Myostatin is a negative regulator of myogenesis, and inactivation of myostatin leads to muscle growth. Here we have used modified RNA oligonucleotides targeting the myostatin mRNA and examined the therapeutic potential in normal and cancer cachexia mouse models. We found that the RNA oligonucleotides could suppress the myostatin expression in vivo, leading to the increase in muscle growth both in normal and cachectic mice. We also established that the effect of myostatin inhibition caused by the RNA oligonucleotides may be through the MyoD pathway, as evidenced by a significant upregulation of MyoD expression. Taken together, these results demonstrate the feasibility using antisense strategy for the treatment of muscle wasting conditions. Gene Therapy (2008) 15, 155-160;

Foxo-1, a member of the Foxo forkhead type transcription factors, is markedly upregulated in skeletal muscle in energy-deprived states such as fasting, cancer and severe diabetes. In this study, we target the Foxo-1 mRNA in a mouse skeletal myoblast cell line C2C12 and in vivo models of normal and cancer cachexia mice by a Foxo-1 specific RNA oligonucleotide. Our results demonstrate that the RNA oligonucleotide can reduce the expression of Foxo-1 in cells and in normal and cachectic mice, leading to an increase in skeletal muscle mass of the mice. In search for the possible downstream target genes of Foxo-1, we show that when Foxo-1 expression is blocked both in cells and in mice, the level of MyoD, a myogenic factor, is increased while a muscle negative regulator GDF-8 or myostatin is suppressed. Taken together, these results show that Foxo-1 pays a critical role in development of muscle atrophy, and suggest that Foxo-1 is a potential molecular target for treatment of muscle wasting conditions.

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Summary The therapeautic efficacy of intact and F(ab')2 fragments of a "'I anti-CEA antibody were compared in an established LS174T colonic xenograft model in nude mice. A single IV dose of either 0.5 mCi (18.5 MBq) intact or 1.0 mCi (37 MBq) F(ab')2 fragments significantly delayed tumour growth, and increased survival time to the same extent. Biodistribution studies showed that the more rapid clearance of the fragments from the circulation improved the tumour:normal tissue ratios found for the intact antibody, but reduced the duration and therefore absolute amount of radioantibody localisation (% injected dose/gram) at the tumour site. The tumours received a similar accumulated beta radiation dose, with 4,065 cGy from 0.5 mCi intact antibody and 4,500 cGy from 1.0 mCi F(ab')2 fragments. The dose rate to the tumour was initially higher for the fragments, but fell off more rapidly as clearance occurred. However, the rapid circulatory clearance resulted in a radiation dose of only 995 cGy to the blood, compared with 2,300 cGy for the intact antibody. This suggests that twice the radiation dose could be delivered to the tumour in the form of fragments for the same blood dose from the intact antibody. Fractionating the 1.0 mCi dose of F(ab')2 into three doses of 0.33 mCi (12.2 MBq), given on days 1, 3 and 5, significantly reduced the therapeutic effect of the treatment. The clinical relevance of these findings is discussed.The successful tumour localisation of radiolabelled antibodies raised against carcinoembryonic antigen (CEA), a tumour associated marker of epithelial carcinomas, has led to the investigation of radioimmunotherapy as a form of cancer treatment in both animal xenograft models (Buchegger et al., 1988;Sharkey et al., 1987;Pedley et al., 1991) and in man (Begent et al., 1989;DeNardo et al., 1988). Antibodies labelled with isotopes emitting medium-to high-energy beta particules such as 13'I and 90Y are promising for solid tumour therapy, because they can deposit their energy over a range of more than 40 cells without requiring either binding to each individual cell or internalisation.However, a major drawback to the use of radioimmunotherapy is the potential damage to normal tissues from the high doses due to circulating radioantibody. The more rapid circulatory clearance and increased tumour penetration normally produced by antibody fragments make them an attractive alternative to intact antibody for tumour localisation and therapy, although they do have the disadvantage of also clearing more rapidly from the tumour itself.We have previously reported on the comparative tumour localisation and clearance patterns of intact IgG and antibody fragments in the nude mouse model (Harwood et al., 1985). The present study compares the therapeutic efficacy of a radiolabelled intact antibody and its F(ab')2 fragments. We have compared the effect of a single dose of "3'I-Fab(ab')2 A5B7, an anti-CEA antibody, with that of the intact antibody on the colonic tumour xenograft LS174T grown in nude (nu/nu) mice, and ha...

Summary Antibody targeted therapy of cancer results in anti-antibody production which prevents repeated treatment. Cyclosporin A (CsA) has been used to suppress this response in patients treated with a radiolabelled antibody to carcinoembryonic antigen (CEA). Patients with CEA producing tumours received a minimum of two courses consisting of an injection of radiolabelled antibody and CsA, 24 mg kg-1 day-1, for 6 days; each course was given at 2 week intervals. Two weeks after the completion of the second course the mean human antimouse antibody (HAMA) levels were 3.5 ug ml -1 (s.d. 2.7) in 3 patients receiving CsA and 1,998 pg ml-1 (s.d. 387) in 3 patients not receiving the drug. Clearance of antitumour antibody was accelerated and tumour localisation absent when HAMA levels exceeded 30 pgml-1. With lower levels of HAMA in the CsA-treated patients, further antitumour antibody accumulated in the tumour after each dose. Further therapy with antitumour antibody and CsA lead to the development of HAMA, but this was less than 25% of the amount in patients not given CsA. In this preliminary study up to 4 times as many doses of antitumour antibody could be usefully given when CsA was used. This increases the potential for effective antibody targeted therapy of cancer.Therapy of cancer with intravenous antitumour mouse monoclonal antibodies alone or conjugated to radionuclides or toxins has produced tumour responses but these are seldom sustained (Meeker et al., 1985, Lenhard et al., 1985, Spitler et al., 1987. Repeated therapy would probably be more effective but is prevented by the formation of human antimouse antibody (HAMA) after one or more injections of antitumour antibody (Meeker et al., 1985, Carrasquillo et al., 1984, Shawler et al., 1985. This causes hypersensitivity reactions and prevents antitumour antibody from localising in the tumour.CsA is a powerful inhibitor of humoral immunity (Borel et al., 1976) and has recently been shown in the accompanying paper to prevent the antibody response to repeated injections of mouse monoclonal antibodies in rabbits (Ledermann et al., 1988 The dose of CsA was adjusted to maintain blood levels between 1,000 and 1,500 ng ml -1 and to prevent a progressive rise in serum creatinine levels. Samples for HAMA were measured before therapy and at intervals after each dose. The distribution of radioactivity in the normal tissues and tumour was determined by planar and single photon emission tomography (SPET) imaging using an IGE Gemini gamma camera.HAMA levels were measured by enzyme immunoassay. Dilutions of serum in PBS containing 0.05% Tween and 0.1% bovine serum albumin were incubated for 2h at room temperature in microtitre wells coated with 10 pgml-l A5B7 in carbonate-bicarbonate buffer, pH 9.6. After washing, wells were incubated with goat anti-human IgG conjugated to alkaline phosphatase (Sigma) followed by the addition of pnitrophenyl phosphate. The absorbance was recorded on a Titertek Multiskan. The assay was standardised using HAMA immunopurified on an A5B7-Sepharose-...