belongs to a family of serine/threonine kinases that play an important role in basic cellular processes and are implicated in the pathogenesis of several diseases. Progress in our understanding of the biological functions of PKD has been limited due to the lack of a PKD-specific inhibitor. The benzoxoloazepinolone CID755673 was recently reported as the first potent and kinase-selective inhibitor for this enzyme. For structure-activity analysis purposes, a series of analogs was prepared and their in vitro inhibitory potency evaluated. © 2011 by the authors; licensee MDPI, Basel, Switzerland.

Purpose: To evaluate the effectiveness of mitigation of acute ionizing radiation damage by mitochondrion-targeted small molecules. Methods and Materials: We evaluated the ability of nitroxide-linked alkene peptide isostere JP4-039, the nitric oxide synthase inhibitor-linked alkene peptide esostere MCF201-89, and the p53/mdm2/mdm4 protein complex inhibitor BEB55 to mitigate radiation effects by clonogenic survival curves with the murine hematopoietic progenitor cell line 32D cl 3 and the human bone marrow stromal (KM101) and pulmonary epithelial (IB3) cell lines. The p53-dependent mechanism of action was tested with p53+/+ and p53-/- murine bone marrow stromal cell lines. C57BL/6 NHsd female mice were injected i.p. with JP4-039, MCF201-89, or BEB55 individually or in combination, after receiving 9.5 Gy total body irradiation (TBI). Results: Each drug, JP4-039, MCF201-89, or BEB55, individually or as a mixture of all three compounds increased the survival of 32D cl 3 (p = 0.0021, p = 0.0011, p = 0.0038, and p = 0.0073, respectively) and IB3 cells (p = 0.0193, p = 0.0452, p = 0.0017, and p = 0.0019, respectively) significantly relative to that of control irradiated cells. KM101 cells were protected by individual drugs (p = 0.0007, p = 0.0235, p = 0.0044, respectively). JP4-039 and MCF201-89 increased irradiation survival of both p53+/+ (p = 0.0396 and p = 0.0071, respectively) and p53 -/- cells (p = 0.0007 and p = 0.0188, respectively), while BEB55 was ineffective with p53-/- cells. Drugs administered individually or as a mixtures of all three after TBI significantly increased mouse survival (p = 0.0234, 0.0009, 0.0052, and 0.0167, respectively). Conclusion: Mitochondrial targeting of small molecule radiation mitigators decreases irradiation-induced cell death in vitro and prolongs survival of lethally irradiated mice. © 2011 Elsevier Inc.

Chemical equations presented. JP4-039 is a novel nitroxide conjugate capable of crossing lipid bilayer membranes and scavenging reactive oxygen species (ROS). An efficient and scalable one-pot hydrozirconation- transmetalation-imine addition methodology has been developed for its asymmetric preparation. Furthermore, this versatile methodology allows for the synthesis of cyclopropyl and fluorinated analogs of the parent lead structure. © 2011 American Chemical Society.

Protein kinase D (PKD) is a member of a novel family of serine/threonine kinases that regulate fundamental cellular processes. PKD is implicated in the pathogenesis of several diseases, including cancer. Progress in understanding the biological functions and therapeutic potential of PKD has been hampered by the lack of specific inhibitors. The benzoxoloazepinolone CID755673 was recently identified as the first potent and selective PKD inhibitor. The study of structure-activity relationships (SAR) of this lead compound led to further improvements in PKD1 potency. We describe herein the synthesis and biological evaluation of novel benzothienothiazepinone analogues. We achieved a 10-fold increase in the in vitro PKD1 inhibitory potency for the second generation lead kb-NB142-70 and accomplished a transition to an almost equally potent novel pyrimidine scaffold, while maintaining excellent target selectivity. These promising results will guide the design of pharmacological tools to dissect PKD function and pave the way for the development of potential anticancer agents. © 2010 American Chemical Society.

Protein kinase D (PKD) belongs to a family of serine/threonine kinases that play an important role in basic cellular processes and are implicated in the pathogenesis of several diseases. Progress in our understanding of the biological functions of PKD has been limited due to the lack of a PKD-specific inhibitor. The benzoxoloazepinolone CID755673 was recently reported as the first potent and kinase-selective inhibitor for this enzyme. For structure-activity analysis purposes, a series of analogs was prepared and their in vitro inhibitory potency evaluated. © 2011 by the authors; licensee MDPI, Basel, Switzerland.

Fanconi anemia (FA) is an inherited disorder characterizedby defective DNA repair and cellular sensitivity to DNAcrosslinking agents. Clinically, FA is associated with highrisk for marrow failure, leukemia and head and necksquamous cell carcinoma (HNSCC). Radiosensitivity in FApatients compromises the use of total-body irradiation forhematopoietic stem cell transplantation and radiationtherapy for HNSCC. A radioprotector for the surroundingtissue would therefore be very valuable during radiotherapyfor HNSCC. Clonogenic radiation survival curves weredetermined for pre- or postirradiation treatment with theparent nitroxide Tempol or JP4-039 in cells of four FApatient-derived cell lines and two transgene-correctedsubclonal lines. FancG-/- (PD326) and FancD2-/- (PD20F)patient lines were more sensitive to the DNA crosslinkingagent mitomycin C (MMC) than their transgene-restoredsubclonal cell lines (both P , 0.0001). FancD2-/- cells weremore radiosensitive than the transgene restored subclonalcell line (n2.0 6 0.7 and 4.7 6 2.2, respectively, P 0.03).In contrast, FancG-/- cells were radioresistant relative to thetransgene-restored subclonal cell line (n 9.4 6 1.5 and 2.26 05, respectively, P0.001). DNA strand breaks measuredby the comet assay correlated with radiosensitivity. Celllines from a Fanc-C and Fanc-A patients showed radiosensitivitysimilar to that of Fanc-D2-/- cells. A fluorophoretaggedJP4-039 (BODIPY-FL) analog targeted the mitochondriaof the cell lines. Preirradiation or postirradiationtreatment with JP4-039 at a lower concentration thanTempol significantly increased the radioresistance andstabilized the antioxidant stores of all cell lines. Tempolincreased the toxicity of MMC in FancD2-/- cells. These dataprovide support for the potential clinical use of JP4-039 fornormal tissue radioprotection during chemoradiotherapy inFA patients. © 2011 by Radiation Research Society.