To overcome proteotoxic stress inherent to malignant transformation, cancer cells induce a range of adaptive mechanisms, with the master transcription factor heat-shock factor 1 (HSF1)-orchestrated response taking center stage. Here we define a novel gain-of-function of mutant p53 (mutp53), whereby mutp53-overexpressing cancer cells acquire superior tolerance to proteotoxic stress. mutp53 via constitutive stimulation of EGFR and ErbB2 signaling hyperactivates the MAPK and PI3K cascades, which induce stabilization and phosphoactivation of HSF1 on Ser326. Moreover, mutp53 protein via direct interaction with activated p-Ser326 HSF1 facilitates HSF1 recruitment to its specific DNA-binding elements and stimulates transcription of heat-shock proteins including Hsp90. In turn, induced Hsp90 stabilizes its oncogenic clients including EGFR, ErbB2 and mutp53, thereby further reinforcing oncogenic signaling. Thus, mutp53 initiates a feed forward loop that renders cancer cells more resistant to adverse conditions, providing a strong survival advantage.

PTX is a potent and efficacious inhibitor of TLR-mediated inflammatory cytokines in newborn cord blood and a promising neonatal anti-inflammatory agent.

IntroductionNeonatal inflammation, mediated in part through Toll-like receptor (TLR) and inflammasome signaling, contributes to adverse outcomes including organ injury. Pentoxifylline (PTX), a phosphodiesterase inhibitor which potently suppresses cytokine production in newborn cord blood, is a candidate neonatal anti-inflammatory agent. We hypothesized that combinations of PTX with other anti-inflammatory agents, the steroid dexamethasone (DEX) or the macrolide azithromycin (AZI), may exert broader, more profound and/or synergistic anti-inflammatory activity towards neonatal TLR- and inflammasome-mediated cytokine production.MethodsWhole newborn and adult blood was treated with PTX (50–200 μM), DEX (10−10–10−7 M), or AZI (2.5–20 μM), alone or combined, and cultured with lipopolysaccharide (LPS) (TLR4 agonist), R848 (TLR7/8 agonist) or LPS/adenosine triphosphate (ATP) (inflammasome induction). Supernatant and intracellular cytokines, signaling molecules and mRNA were measured by multiplex assay, flow cytometry and real-time PCR. Drug interactions were assessed based on Loewe's additivity.ResultsPTX, DEX and AZI inhibited TLR- and/or inflammasome-mediated cytokine production in newborn and adult blood, whether added before, simultaneously or after TLR stimulation. PTX preferentially inhibited pro-inflammatory cytokines especially TNF. DEX inhibited IL-10 in newborn, and TNF, IL-1β, IL-6 and interferon-α in newborn and adult blood. AZI inhibited R848-induced TNF, IL-1β, IL-6 and IL-10, and LPS-induced IL-1β and IL-10. (PTX+DEX) synergistically decreased LPS- and LPS/ATP-induced TNF, IL-1β, and IL-6, and R848-induced IL-1β and interferon-α, while (PTX+AZI) synergistically decreased induction of TNF, IL-1β, and IL-6. Synergistic inhibition of TNF production by (PTX+DEX) was especially pronounced in newborn vs. adult blood and was accompanied by reduction of TNF mRNA and enhancement of IL10 mRNA.ConclusionsAge, agent, and specific drug-drug combinations exert distinct anti-inflammatory effects towards TLR- and/or inflammasome-mediated cytokine production in human newborn blood in vitro. Synergistic combinations of PTX, DEX and AZI may offer benefit for prevention and/or treatment of neonatal inflammatory conditions while potentially limiting drug exposure and toxicity.

Neonatal sepsis and its accompanying inflammatory response contribute to substantial morbidity and mortality. Pentoxifylline (PTX), a phosphodiesterase inhibitor which suppresses transcription and production of proinflammatory cytokines, is a candidate adjunctive therapy for newborn sepsis.

Introduction: Neonatal sepsis triggers an inflammatory response that contributes to mortality and multiple organ injury. Pentoxifylline (PTX), a phosphodiesterase inhibitor which suppresses pro-inflammatory cytokines, is a candidate adjunctive therapy for newborn sepsis. We hypothesized that administration of PTX in addition to antibiotics decreases live bacteria-induced pro-inflammatory and/or enhances anti-inflammatory cytokine production in septic neonatal mice without augmenting bacterial growth. Methods: Newborn C57BL/6J mice (<24 h old) were injected intravenously with 10 5 colony forming units (CFUs)/g weight of a bioluminescent derivative of the encapsulated clinical isolate Escherichia coli O18:K1. Adequacy of intravenous injections was validated using in vivo bioluminescence imaging and Evans blue. Pups were treated with gentamicin (GENT), PTX, (GENT + PTX) or saline at 0, 1.5, or 4 h after sepsis initiation, and euthanized after an additional 4 h. CFUs and cytokines were measured from blood and homogenized organ tissues. Results: GENT alone inhibited bacterial growth, IL-1β, and IL-6 production in blood and organs. Addition of PTX to GENT profoundly inhibited E. coli-induced TNF and enhanced IL-10 in blood of newborn mice at all timepoints, whereas it primarily upregulated IL-10 production in peripheral organs (lung, spleen, brain). PTX, whether alone or adjunctive to GENT, did not increase microbial colony counts in blood and organs. Conclusion: Addition of PTX to antibiotics in murine neonatal E. coli sepsis promoted an anti-inflammatory milieu through inhibition of plasma TNF and enhancement of IL-10 production in plasma and organs without increasing bacterial growth, supporting its utility as a potential adjunctive agent for newborn sepsis.