MEK inhibition enhances paclitaxel-induced tumor apoptosis.

The anti-cancer drug paclitaxel (Taxol) alters microtubule assembly and activates pro-apoptotic signaling pathways. Previously, we and others found that paclitaxel activates endogenous JNK in tumor cells, and the activation of JNK contributes to tumor cell apoptosis. Here we find that paclitaxel activates the prosurvival MEK/ERK pathway, which conversely may compromise the efficacy of paclitaxel. Hence, a combination treatment of paclitaxel and MEK inhibitors was pursued to determine whether this treatment could lead to enhanced apoptosis. The inhibition of MEK/ERK with a pharmacologic inhibitor, U0126, together with paclitaxel resulted in a dramatic enhancement of apoptosis that is four times more than the additive value of the two drugs alone. Enhanced apoptosis was verified by the terminal transferase-mediated dUTP nick end labeling assay, by an enzyme-linked immunosorbent assay for histone-associated DNA fragments, and by flow cytometric analysis for DNA content. Specificity of the pharmacologic inhibitor was confirmed by the use of (a) a second MEK/ERK inhibitor and (b) a transdominant-negative MEK. Enhanced apoptosis was verified in breast, ovarian, and lung tumor cell lines, suggesting this effect is not cell type-specific. This is the first report of enhanced apoptosis detected in the presence of paclitaxel and MEK inhibition and suggests a new anticancer strategy.

Paclitaxel up-regulates interleukin-8 synthesis in human lung carcinoma through an NF-kappaB- and AP-1-dependent mechanism.

Lung cancer is a leading cause of cancer-related death in the United States. For this reason we chose to study the specific cellular effects that one chemotherapeutic agent, paclitaxel, has on lung carcinoma. In addition to its known mechanism of action, which is to stabilize microtubules, paclitaxel has been shown to have other interesting and relevant cellular effects. In this report, we demonstrate that a subset of human lung carcinoma cell lines respond to paclitaxel treatment with an up to a fivefold increase in the production of interleukin-8 (IL-8). We demonstrate that this increased production is specific to IL-8 but not to other chemokines, and is both dose- and time-dependent. Increased IL-8 mRNA is seen as early as 45 min with a peak at 4 h after paclitaxel treatment. This increase in mRNA is due to transcriptional activation because actinomycin D treatment blocked the increase. Paclitaxel also activates the mitogen-activated protein kinase family member, JNK1, in dose-dependent fashion. IL-8 enhancement is completely abolished with the use of an inhibitor of NF-kappaB, the super-repressor IkappaB. Similar results were obtained upon the inhibition of AP-1 activation with the MEK1/2 inhibitor, U0126. By gaining a better understanding of the differences in cellular response to paclitaxel chemotherapy, these findings might lead to either improved patient selection or to the development of adjuvant therapy targeted at specific-cell signaling proteins.

Group B streptococcal colonization in a developing country: its association with sexually transmitted disease and socioeconomic factors.

Group B Streptococcus (GBS) is an important infectious organism in pregnant women and their neonates. Although excellent data are available from the developing world, little epidemiologic information is available from Latin America. To evaluate the prevalence of GBS colonization in a developing country, a prospective study was performed in Lima, Peru. We found a relatively low prevalence of GBS colonization of 6.0% in parturient women and 10.6% in nonpregnant women. No association of GBS colonization was made with previously identified risk factors such as age, parity, or birth control practices. We did find a positive association between GBS colonization and chlamydial carriage (P < 0.05). We also report an even distribution of GBS serotypes: Ia/c = 35%, IIc = 18%, III = 29%, and V = 18%. Our study provides evidence for a low prevalence of GBS maternal carriage in this urban Latin American population.

Hemolytically inactive C4B complement allotype caused by a proline to leucine mutation in the C5-binding site.

The fourth component of complement (C4) is encoded by two highly homologous genes, C4A and C4B. Only one hemolytically inactive C4A allotype (C4A6) has been reported. No hemolytically inactive C4B allotype has been described. We report the first hemolytically inactive (hi) allotype of C4B, C4B*1 (hi). This unique variant was first recognized by hemolytic overlay assays and confirmed to segregate in the affected pedigree with the major histocompatibility complex haplotype A28,B35,CW4,DR6, C4A3,C4B1(hi), BFF,C2C. By single strand conformational polymorphism, we detected only a migration variant in exon 12 caused by a C to T transition in the second base of codon 459. This mutation results in a leucine substitution for proline (P459L) 1 residue downstream of a residue known to contribute to the C5-binding site. Allele-specific oligonucleotide analysis of samples demonstrated cosegregation of the mutation with the hemolytically inactive allotype in the affected pedigree. Site-directed mutagenesis and expression studies showed that the P459L mutation causes loss of hemolytic function. C4B*1(hi) is the first example of a circulating C4B protein lacking detectable hemolytic activity and the P459L mutation expands our knowledge of the C5-binding site of C4.

Soluble human complement receptor type 1 inhibits complement-mediated host defense.

Soluble complement receptor type 1 (sCR1) is a powerful inhibitor of complement activation. Because of this ability, sCR1 may prove to be an important therapeutic agent that can be used to block the immunopathologic effects of uncontrolled complement activation in a variety of clinically significant disorders. Although several previous studies have examined the ability of sCR1 to inhibit complemented-mediated immunopathologic damage, there is no information on its ability to interfere with the host's defense against infection. In the current experiments sCR1 exerted a concentration-dependent inhibitory effect on the phagocytosis of Streptococcus pneumoniae by human polymorphonuclear leukocytes in vitro. Not only di sCR1 inhibit complement-dependent opsonization of the pneumococcus but at higher concentrations it also inhibited the ingestion of bacteria which had been previously opsonized. Furthermore, when rats were injected with sCR1, it inhibited both their serum hemolytic activity and serum opsonic activity in a dose-dependent fashion. Finally, for rats treated with sCR1, the 50% lethal dose was S. pneumoniae and Pseudomonas aeruginosa. These data demonstrate that sCR1 significantly inhibits complement-mediated host against bacterial infection.

Neonatal tetanus in Peru: risk assessment with modified enzyme-linked immunosorbent assay and toxoid skin test.

We used a modified enzyme-linked immunosorbent assay (ELISA) to investigate tetanus immunity in 232 pregnant Peruvian women. One hundred forty-two (61.2%) had protective antitoxin titers (> or = 0.01 IU/mL). Protective titers correlated positively with the number of toxoid doses reported during the current pregnancy. A majority of women reporting no toxoid doses during the current pregnancy had at least one prenatal health care visit. We evaluated a toxoid skin test in 44 of the subjects, but it correlated poorly with the ELISA. The modified ELISA is a useful in vitro method for studying tetanus immunity in the developing world.