Acute phase protein response and changes in lipoprotein particle size in dogs with systemic inflammatory response syndrome.

BACKGROUND: Improved methodology to measure acute phase proteins and determination of lipoprotein particle-size distribution (PSD) could be clinically useful in dogs with systemic inflammatory processes. OBJECTIVES: Evaluate an immunoturbidometric assay for serum amyloid A (SAA) and lipoprotein PSD in dogs with sepsis, nonseptic systemic inflammation, and in healthy controls. Correlate dyslipidemic changes with SAA and C-reactive protein (CRP) concentrations. ANIMALS: Twenty-five dogs with sepsis, 15 dogs with nonseptic systemic inflammation, and 22 healthy controls. METHODS: Prospective, case-control study. Variables included SAA, CRP, and electrophoretic subfractionation of high- and low-density lipoproteins (HDL, LDL). Continuous variables were compared using ANOVA or Kruskal-Wallis tests with linear regression or Spearman's rank correlation used to assess relationships between variables. RESULTS: Median SAA and CRP concentrations were greater in dogs with sepsis (SAA 460 mg/L, interquartile range [IQR] 886 mg/L; CRP 133.2 mg/L, IQR 91.6 mg/L) and nonseptic inflammation (SAA 201 mg/L, IQR 436 mg/L; CRP 91.1 mg/L, IQR 88.6 mg/L) compared to healthy dogs (SAA 0.0 mg/L, IQR 0.0 mg/L; CRP 4.9 mg/L, IQR 0.0 mg/L) P < .0001. A cutoff of >677.5 mg/L SAA was 43.2% sensitive and 92.3% specific for sepsis. Low-density lipoprotein was higher in dogs with sepsis 29.6%, (mean, SD 14.6) compared to 14.4% (mean, SD 5.6) of all lipoproteins in healthy controls (P = .005). High-density lipoprotein was not associated with CRP but was negatively correlated with SAA (rs -0.47, P < .0001). Subfractions of LDL and HDL differed between groups (all P < .05). CONCLUSIONS AND CLINICAL IMPORTANCE: Measurement of SAA using the immunoturbidometric assay evaluated in this study and lipoprotein PSD in dogs with inflammation might help distinguish septic from nonseptic causes of inflammation.

Cotreatment with BCL-2 antagonist sensitizes cutaneous T-cell lymphoma to lethal action of HDAC7-Nur77-based mechanism.

Pan-histone deacetylase inhibitors, for example, vorinostat and panobinostat (LBH589; Novartis Pharmaceuticals, East Hanover, NJ), have shown clinical efficacy against advanced cutaneous T-cell lymphoma (CTCL). However, the molecular basis of this activity remains unclear. HDAC7, a class IIA histone deacetylase (HDAC), is overexpressed in thymocytes, where it represses expression of the proapoptotic nuclear orphan receptor Nur77. Here, we demonstrate that treatment with panobinostat rapidly inhibits the in vitro and intracellular activity, as well as the mRNA and protein levels of HDAC7, and induces expression and translocation of Nur77 to the mitochondria. There, Nur77 converts death resistance protein Bcl-2 into a killer protein, promoting cell death of cultured and patient-derived human CTCL cells. Treatment with panobinostat improved survival of athymic nude mice implanted with human CTCL cells. Ectopic expression of Nur77 induced apoptosis and sensitized HH cells to panobinostat, whereas combined knockdown of Nur77 and its family member Nor1 was necessary to inhibit panobinostat-induced apoptosis of CTCL cells. Cotreatment with the Bcl-2/Bcl-x(L) antagonist ABT-737 decreased resistance and synergistically induced apoptosis of human CTCL cells. These findings mechanistically implicate HDAC7 and Nur77 in sensitizing human CTCL cells to panobinostat as well as suggest that cotreatment with an anti-Bcl-2 agent would augment the anti-CTCL activity of panobinostat.

Molecular and biologic characterization and drug sensitivity of pan-histone deacetylase inhibitor-resistant acute myeloid leukemia cells.

Hydroxamic acid analog pan-histone deacetylase (HDAC) inhibitors (HA-HDIs) have shown preclinical and clinical activity against human acute leukemia. Here we describe HA-HDI-resistant human acute myeloid leukemia (AML) HL-60 (HL-60/LR) cells that are resistant to LAQ824, vorinostat, LBH589, and sodium butyrate. HL-60/LR cells show increased expression of HDACs 1, 2, and 4 but lack HDAC6 expression, with concomitant hyperacetylation of heat shock protein 90 (hsp90). Treatment with HA-HDI failed to further augment hsp90 acetylation, or increase the levels of p21 or reactive oxygen species (ROSs), in HL-60/LR versus HL-60 cells. Although cross-resistant to antileukemia agents (eg, cytarabine, etoposide, and TRAIL), HL-60/LR cells are collaterally sensitive to the hsp90 inhibitor 17-AAG. Treatment with 17-AAG did not induce hsp70 or deplete the hsp90 client proteins AKT and c-Raf. HL-60/LR versus HL-60 cells display a higher growth fraction and shorter doubling time, along with a shorter interval to generation of leukemia and survival in nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice. Thus, resistance of AML cells to HA-HDIs is associated with loss of HDAC6, hyperacetylation of hsp90, aggressive leukemia phenotype, and collateral sensitivity to 17-AAG. These findings suggest that an hsp90 inhibitor-based antileukemia therapy may override de novo or acquired resistance of AML cells to HA-HDIs.

Cotreatment with vorinostat enhances activity of MK-0457 (VX-680) against acute and chronic myelogenous leukemia cells.

PURPOSE: We determined the effects of vorinostat (suberoylanalide hydroxamic acid) and/or MK-0457 (VX-680), an Aurora kinase inhibitor on the cultured human (HL-60, OCI-AML3, and K562) and primary acute myelogenous leukemia (AML) and chronic myelogenous leukemia (CML), as well as on the murine pro-B BaF3 cells with ectopic expression of the unmutated and mutant forms of Bcr-Abl. EXPERIMENTAL DESIGN: Following exposure to MK-0457 and/or vorinostat, apoptosis, loss of viability, as well as activity and levels of Aurora kinase and Bcr-Abl proteins were determined. RESULTS: Treatment with MK-0457 decreased the phosphorylation of Aurora kinase substrates including serine (S)10 on histone H3 and survivin, and led to aberrant mitosis, DNA endoreduplication as well as apoptosis of the cultured human acute leukemia HL-60, OCI-AML3, and K562 cells. Combined treatment with vorinostat and MK-0457 resulted in greater attenuation of Aurora and Bcr-Abl (in K562) kinase activity and levels as well as synergistically induced apoptosis of OCI-AML3, HL-60, and K562 cells. MK-0457 plus vorinostat also induced synergistic apoptosis of BaF3 cells with ectopic overexpression of wild-type or mutant Bcr-Abl. Finally, cotreatment with MK-0457 and vorinostat induced more loss of viability of primary AML and imatinib-refractory CML than treatment with either agent alone, but exhibited minimal toxicity to normal CD34+ progenitor cells. CONCLUSIONS: Combined in vitro treatment with MK-0457 and vorinostat is highly active against cultured and primary leukemia cells. These findings merit in vivo testing of the combination against human AML and CML cells, especially against imatinib mesylate-resistant Bcr-AblT315I-expressing CML Cells.