[The immunological basis of the administration of DTP-polio vaccine]. / De immunologische basis van de toepassing van DKTP-vaccin.

The main subject of this essay is the analysis of the immunizing properties and possible side effects of DTP (whole cell)-polio vaccine, which since 1964 is the backbone of the immunization programme in the Netherlands. The concept, that the basis of the immunity conferred by diphtheria and tetanus toxoid and inactivated poliovirus is humoral, is now generally accepted. Assessment of the potency of these three components is based on their binding properties to specific immunoglobulines in standard-antisera. An example is the testing of neutralising antibodies against the three types of poliovirus in sera obtained from different groups of infants injected with increasing doses of this vaccine. Side-effects after injecting diphtheria- and tetanus toxoids and three types inactivated poliovaccine are negligible. For the pertussis component of the vaccine, this is not the case. The standardization of the potency of cellular pertussis vaccine is still based on the analysis of the protective capacity of a number of plain vaccine lots in the intercerebral mouse-protection test relative to the protection they confer in children. An international standard vaccine is available for this purpose, and the potency of individual lots can be expressed in International Units. At present acellular pertussis vaccines are also available for immunization against this disease. The components of such vaccines are at least for a number of components, arbitrarily chosen. Recently in a trial in Sweden, a five-component pertussis vaccine has given a similar protection of infants as a cellular vaccine of British origin. In addition this acellular vaccine induced less febrile reactions. However, acellular vaccines do not protect mice against intracerebral infection. because producers have decided to omit the "Outer Membrane Complex (OMC) as a component of the vaccine. In addition it has recently been shown that the presence of the cellular pertussis component in DPT vaccine will almost completely suppress tetanus antitoxins in the IgE fraction of the serum. This observation could indicate that this suppression is the result of a cellular immunity reaction by components in the bacteria.

Pharmacologic reversal of pertussis toxin-induced thermal allodynia in mice.

We have previously demonstrated that the intrathecal administration of pertussis toxin produces a long-lasting thermal allodynia in mice. The purpose of the present studies was to compare the antinociceptive and the antiallodynic effects of drugs that are commonly used in treating neuropathic allodynia in untreated mice and in mice which had been administered vehicle or pertussis toxin intrathecally 7 days previously. In untreated mice, morphine, fentanyl, clonidine, oxymetazoline, desipramine and lidocaine, but not MK801, produced dose-related antinociception when tested using a 55 degrees C water tail-flick test. However, 7 days after the intrathecal injection of pertussis toxin, which induced a condition of thermal allodynia when tested using a 45 degrees C water bath, the full opioid and the full alpha(2)-adrenergic receptor agonists fentanyl and clonidine, but not the partial opioid nor the partial alpha(2)-adrenergic receptor agonists morphine and oxymetazoline, reversed the pertussis toxin-induced thermal allodynia. Moreover, lidocaine, desipramine, carbamazepine and MK801 failed to reverse the pertussis toxin-induced thermal allodynia. The present results suggest that decrements in G(i)/G(o)-protein function may be involved in initiating and/or maintaining some neuropathic pain states. Moreover, the results of the present study suggest that the use of full, but not partial, opioid or alpha(2)-agonists may be useful in the treatment of thermal allodynic pain states which may be due at least in part to inhibitory second messenger system dysfunction. Further, the underlying biochemistry of the apparent allodynic pain state induced by intrathecal administration of pertussis toxin warrants further investigation.

Tumor cell motility. A novel therapeutic target in bladder carcinoma, experimental and clinical results.

Surgery, chemotherapy and radiotherapy are the current modalities of tumor management. However, in systemic disease, predicted patients' cure can be achieved in but a few tumor diseases. Based on previous basic research we have highlighted that the loss of cell-cell adhesion in association with an increased tumor cell motility is an essential feature of the malignant potential of bladder tumors. Thus, we have attempted therapeutical methods differing from hitherto existing treatments by focusing on a tumor cell function we call cell motility. Characterization of so-called anti-motility drugs was performed biochemically as well analyzed by in vitro by using in established bladder carcinoma cell lines. We evaluated the potential therapeutic benefit in a model of chemically induced bladder carcinoma followed by a phase I/II trial applying anti-motility drugs in patients which were chemotherapy-resistant and having metastatic bladder cancer. Both basic research as well as the results of first translational clinical trials confirmed, that advanced bladder carcinomas can be favorably affected by inhibition of tumor cell motility.

Whole-cell and acellular pertussis vaccines.

OBJECTIVE: To provide a review of pertussis vaccines, including information on efficacy, adverse reactions, and antibody production following administration of both whole-cell and acellular pertussis vaccines. DATA SOURCES: A MEDLINE search and extensive review of journals was conducted to identify the information for this review. DATA EXTRACTION: Pertinent studies reporting experience with pertussis vaccinations were reviewed. DATA SYNTHESIS: The differences in efficacy, adverse reactions, and antibody responses between whole-cell and acellular pertussis vaccines are emphasized. The status of acellular pertussis vaccination in the US is defined. CONCLUSIONS: Acellular (chemically detoxified or recombinant) pertussis vaccine formulation appears to cause fewer adverse reactions than whole-cell vaccine in most studies. Clinical efficacy and safety in the very young has not been well established. Thus, acellular pertussis vaccine is reserved for the 4th and 5th doses in the US. Oral or intranasal formulations of the pertussis vaccine are being evaluated.

Severe reactions associated with diphtheria-tetanus-pertussis vaccine: detailed study of children with seizures, hypotonic-hyporesponsive episodes, high fevers, and persistent crying.

OBJECTIVE: The pathophysiology of severe reactions to diphtheria-tetanus-pertussis (DTP)vaccine is not well understood. Active pertussis toxin in DTP vaccine has been proposed to cause severe DTP vaccine reactions. Large doses of pertussis toxin cause hyperinsulinemia and hypoglycemia as well as leukocytosis with a predominant lymphocytosis in animal models. To learn more about the causes of and risk factors for severe DTP vaccine reactions, children experiencing severe DTP vaccine reactions were studied. DESIGN: Prospective, referral-based surveillance. SETTING: Los Angeles, CA. SUBJECTS: Children experiencing severe reactions within 48 hours of DTP immunization and evaluated within 24 hours of the reaction. Severe reactions included encephalopathy, persistent crying > or = 3 hours, hypotonic-hyporesponsive episodes (collapse episodes), fever > or = 40.5 degrees C, or seizures. Some comparisons were made between children with DTP vaccine-associated seizures and a comparison group of children experiencing febrile seizures unrelated to immunization. OUTCOME MEASURES: A history and physical examination were performed. Follow-up examinations were performed 1 month later. Blood was collected for complete blood cell count with leukocyte differential count, serum chemistry measurements, and insulin and glucose values. Serum was assayed for active pertussis toxin, both in free and immune-complex masked states. RESULTS: Sixty children experienced severe reactions within 48 hours of DTP immunization: 32 children had seizures only, 14 subjects had hypotonic-hyporesponsive episodes, 2 subjects had fever > or = 40.5 degrees C only, 4 subjects had persistent crying > or = 3 hours, 6 children had seizures and fever > or = 40.5 degrees C, and 2 children had persistent crying and seizures. The children with seizures had a high rate of personal and family histories of seizures, and 90% had documented fevers (> or = 38 degrees C). Persistent crying was associated with painful local reactions. Effects that may have been due to vaccine pertussis toxin were not found. Lymphocytosis did not occur, nor did hypoglycemia. Some relatively elevated insulin values were noted; however, this finding was also noted in the comparison group of children experiencing febrile seizures unrelated to immunization. No biologically active pertussis toxin was found in the acute sera of children experiencing severe DTP vaccine reactions. CONCLUSIONS: Seizures associated with DTP vaccine have similar clinical characteristics as febrile seizures, and persistent crying is initiated by painful local reactions. Vaccine endotoxin is a cause of febrile DTP vaccine reactions. We found no evidence that DTP vaccine pertussis toxin plays a role in severe DTP vaccine reactions.

Sensitization to psychostimulants and stress after injection of pertussis toxin into the A10 dopamine region.

An augmentation of psychostimulant-induced motor activity, termed sensitization, occurs with daily treatment and can last for months or years. At least in part, sensitization results from a long-term change in mesocorticolimbic dopamine transmission and may involve a disinhibition of dopamine neurons. Dopamine D2 autoreceptors and gamma-aminobutyric acidB (GABAB) receptors provide tonic inhibition to dopamine neurons via a G protein-mediated increase in K+ efflux. To evaluate the role of these inhibitory mechanisms in sensitization, pertussis toxin (PTX) was injected into the A10 dopamine region to uncouple the receptors via ADP-ribosylation of G proteins. In this study we demonstrated a significant augmentation in cocaine-stimulated motor activity, at doses greater than 3.0 mg/kg, 14 days after intra-A10 injection of PTX. Also, amphetamine-, but not morphine- or caffeine-stimulated motor activity was significantly augmented 2 weeks after PTX pretreatment. In vivo microdialysis revealed an augmentation of cocaine-induced increases in extracellular dopamine in the nucleus accumbens 14 days after PTX pretreatment. Pretreatment in the A10 region with the GABAB agonist baclofen, blocked cocaine-stimulated motor activity in control animals, but not in PTX-pretreated animals, indicating that the PTX treatment had uncoupled the GABAB receptor. Footshock stress activates mesocortical dopamine transmission, and postmortem tissue levels of dihydroxyphenylacetic acid and homovanillic acid in the prefrontal cortex were increased in PTX-pretreated animals. We hypothesize that the sensitized responses to cocaine, amphetamine and stress produced by PTX results from a decrease in dopamine D2 and GABAB-mediated inhibitory control of A10 dopamine neurons.

Metabolic, humoral, and cellular responses in adult volunteers immunized with the genetically inactivated pertussis toxin mutant PT-9K/129G.

PT-9K/129G, a nontoxic mutant of pertussis toxin (PT) obtained by genetic manipulation, has been shown in animal models to be a promising candidate for new vaccines against whooping cough. To assess the safety and the immunogenicity of PT-9K/129G in humans, a pilot study has been performed in adult volunteers. The protein was found to be safe, capable of inducing high titers of toxin-neutralizing antibodies, and capable of generating immunological memory. In fact, vaccination caused an increase of cell-mediated response to PT, PT-9K/129G, S1 subunit, and B oligomer, indicating that memory T cells are induced by the vaccine. Since PT-9K/129G is mitogenic for T lymphocytes in vitro, it was investigated whether this activity is also present in vivo. No variation was observed in the proportion of T cells (CD3+), T helper cells (CD4+), and cytotoxic T cells (CD8+), as well as in that of other lymphoid populations, by FACS analysis. Interestingly, no thorough correlation was found between humoral and cellular responses. In one case, a very high cellular response was present in absence of detectable antibodies, suggesting that the antibody response, which is the only parameter measured in most clinical trials, may not give a complete picture of the response induced by a vaccine.

Immunogenicity and safety of a pertussis vaccine composed of pertussis toxin inactivated by hydrogen peroxide, in 18- to 23-month-old children.

A new pertussis vaccine, composed of purified pertussis toxin inactivated by hydrogen peroxide and adsorbed onto aluminum hydroxide (NICHD-Ptxd), was injected into 60 children aged 18 to 23 months without a history of pertussis or pertussis vaccination. Two doses of toxoid, 10 and 50 micrograms, were used. Two injections, given 8 to 12 weeks apart, elicited increases in serum levels of antitoxin and IgG antibodies in 56 children who had no detectable antitoxin (less than 5 units) before vaccination. Four children with detectable antitoxin (greater than or equal to 5 units) before the first vaccination had pronounced antibody increases after the first dose. After the second dose, the geometric mean antitoxin concentration was 29 units with the 50 micrograms dosage and 10 units with the 10 micrograms dosage (p less than 0.001). Serum antibody levels elicited by two injections of 50 micrograms were similar to those in patients convalescing from pertussis. A third injection given to seven children 9 to 10 months after the second injection gave a booster response, with high levels of antitoxin (160 to 1280 units) and of IgG antibodies. With few exceptions the antibody response was restricted to the IgG class. Transient local reactions greater than or equal to 2 cm in diameter occurred in 14% of the children after the first dose and in 44% after the second and third doses. Moderate fever was recorded after 6% of all injections. There were no changes in peripheral blood leukocyte counts or fasting blood glucose levels measured before and 24 hours after the first injection. We conclude that NICHD-Ptxd is immunogenic in children. No serious adverse effects were noted.

Characterization and clinical study on the acellular pertussis vaccine produced by a combination of column purified pertussis toxin and filamentous hemagglutinin.

Bordetella Pertussis Tohama phase I was cultured in a 300-liter fermentor using a medium containing 0.1% heptakis (2,5-0-dimethyl) beta-cyclodextrin (MeCD). Pertussis toxin (PT) and filamentous hamagglutinin (FHA) were purified using affinity and ion exchange gel column chromatographies. Endotoxin contents of these antigens (10 micrograms PN/ml) were less than 10 ngLPS/ml. PT and FHA were independently treated with formalin in the presence of amino acid and were mixed at a protein concentration ratio of 1:4, the same ratio of our commercialized acellular pertussis vaccine. PDT vaccine containing 2 micrograms PN of PT and 8 micrograms PN of FHA per milliliter was prepared. This PDT vaccine satisfied all the items of the Japanese Minimum Requirements including potency and toxicity tests. Even after this vaccine was incubated for 4 weeks at 37 degrees C, no deaths of the inoculated mice were observed after challenge with 4 mg of histamine on the 4th and 12th day of the inoculation. Compared with the conventional vaccine, this new vaccine caused less swelling in the mouse footpad test. A field trial of our two vaccines, one manufactured by the conventional method (lot No. 21A) and the other produced by the new method (lot No. KC8702), revealed that children receiving KC8702 showed almost the same anti-PT and anti-FHA antibody levels as those given 21A. Those who received KC8702 suffered from less local side effects such as redness, swelling or induration than those given 21A. Our new method for the production of acellular pertussis vaccine permits us the economical manufacturing of the vaccine with uniform quality in a closed system.

Synergistic teratogenic effect produced in mice by whole cell pertussis vaccine.

Pertussis whole cell bacterial vaccine was injected in mice during early pregnancy to disclose any teratogenic effect on the brain of the fetuses. Cytochalasin D by itself induced exencephaly in a dose dependent way in fetal mice. When pregnant mice received a single injection of pertussis vaccine on day 8 of gestation and a subteratogenic dose of cytochalasin D on days 8, 9 and 10 of gestation a synergistic teratogenic action of pertussis vaccine and cytochalasin D in mice was observed. When autopsy was performed after a further 9 to 10 days a significant number of brain malformations was found. In order to analyse which component in the vaccine might be responsible for the co-teratogenic effect, purified pertussis components, pertussis toxin and filamentous haemagglutinin were used in combination with cytochalasin D, but no malformations occurred. The same results were obtained by using diphtheria-tetanus-polio (DiTePol) vaccine and acellular pertussis component vaccine, whereas the use of whole cell typhoid vaccine resulted in a high rate of fetuses with exencephalies. Experiments with purified Bordetella pertussis and Escherichia coli lipopolysaccharides indicated that lipopolysaccharides in whole cell pertussis vaccine as well as in typhoid vaccine were the factors causing teratogenicity in fetal mice.

Bacterial toxins and the role of ADP-ribosylation.

Studies on the pathogenesis of "Whooping Cough" and cholera have resulted in the discovery of important pathways in the regulation of cellular metabolism leading to the realization of a complex family of proteins that appear to play central roles in the regulation of hormonal responses and which utilize guanine nucleotides in their mechanism of action. The fact that these bacterial toxins interfere so precisely with the complex regulation of eukaryotic cellular metabolism and the discovery of analogous enzymes within the cytosol of eukaryotic cells suggests that ADP-ribosylation may be an important pathway through which the cell can establish its responsiveness to its environment. Clearly, future work directed towards the role of ADP-ribosylation and towards the mechanisms of the regulation of these endogenous ADP-ribosyltransferases and lyases may provide great insights into the mechanisms of hormone action.