|Budget Amount *help
¥1,800,000 (Direct Cost: ¥1,800,000)
Fiscal Year 1988: ¥200,000 (Direct Cost: ¥200,000)
Fiscal Year 1987: ¥1,600,000 (Direct Cost: ¥1,600,000)
The effect partially purified rat tumor necrosis factor (TNF) was tested against 9L rat brain tumor both in vivo and in vitro. The TNF-containing serum (TNS) was produced by intravenous injection of OK432 and lipopolysaccharide (LPS). Injection of TNS significantly (p<0.05) prolonged the survival time of brain tumor-bearing rats (29.9 12.6 days after tumor cell inoculation, as compared to 20.8 4.4 days in the untreated group).
In the in vitro assay, medium containing 50% TNS significantly decreased the viability of 9L brain tumor cells, by 57.6%, 50.0%, and 57.0% at 3, 5, and 7 days after the beginning of culture, respectively. TNS also displayed significant inhibition of cell growth, indicating a cytostatic effect. To verify TNS activity, TNS was partially purified by means of the DEAE-Sephadex a 50 batch ion exchange method and Sephadex G 200 column chromatography. Four fractions were tested in TNF-sensitive L(S) cells, TNF-resistant L(R) cells, and 9L brain tumor cells. Fraction 4 of TNS demonstrated 37.5%, 88.1%, and 43.2% cell viability against L(S), L(R), and 9L cells, respectively. On the other hand, fraction 4 of normal rat serum showed 87.5%, 87.8%, and 82.2% cell viability, respectively. These results strongly suggest the presence of TNF in the TNS produced by OK432 and LPS.
The effect of recombinant human TNF( -hTNF, kindly supplyed by Dainippon Pharmaceutical Co, LTD.) was also tseted against 9l brain tumor in vivo. When the 9L tumor reached to the 10mm in diameter, 5000unit of -hTNF with or without lymphokine activated killer cell was injected intraven ously or intratumorally. Fine injections were given to each mouse one week interval. The result was that the -htnf with LAK cells siginificantly prolonged their survival time (P<0.005) compared to other groups.