Biomedical subjects
Publications and source records attributed to L Kangas.
The subrenal capsule assay (SRCA) as an in vivo predictive test is reviewed. The main topics include the technique, the evaluability, the reliability (both histological and clinical aspects) and the usefulness of the SRCA in fresh tumor samples.
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The chemotherapeutic potential of 5-hydroxymethyl-2'-deoxyuridine (5HmdUrd) was examined in vitro and in vivo. The compound was toxic in 2-day cultures; 7, 66 and 88% inhibition in the growth of L1210 cells was achieved with 1, 10 and 100 microM 5HmdUrd, respectively. The maximal plasma concentration of 5HmdUrd at 15 min after a single i.p. injection (100 mg/kg) in DBA/2 mice was 193-244 mumol./l and the compound had a logarithmic disappearance curve with a half-life of 20 min. Chemotherapy given as two daily i.p. injections of 5HmdUrd (100 mg/kg) for five successive days resulted in a 239% increase in median lifespan and 2/6 long-term survivals among DBA/2 mice bearing leukemia L1210. This treatment resulted in temporary neutropenia and thrombocytopenia, which were followed by rebound thrombocytosis and neutrophilia of short duration. Our data indicate that 5HmdUrd can successfully be used in experimental cancer chemotherapy in vivo.
After a single intramuscular dose of flunitrazepam 0.015 mg kg-1 (n = 14) in women 37 to 41 weeks pregnant, the concentrations in the umbilical artery and amniotic fluid were significantly lower than in maternal venous plasma. Although the difference between the maternal venous and umbilical venous plasma concentrations was not significant, the mean fetomaternal ratio was 0.7. The plasma protein binding of flunitrazepam was 80 +/- 4% in the mother and 79 +/- 5% in the umbilical circulation. Both mothers and midwives subjectively estimated intramuscular flunitrazepam as a valuable sedative-anxiolytic agent during the first stage of labour.
From 1982 through 1986 the subrenal capsule assay (SRCA) was used to test the sensitivity of 304 gynecological malignant tumors to cytotoxic drug combinations. Most of the tumors were ovarian cancers. Ninety-five percent of the assays were evaluable. Histological and ultrastructural studies were performed on transplants derived from ovarian cancers. Besides cytotoxic drugs, the activity of interferons was also studied by SRCA. Clinical comparisons were available in 43 patients with advanced disease; the predictive accuracy rate of the assay was 77%. The histological, ultrastructural and clinical studies indicated that SRCA is a reliable predictive test, which aids in the selection of proper therapy for individual patients.
The sensitivity to medroxyprogesterone acetate (MPA) and tamoxifen (T) and their combination was assayed in 13 patients with untreated endometrial cancer by an in vitro ATP-bioluminescence method. The method measures the levels of adenosinetriphosphate (ATP), the basic energy source of living cells. A tumor was considered to respond to the drug, if the proportion of living cells after manipulation was 50% or less from unmanipulated control culture. Estrogen (ER) and progesterone (PR) receptors were assayed by the DCC-method and the results calculated by Scatchard-analysis. ER (greater than or equal to 3 fmol/kg cytosol protein) was present in all tumors and PR (greater than or equal to 10 fmol/kg cytosol protein) in 85% of the tumors. The response rate in vitro to MPA was 67% (8 out of 12 tumors), to T 18% (2 out of 11) and to their combination 69% (9 out of 13). The G1 tumors responded statistically significantly better to MPA (p less than 0.01) and MPA + T (p less than 0.02) as compared to T. MPA produced higher cell kill of G1 than G2 tumors (p less than 0.05). The ER content correlated with the effect of MPA in vitro in 67%, with the effect of T in 18% and with that of their combination in 69% of the tumors. The PR content correlated with the effects of MPA in vitro in 83%, with the effect of T in 36% and with that of their combination in 54% of the tumors. It was concluded that the in vitro ATP-bioluminescence method provides valuable information besides steroid receptor determinations for sensitivity testing of endometrial cancer to hormones.
The toxicity and metabolism of a thymidine analogue, 5-hydroxymethyl-2'-deoxyuridine (5HmdUrd) were studied with human leukemia cells (HL-60) and with human platelets. 3 X 10(-5) M 5HmdUrd caused a 50% inhibition in the proliferation of HL-60 cells. The compound was hydrolyzed to 5-hydroxymethyluracil (5HmUra) by the enzyme thymidine phosphorylase (EC 2.4.2.4) present in leukemia cells; this catabolic product was non-toxic. The catabolism of 5HmdUrd by human platelet thymidine phosphorylase could be inhibited by 6-aminothymine. The toxicity of 5HmdUrd was effectively reversed by deoxycytidine and 5HmdUrd increased the incorporation of deoxycytidine into dCTP and DNA several fold. The two latter phenomena are explicable in terms of a feedback action to ribonucleotide reductase, resulting in deoxycytidylate starvation, which is a known effect of excess thymidine. We report here also our preliminary observations that 5HmdUrd is active against mouse leukemia in vivo.
The basic pharmacological and biochemical properties of a new antiestrogen, Fc-1157a, are described. Fc-1157a is bound specifically and with high affinity to estrogen receptors. The binding is competitive with estradiol. Fc-1157a treatment induces translocation of estrogen receptors from cytoplasm to nucleus. The turnover rate of nuclear estrogen receptors is markedly lower than with estradiol, but is more rapid than after tamoxifen. Fc-1157a is an almost pure antiestrogen in rat uterus, but has intrinsic estrogenic activity in mouse uterus. In animal experiments Fc-1157a has shown antitumor properties, which are described in the companion paper.
The antitumor effects of a new antiestrogen, Fc-1157a have been studied in vitro and in vivo. In vitro the effect of Fc-1157a was comparable to that of tamoxifen. The effect was dose-dependent, and at concentrations higher than 10(-6) mol/1 Fc-1157a induced real cell death of the MCF-7 cells. In DMBA-induced mammary cancer in rats Fc-1157a decreased the number of new tumors and inhibited the growth of existing tumors, these effects being statistically highly significant. The ratio of growing tumors to stable and regressing tumors was significantly decreased. Although these effects were slightly stronger with Fc-1157a than with tamoxifen, the difference between these two compounds was not statistically significant. Murine uterine sarcoma, an estrogen receptor-negative tumor, was resistant to tamoxifen, but was statistically significantly inhibited by high doses (100 and 200 mg/kg-1 day-1 for 5 days) of Fc-1157a. The antitumor effects of Fc-1157a are due mainly to the antiestrogenic activity. At high concentrations in vitro and at high doses in vivo Fc-1157a exerts antitumor effects some of which are different from those of tamoxifen and are directed even against estrogen receptor-negative tumors. The exact mechanism of the observed cytolytic effect at high doses is unknown.
The effects of pH on the antitumour activity of six cytostatics--cisplatin, doxorubicin, ethoglucid, mitomycin C, thiotepa and VM-26--used in intravesical instillations were tested in vitro. The activity of cytostatics was determined on the Walker 256 carcinosarcoma cell line by using ATP-bioluminescence method. Cytostatics were incubated at different pH's (5.0-7.4) for 2 and 4 h. The activity of most of the cytostatics was not affected by pH, but mitomycin C had somewhat lower activity at pH 5.0 at 4 h and cisplatin at pH 6-7 at 2 and 4 h. Because the pH of normal urine is within the range used in this test, the use of a buffer solution during the intravesical instillation therapy is not necessary. However, according to our clinical experience, buffering is recommended to avoid chemical cystitis.
Rous sarcoma cells were implanted into the kidney of rats. After 5 days of growth the renal tumor was used for comparing histology with glucose 6-phosphatase (G6Pase) enzyme histochemistry (EHC) and 18F-fluoro-2-deoxyglucose (FDG) auroradiography (ARG). It was found that the regions of the kidney tumor that had retained normal kidney structures were devoid of FDG, whereas there was histochemical staining of normal cortical areas. Regions of tumor growth, on the other hand, retained FDG and lacked G6Pase. Necrotic areas did not accumulate FDG. There was a dramatic decrease in the areas of G6Pase activity as a result of tumor infiltration in the kidney. The results show that FDG, currently being evaluated as a tumor detecting radiopharmaceutical indeed accumulates into areas of vital malignant growth, and they indicate that FDG positron emission tomographic (PET) images reveal the true anatomic location of malignant tissue.
The subrenal capsule assay (SRCA) was used to study the sensitivity of breast cancer to cytostatic drug combinations. The results were compared with steroid receptor status. Forty-five of 46 SRCAs (98%) were macroscopically evaluable. However, a histological study implied that the transplants should also be evaluated histologically, because in only 14/21 (67%) of the control SRCAs examined were histologically viable tumor cells seen. An inflammatory cell reaction was noticed in half of the cases. In the groups treated with cytostatics only 3/21 (14%) had vital tumor cells, whereas inflammation was evident in 4/21 (19%) of the cases. The rate of resistance to A + CTX was 30%. By testing several drug combinations against each tumor the rate of chemoresistance was reduced to 10%. The differences between A + CTX and the best combination were statistically significant (P less than 0.05). Of the tumors 79% were ER-positive and 67% PR-positive. Receptor-negative tumors tended to be more sensitive to cytostatics than receptor-positive tumors (100 vs 85%). The difference was not, however, statistically significant. It can be concluded that the SRCA under standardized conditions is a good method for studying the response of individual breast cancers to chemotherapy.
The 6-day subrenal capsule assay (SRCA) in normal immunocompetent mice was used to assess response of vulvar and cervical cancers to chemotherapy. Twenty-five out of 31 assays (81%) were evaluable. The previously treated tumors tended to be less sensitive than the untreated tumors (20 vs 33%). Three to five cytotoxic drugs or drug combinations were tested against each individual tumor. In the whole material the combination of cisplatin and etoposide (VP-16) was significantly more effective than bleomycin (P less than 0.01, chi 2 test). The combinations adriamycin + cyclophosphamide + cisplatin and cyclophosphamide + methotrexate were also rather effective. Preliminary clinical correlations were positive. The reliability of the SRCA was discussed, and it was concluded that the assay is a promising predictive method for individualizing chemotherapy.
Specimens from 16 freshly biopsied human tumors, two mammary adenocarcinomas, ten ovarian adenocarcinomas, two squamous cell carcinomas, one malignant histiocytoma and one chondrosarcoma of the bone, two human ovarian adenocarcinomas established by transplantation into nude mice and two adenocarcinomas induced in rat mammary gland were transplanted under the renal capsule of 510 normal immunocompetent mice and 180 rats and the effects of chemotherapy were evaluated. The results showed successful transplantation of all types of tumors in both animal species. Morphological analysis revealed preserved glandular structures with surface microvilli, mucin and CEA production and partially preserved basement membranes. Treatment with cyclophosphamide, vinblastine, adriamycin and cisplatin caused cell shrinkage, degradation and partial or total disappearance of the tumor cells. Vascularization was distinct in all specimens. A cellular infiltrate was found frequently but not consistently. A common end stage was a fibrotic scar with no cellular activity, occasionally giving a misleading impression of a growing tumor on gross observation. The results were obtained rapidly and suggest that the subrenal capsule assay would be useful for evaluating the sensitivity of human tumors to therapeutic manipulation, but needs supplementary histological examination.
Neither doxorubicin (ADM) nor mitomycin C (MMC) were well absorbed through the intact or damaged rabbit bladder wall into the blood stream. Preliminary clinical investigations indicating negligible systemic side effects of ADM and MMC agree with the experimental results.
Combinations of interferons (IFNs) and cytostatic drugs were studied in subrenal capsule assay (SRCA) using fresh human ovarian cancer samples. Slight tumour growth inhibition was observed with IFN-alpha and IFN-gamma. The effect was more pronounced if combination IFN-alpha + IFN-gamma was used. Cytostatic drug combination cyclophosphamide + cisplatin + doxorubicin (combination I) was more effective than interferons, and combination methotrexate + tegafur (combination II) was about equal to IFN-alpha. When IFN-alpha was given together with cytostatic drugs, the antitumour effect was diverse: both synergistic and antagonistic effects were found, as well as no evidence of interaction. IFNs did not increase the toxicity of cytostatic drugs. If there is a good correlation between SRCA and clinical results, the combination of IFNs and cytotoxic drugs should be used clinically in connection with a predictive test.
The effect of human interferons alpha and gamma alone and in combination with a novel antiestrogen toremifene were studied in vitro using MCF-7 cell line, an estrogen receptor positive and antiestrogen sensitive cell line. The effects were evaluated by a simple bioluminescence method with which the number of living cells was obtained as cellular adenosine triphosphate (ATP) content. The growth of MCF-7 cells was inhibited both by interferon alpha and interferon gamma. At least additive effect was evident when the cells were exposed to combination of interferons and toremifene: the combination was additive with interferon gamma + toremifene and synergistic with interferon alpha + toremifene. The combination of toremifene and interferons may have clinical importance.