Towards novel paradigms for cancer therapy.

Cancer is a complex progressive multistep disorder that results from the accumulation of genetic and epigenetic abnormalities, which lead to the transformation of normal cells into malignant derivatives. Despite enormous progress in the understanding of cancer biology including the decryption of multiple regulatory networks governing cell growth and death, and despite the possibility of analyzing (epi)genetic deregulation at the genome-wide scale, cancer-targeted therapy is still the exception. In fact, to date there are still far too few examples of therapies leading to cure; treatment-derived toxicity is a major issue, and cancer remains to be one of the largest causes of death worldwide. The purpose of this review is to discuss the state of the art of cancer therapy with respect to the key issue of any treatment, namely its target selectivity. Therefore, we recapitulate and discuss current concepts and therapies targeting tumor-specific features, including oncofusion proteins, aberrant kinase activities and epigenetic tumor makeup. We analyze strategies designed to induce tumor-selective death such as the use of oncolytic virus, tumoricidal proteins (NS1, Eorf4, apoptin, HAMLET (human α-lactalbumin made lethal to tumor cells)) and activation of signaling pathways involved in tumor surveillance. We emphasize the potential of the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) pathway, an essential component of the evolutionary developed defense systems that eradicate malignant cells. Finally, we discuss the necessity of targeting tumor-initiating cells (TICs) to avoid relapse and increase the chances of complete remission, and describe emerging concepts that might provide novel avenues for cancer therapy.

Comparative study of poliovirus excretion after vaccination of infants with two oral polio vaccines prepared on Vero cells or on primary monkey kidney cells.

A comparative study was designed to assess the bioequivalence of 2 oral poliovaccines (OPV) produced on 2 different cell systems: primary monkey kidney (PMK) cells and the Vero cell line. The Vero cell line has been used to overcome the problem of obtaining a regular supply of high quality monkeys that are devoid of latent viruses. For this study, 9 children were vaccinated with PMK-OPV and 12 children with Vero-OPV. The comparison covered poliovirus excretion, reversion of polioviruses in the 5'-noncoding region, and immunogenicity. Major molecular markers in the 5'-noncoding region related to neurovirulence already had been identified at position 480 for type 1, position 481 for type 2, and position 472 for type 3 poliovirus. Two nucleic-acid based methods were designed for studying these positions: a RT-PCR followed by sequencing, which required preliminary culture and cloning; and a type-specific nested PCR followed by sequencing, which enabled direct detection and genotyping of polioviruses. Twenty-eight stool specimens were analyzed by this second method with no PCR inhibition problem. The use of Vero cell line did not modify the global pattern of poliovirus excretion, reversion frequency, or seroconversion. These results provide additional support for the use of the well-characterized Vero cell line in OPV manufacturing.

Protection of non-murine mammals against encephalomyocarditis virus using a genetically engineered Mengo virus.

Genetically engineered Mengo viruses with artificial deletions in the 5' noncoding poly(C) tracts are highly attenuated for pathogenicity when introduced as live vaccines into the natural murine host. Inoculation produces lifelong protective immunity without disease or viral persistence. This report extends the vaccination studies to non-murine hosts, including baboons, macaques and domestic pigs, all of which are susceptible to severe cardiovirus epizootics. All animals of these species that were inoculated with vMC24, an engineered strain of Mengo, seroconverted. When the immunized animals were challenged, they were protected against lethal doses of encephalomyocarditis virus (EMCV) derived from currently circulating epizootic strains. In baboons, the neutralizing antibody titers induced by vMC24 were significantly higher than from an inactivated EMCV vaccine. Moreover, terminal histopathology on baboons (inoculated intramuscularly), macaques (inoculated intracerebrally), and pigs (inoculated intramuscularly) showed few, if any, gross lesions characteristic of EMCV-like disease, in the vMC24 vaccinates. We suggest that genetically engineered, short poly(C) Mengo viruses may be universally potent attenuated vaccines for many types of animals and can possibly provide safe, efficacious protection against all cardioviruses of the EMCV serotype.

Expression and tumorigenicity of the Epstein-Barr virus BARF1 gene in human Louckes B-lymphocyte cell line.

We previously showed that the Epstein-Barr virus, which encodes the BARF1 gene, could transform rodent fibroblasts. In this work, the expression of the BARF1 gene was studied in the human Louckes B-lymphocyte cell line. Introduction of the BARF1 open reading frame under the control of the Mo-MuLV LTR promotor into nontumorigenic Louckes lymphoid cells led to the activation of the c-myc protooncogene and increased expression of the B-cell surface proteins, the transferrin receptor, CD21, and CD23. BARF1-expressing cells induced a diffuse lymphoma-like tumor in newborn rats treated with anti-thymocyte serum that was, however, transient and regressed after 3-4 weeks as the immune system recovered. The tumor induction was similar to that observed with lymphoid cell lines in vitro generated by infection with the B95-8 virus strain, in which lytic antigens are expressed at low levels. After long-term culture, Louckes cell clones lost expression of the BARF1 gene and were unable to induce tumors.

Qualification of working cell banks for the Vero cell line to produce licensed human vaccines.

Since 1980, Merieux Institute has prepared on microcarriers four working cell banks from Vero Cells (137th p.) received from the ATCC in May 1979 (at 124th p.). The lots have been or are used for the production of rabies and inactivated poliomyelitis vaccines. Three lots were controlled according to WHO requirements described in the technical report 673, 1982. For the fourth lot, we have followed the WHO requirements corresponding to the technical report 745, 1987. All the tests required us to demonstrate: i) Safety and purity (tests in animals and eggs, sterility tests, cocultures with human cells and other electron microscopic observations). ii) The absence of tumorigenicity (tests in newborn rats treated with antihymocyte serum at the WBC level and on the cells propagated to at least 10 population doublings beyond the maximum passage level used for production. Assays of cell transformation with DNA from the Vero line in the standard 3T3 assay system). iii) Identity (isoenzyme technique). All were satisfactory.

Intravenous immunoglobulins: can the blood pressure response in laboratory animals be considered a predictive criterion for good clinical tolerance?

Having noted that the only physico-chemical and biological test recommended cannot ensure good tolerance of i.v. IgG in man, we studied a physiological test consisting of evaluating blood pressure during infusion in conscious dogs (10 mg/kg/min) or after i.v. injection in rats (250 mg/kg in 12 sec.). The data were obtained on more than 100 dogs and 500 rats. For preparations known to be well tolerated in man or inducing a few clinical intolerances, the correlation with hypotension in rats and dogs seems good. Therefore, we think this test has its place in the battery of i.v. IgG qualification tests carried out before passage to man. Institut Mérieux has developed a new intact i.v. IgG equilibrated in sub-classes, without PKA, low in IgA and not hypotensive in these two species.