A comparison of efficacy and toxicity between electroporation and adenoviral gene transfer

BACKGROUND: Electroporation of skeletal muscle after injection of naked DNA was shown by others to increase transgene expression. Information regarding tissue damage caused by electroporation is conflicting. It is also not well known how plasmid electroporation compares with transfection by adenoviral vectors. To investigate these questions the most used protocol for muscle electroporation was used, i.e. 8 pulses of 200 V/cm and 20 ms at a frequency of 1 Hz. RESULTS: Intra-muscular DNA transfer of pLuciferase was increased by 2 logs after electroporation, confirming data described by others. However, the blood levels of the encoded protein were still lower than those obtained after injection of first generation adenoviral vectors. Also, the electroporation procedure, on its own, caused severe muscle damage consisting of rhabdomyolysis and infiltration, whereas the adenoviral vectors caused only a slight infiltration. As damage of targeted tissue may be an advantage in the case of tumour transfection, we also compared the two transfection methods in tumour tissue. In case of poorly permissive tumours, adenoviral vectors cannot transfect more than 2% of the tumour tissue without inducing significant liver damage. In contrast, the electroporation seems to offer a wider therapeutic window since it does not cause any systemic toxicity and still induce's significant transfection. CONCLUSIONS: Plasmid electroporation of the muscle induce severe local damage and is of no advantage over adenoviral vectors for obtaining high blood levels of a vector encoded protein. In contrast, electroporation of tumours might be safer than adenoviral gene transfer

Pharmacogenetic heterogeneity of transgene expression in muscle and tumours

BACKGROUND: Recombinant adenoviruses are employed to deliver a therapeutic transgene in the liver, muscle or tumour tissue. However, to rationalise this delivery approach, the factors of variation between individuals need to be identified. It is assumed that differences between inbred strains of laboratory animals are considered to reflect differences between patients. Previously we showed that transgene expression in the liver of different rat strains was dependent on the transcription efficiency of the transgene. In the present paper we investigated if transfection of muscle and tumour tissue were also subject to such variations. METHODS: Variation, in transgene expression, after intramuscular gene delivery was determined in different rodent strains and gene expression in tumours was investigated in different human and rodent cell lines as well as in subcutaneously implanted rodent tumours. The molecular mechanisms involved in transgene expression were dissected using an adenovirus encoding luciferase. The luciferase activity, the viral DNA copies and the luciferase transcripts were assessed in cultured cells as well as in the tissues. RESULTS: Large differences of luciferase activity, up to 2 logs, were observed between different rodent strains after intramuscular injection of Ad Luciferase. This inter-strain variation of transgene expression was due to a difference in transcription efficiency. The transgene expression level in tumour cell lines of different tissue origin could be explained largely by the difference of infectibility to the adenovirus. In contrast, the main step responsible for luciferase activity variation, between six human breast cancer cell lines with similar phenotype, was at the transcriptional level. CONCLUSION: Difference in transcriptional efficiency in muscles as observed between different inbred strains and between human breast cancer cell lines may be expected to occur between individual patients. This might have important consequences for clinical gene therapy. The variation between tumour types and tissues within a species are mainly at the levels of infectivity

Adenoviral gene transfer of angiostatic ATF-BPTI inhibits tumour growth

BACKGROUND: The outgrowth of new vessels – angiogenesis – in the tumour mass is considered to be a limiting factor of tumour growth. To inhibit the matrix lysis that is part of the tumour angiogenesis, we employed the chimeric protein mhATF-BPTI, composed of the receptor binding part of the urokinase (ATF) linked to an inhibitor of plasmin (BPTI). METHODS: For delivery, recombinant adenovirus encoding the transgene of interest was injected intravenously or locally into the tumour. The anti tumour effect of this compound was compared to that of human endostatin and of mhATF alone in two different rat bronchial carcinomas growing either as subcutaneous implants or as metastases. RESULTS: Significant inhibition of the tumour growth and decrease of the number of lung metastasis was achieved when the concentration of mhATF-BPTI at the tumour site was above 400 of ng / g tissue. This concentration could be achieved via production by the liver, only if permissive to the recombinant adenovirus. When the tumour cells could be transduced, local delivery of the vector was enough to obtain a response. In the case of metastasis, the capacity of the lung tissue to concentrate the encoded protein was essential to reach the required therapeutic levels. Further, endostatin or mhATF could not reproduce the effects of mhATF-BPTI, at similar concentrations (mhATF) and even at 10-fold higher concentration (endostatin). CONCLUSION: The ATF-BPTI was shown to inhibit tumour growth of different rat lung tumours when critical concentration was reached. In these tumour models, endostatin or ATF induce almost no tumour response

Peripheral precocious puberty in Li–Fraumeni syndrome: a case report and literature review of pure androgen-secreting adrenocortical tumors

Abstract Introduction Pure androgen-secreting adrenocortical tumors are a rare but important cause of peripheral precocious puberty. Case presentation Here, we report a pure androgen-secreting adrenocortical tumor in a 2.5-year-old boy presenting with penile enlargement, pubic hair, frequent erections, and rapid linear growth. We confirmed the diagnosis through laboratory tests, medical imaging, and histology. Furthermore, genetic testing detected a pathogenic germline variant in the TP53 gene, molecularly confirming underlying Li–Fraumeni syndrome. Discussion Only 15 well-documented cases of pure androgen-secreting adrenocortical tumors have been reported so far. No clinical or imaging signs were identified to differentiate adenomas from carcinomas, and no other cases of Li–Fraumeni syndrome were diagnosed in the four patients that underwent genetic testing. However, diagnosing Li–Fraumeni syndrome is important as it implies a need for intensive tumor surveillance and avoidance of ionizing radiation. Conclusion In this article, we emphasize the need to screen for TP53 gene variants in children with androgen-producing adrenal adenomas and report an association with arterial hypertension