Cooperative cytotoxicity of proteasome inhibitors and tumor necrosis factor-related apoptosis-inducing ligand in chemoresistant Bcl-2-overexpressing cells.

PURPOSE: Bcl-2 overexpression is frequently detected in lymphoid malignancies, being associated with poor prognosis and reduced response to therapy. Here, we evaluated whether Bcl-2 overexpression affects the cytotoxic activity of proteasome inhibitors taken alone or in association with conventional anticancer drugs or tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). EXPERIMENTAL DESIGN: Jurkat cells engineered to overexpress Bcl-2 were treated with proteasome inhibitors (MG132, epoxomicin, and bortezomib), anticancer drugs (etoposide and doxorubicin), TRAIL, or combinations of these compounds. Cell death and loss of mitochondrial transmembrane potential were detected by flow cytometry. Cytosolic relocalization of cytochrome c and SMAC/Diablo, caspase cleavage, and Bcl-2 and Mcl-1 levels were determined by immunoblotting. Nuclear factor-kappaB inhibition was done by retroviral transduction with a dominant-negative mutant of IkappaBalpha. RESULTS: Bcl-2 overexpression results in significant inhibition of apoptosis in response to proteasome inhibitors, antiblastics, and TRAIL. Addition of TRAIL to proteasome inhibitors results in a synergistic cytotoxic effect in Bcl-2-overexpressing cells, whereas this result is not reproduced by the combination of proteasome inhibitors with antiblastic drugs. Importantly, proteasome inhibitors plus TRAIL induce mitochondrial dysfunction irrespective of up-regulated Bcl-2. Bcl-2 cleavage to a fragment with putative proapoptotic activity and elimination of antiapoptotic Mcl-1 may both play a role in proteasome inhibitors-TRAIL cooperation. Conversely, nuclear factor-kappaB inhibition by proteasome inhibitors is per se insufficient to explain the observed synergy. CONCLUSIONS: Combined proteasome inhibitors and TRAIL overcome the apoptotic threshold raised by Bcl-2 and may prove useful in the treatment of chemoresistant malignancies with up-regulated Bcl-2.     

Bioreversible quaternary N-acyloxymethyl derivatives of the tertiary amines bupivacaine and lidocaine—synthesis, aqueous solubility and stability in buffer, human plasma and simulated intestinal fluid

Design of water-soluble prodrugs may constitute a means to improve the oral bioavailability of drugs suffering from dissolution rate-limited absorption. The model drug bupivacaine containing a tertiary amine function has been converted into bioreversible quaternary N-acyloxymethyl derivatives. The pH-independent solubility of the N-butanoyloxymethyl derivate exceeded 1000 mg ml⁻¹ corresponding approximately to a 10,000-fold increase in water solubility compared to that of bupivacaine base. The kinetics of hydrolysis of the prodrugs was studied in the pH range 0.1–9.8 (37 °C). Decomposition was found to follow first-order kinetics and U-shaped pH-rate profiles were constructed. The observed differences between the hydrolytic lability of the derivatives might most likely be ascribed to steric effects. In most cases, the prodrugs were quantitatively converted into bupivacaine. However, for the hydrolysis of the N-butanoyloxymethyl derivative at neutral to slightly alkaline pH parallel formation of bupivacaine (80%) and an unknown compound X (20%) was observed. LC–MS analysis of the latter compound suggests that an aromatic imide structure has been formed from an intramolecular acyl transfer reaction involving a nucleophilic attack of the amide nitrogen atom on the ester carbonyl carbon atom. Whereas the derivatives were poor substrates for plasma enzymes; they were hydrolyzed rapidly to parent bupivacaine in the presence of pancreatic enzymes (simulated intestinal fluid) at 37 °C. The data indicate that such prodrugs possess sufficient stability in the acidic environment of the stomach to reach the small intestine in intact form where they can be cleaved efficiently by action of pancreatic enzymes prior to drug absorption. Thus, the N-acyloxymethyl approach might be of potential utility to enhance oral bioavailability of tertiary amines exhibiting pK_a values below approximately 6 and intrinsic solubilities in the low μM range.     

Increased proportions of B cells with spontaneous production of interleukin‐10 in HIV‐infected individuals are normalized during combination antiretroviral therapy: a longitudinal study

Interleukin‐10 (IL‐10)‐producing B cells (B10 cells) may inhibit HIV‐specific T cells and are elevated in untreated HIV infection. We aimed to determine the effect of combination antiretroviral treatment (cART) on the proportion of B10 cells. Furthermore, we compared B10‐cell proportions in HIV‐infected progressors and viremic controllers. This was a prospective study including HIV‐infected progressors, viremic controllers and healthy controls. Progressors initiating cART were followed for 6 months. Purified B cells were stimulated with CpG, alone or in combination with HIV gp120, and the proportion of B10 cells was measured by flow cytometry. Without stimulation, the B10‐cell proportion was higher in progressors than in healthy controls, while viremic controllers and healthy controls had comparable proportions. Moreover, the proportion of CD24^hiCD38^hi transitional B cells was higher in progressors than in healthy controls. After initiation of cART, the proportion of B10 cells and transitional B cells decreased. In conclusion, progressors had elevated B10‐cell proportions, while viremic controllers displayed normal proportions. After initiation of cART, the B10‐cell proportion decreased. This could limit B10‐cell‐mediated suppression of specific CD8⁺ T‐cell responses.     

A non‐fatal case of invasive zygomycete (Lichtheimia corymbifera) infection in an allogeneic haematopoietic cell transplant recipient

Post‐transplant infections in allogeneic haematopoietic cell transplant (allo‐HCT) recipients often have severe consequences. This is especially the case when dealing with zygomycete infections where the result is often fatal. A major problem when dealing with zygomycete infections is the need for an accurate and fast diagnosis as the phylum is highly resistant towards the conventional antifungals. We herein describe a non‐fatal case of Lichtheimia corymbifera infection in an allo‐HCT recipient.