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Biomedical subjects

M E Lippman

Publications and source records attributed to M E Lippman.

At least 217 records · Page 12Linked to original sources
PubMed1985

Estrogen receptor separation employing sucrose deuterium oxide gradients generated during sample centrifugation.

A quick and easy method of sucrose density gradient analysis is described in which prior preparation of gradients for protein separation is not required. We compare the refractive indices as well as the separation pattern of radioactively labeled proteins with either preformed gradients (prepared by layering or by gradient marker) or gradients that establish themselves during centrifugation. Our results show that an identical pattern of protein separation can be achieved by layering the nuclear extracted estrogen receptor either on top of one single sucrose concentration or on a preformed gradient. Centrifugation times are 40 hr in a swinging bucket type rotor (SW60) or 15 hr in a vertical tube rotor (VTi 65). This method avoids the onerous task of gradient preparation, and the separation itself can be achieved in a 15 hr centrifugation by use of a vertical tube rotor.

Breast Neoplasms↗
PubMed1985

Height, weight, and risk of breast cancer relapse.

The records of 231 patients with primary breast cancer and 85 patients with metastatic breast cancer were reviewed to determine whether indices of body weight were associated with prognosis. Results support previous reports that indices of body weight are relatively weak, but real, prognostic factors.

Adult↗
PubMed1985

Increasing the response rate to cytotoxic chemotherapy by endocrine means.

Cloned cell lines of human breast cancer can be growth inhibited by tamoxifen and this inhibition can be reversed by estrogen. We wondered whether tamoxifen inhibition of breast cancer followed by estradiol reversal would increase the efficacy of chemotherapy by increasing the fraction of rapidly cycling cells. We describe a clinical trial in which 110 patients were prospectively randomized to chemotherapy consisting of cytoxan 750 mg/m2 and adriamycin 30 mg/m2 on Day 1 plus 5-FU 500 mg/m2 and methotrexate 40 mg/m2 on Day 8 vs the same chemotherapy plus tamoxifen 20 mg/m2 Days 2-6 and premarin 0.625 mg Q 12-H X 3 on Day 7. Chemotherapy was given in 21-day cycles. 108 patients were evaluable. No difference exist for any important prognostic variables. The first 55 patients were randomized to a regimen in which 5-FU preceded methotrexate by 24 h; thereafter, all patients received methotrexate followed in 1 h by 5-FU. No difference in any response parameter was seen between these two 5-FU methotrexate schedules. No differences in percent of protocol chemotherapy administered or observed toxicity was seen between the 2 regimens. Objective response rate was nearly identical--57% without and 64% with additional hormones. Prior adjuvant chemotherapy with L-PAM had no observable effect on response rate, response duration or survival. In a limited number of patients with inflammatory breast cancer we saw a significantly higher response rate (93 vs 61%; P = 0.03) than in patients with recurrent metastatic disease. Time to progression (13 vs 17 months) and survival (17 vs 23 months) of responders significantly favored the treatment arm including tamoxifen and premarin. Greater benefits of additional tamoxifen and premarin were seen in partial vs complete responders. This may have resulted from lower doses of chemotherapy given to patients achieving a complete remission. An additive effect of hormones plus chemotherapy cannot be entirely excluded as the explanation for the improved results seen with the addition of tamoxifen for 4 days plus 1 day of premarin. We believe that our results suggest that further efforts to increase the efficacy of chemotherapy by perturbing tumor growth rates may be worthwhile.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1985

Onset of androgen action in MCF-7 human breast cancer cells is not accompanied by receptor depletion.

Quantitative and qualitative changes in estrogen receptor follow addition of estradiol to estrogen responsive MCF-7 human breast cancer cells. We asked whether similar changes would accompany treatment of these cells with physiologically relevant concentrations of androgens. Androgen receptor sites were quantified by competitive protein binding assays on whole cells or extracts at various times following hormone addition. Both direct and exchange assays were employed. The androgen receptor in all of these experiments remained in a form which is completely exchangeable and approx 85% salt extractable. Quantity of receptor was unchanged (30,000 sites/cell, Kd 0.1 nM). Responsiveness to hormone treatment was demonstrated by antagonizing the estrogen dependent augmentation of cytoplasmic progesterone receptor in the MCF-7 cells with androgens. Thus, the androgen receptor was shown to be biologically active, but no time dependent quantitative or qualitative changes were observed during the first 6 h following androgen treatment.

Androgens↗
PubMed1985

Biologic implications of steroid hormone receptors in cancers of the colon.

In follow-up investigations on 41 patients with primary cancer of the colon that had been assayed for the presence of steroid-binding activity, six of eight patients whose tumors showed steroid-binding activity for at least one steroid were free of disease one to three years postoperatively. In contrast, only two of 12 patients who had negative binding assay (16%) were free of disease in the same interval. The finding of steroid receptors in human colon cancers does not immediately imply therapeutic advantage. These binding activities may not be receptors, and even if they are, cytoplasmic receptors are a necessary, but not sufficient condition for hormone response. There are, however, sufficient epidemiologic, etiologic, and nutritional observations that seem to link breast and colon cancers to indicate further investigation into the biologic significance of the association of steroid-binding protein in human colon cancer.

Adult↗
PubMed1985

Tamoxifen flare in advanced endometrial carcinoma.

Tamoxifen citrate has recently been shown to be effective palliative therapy in advanced endometrial carcinoma. Flare reactions from tamoxifen are well known in treatment of metastatic breast carcinoma and are not an indication for stopping therapy. We report a patient being treated with tamoxifen for advanced endometrial cancer who developed acute abdominal pain as a manifestation of a flare reaction and the significance of this observation.

Abdomen↗
PubMed1985

Plasma melatonin and the hormone-dependency of human breast cancer.

We studied the 24-hour plasma melatonin profile in three groups of women: normal individuals, women with breast cancer, and women at high risk for breast cancer, to determine the relationship of plasma melatonin to this malignancy. The mean daytime (nadir) and mean nighttime (peak) plasma levels for the normal subjects were 9.1 pg/mL and 70.9 pg/mL, respectively. The mean daytime and nighttime plasma levels, and the range of melatonin day to night differences for women with breast cancer and women at high risk for breast cancer were comparable to each other and to the normal subjects, with no statistically significant differences noted. The patients with breast cancer demonstrated a striking correlation between the melatonin diurnal rhythm and the steroid receptor content of the primary tumor. Women with estrogen (ER) or progesterone (PR) receptor-positive tumors had a significantly lower mean plasma melatonin day to night difference than did patients with ER- or PR-negative tumors. Further, a strong inverse correlation was observed between the plasma melatonin concentration and the quantities of ER and PR in the primary tumor: the lower the plasma melatonin concentration the greater the amount of either receptor in the primary tumor. Plasma melatonin did not correlate with tumor glucocorticoid receptor content or stage of breast cancer among these patients, or with menopausal status, age, parity, or the plasma levels of estrone, estradiol, progesterone, follicle-stimulating hormone (FSH), or luteinizing hormone (LH) among all individuals studied. Plasma melatonin was also independent of the degree of risk for breast cancer among the high-risk patients. These findings suggest an important relationship between the plasma melatonin diurnal rhythm and the hormone dependency of human breast cancer, and may have implications for both the prognosis and treatment of this malignancy.

Adolescent↗
PubMed1985

Selection and characterization of a breast cancer cell line resistant to the antiestrogen LY 117018.

We have selected and cloned a stable variant of the MCF-7 human breast cancer cell line (LY 2) that is resistant to LY 117018 (LY), a potent antiestrogen that inhibits cell growth at concentrations as low as 10(-10) M. The cell line was selected by increasing the concentration of LY in the growth medium in a stepwise manner from 10(-8) to 10(-6) M as the cells become resistant. LY2 has been cloned in soft agar and carried for over 50 passages with no change in resistance. Other antiestrogens, such as tamoxifen and 40-hydroxytamoxifen no longer inhibit cell proliferation of LY 2. The cell line is still responsive to estrogen in a cell proliferation assay, but contains somewhat less estrogen receptors than MCF-7. The cytosolic estrogen receptor sediments to a 4S position on high salt sucrose density gradient centrifugation and is completely shifted to a denser gradient region when the receptor is incubated with a monoclonal antiestrophilin. The nuclear estrogen receptor when covalently labeled with [3H]tamoxifen aziridine has the same mol wt (62,000) in both MCF-7 and LY2 cells, when determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In a competitive binding assay, LY 117018 competes for [3H]estradiol binding to its cytosol receptor with the same Ki in both MCF-7 and LY2 cells. When the induction of estrogen-specific proteins was examined, no detectable progesterone receptor could be detected in either estrogen-induced or control LY2 cells, in contrast to MCF-7 cells. However, both 52,000- and 160,000-dalton proteins were estrogen inducible in the medium of LY2 and MCF-7 cells, as measured by labeling with [35S]methionine. The phenotypic stability of the antiestrogen resistance in LY2 cells coupled with the cross-resistance the antiestrogens of widely different structures make this cell line an ideal model system for the study of hormone resistance in human breast cancer. In addition, while the mechanism of resistance is currently not elucidated, the selective loss of estrogen-inducible functions in this cell line may provide powerful clues for future study.

Binding, Competitive↗
PubMed1985

Response to doxorubicin of cultured normal and cancerous human mammary epithelial cells.

Epithelial cells were isolated and cultured from a number of human mammary specimens of both cancerous and noncancerous origin. Doxorubicin (Dx) sensitivity was measured at second passage with the use of a highly efficient clonogenic assay. For 23 different tumor specimens derived from patients without previous chemotherapy, the drug concentrations required to kill 50% of the cells varied approximately 35-fold. In contrast, for 11 tumor specimens from patients who relapsed after regimens containing Dx, the drug concentration for 50% survival varied only fivefold and the dose-response curves for these specimens clustered at the more resistant end of the spectrum. A wide range of sensitivities was also observed among 13 noncancerous mammary specimens; however, tumor tissue and noncancerous tissue from the same donor were similar. When cultures were subjected to drug incubation periods of 1 and 4 hours, dose-response curves were superimposable when plotted as a function of drug concentration multiplied by time.

Breast↗
PubMed1985

Clonogenic and nonclonogenic in vitro chemosensitivity assays.

We need practical laboratory methods for predicting the chemosensitivity of human neoplasms. Over the past 20 years, numerous investigators have implied or stated with increasing certitude that clonogenic assays are the most valid (or only valid) approach to predictive chemosensitivity testing. We feel that this point of view may have insufficient scientific validity and may be harmful to progress in this area. In addition to well-known technical problems, there are serious theoretical problems with clonogenic assays. These include the disruption of normal cell-to-cell interactions, the possibility that true tumor stem cells may be largely nondividing (G0) cells, while cells forming colonies are exclusively dividing cells, the possibility that clonogenic cells may largely represent cells which are not true stem cells, and the fact that clonogenic assays have the ability to measure cell kill over a narrow log range, while meaningful clinical responses require a multiple-log cell kill. This latter fact mandates the use of unrealistically low drug concentrations to avoid excessive false-positive results. Neither theoretical concepts, direct experimental data, nor clinical correlations support the alleged superiority of clonogenic assays. Clonogenic assays may not be advantageous compared to other more practical methods of estimating the general chemosensitivity of proliferating cells. In contrast, there is a growing body of literature which indicates that early evidence of cell damage in the entire tumor cell population may accurately predict for a multiple-log stem cell kill and meaningful clinical response. Future studies should continue to develop and test assays based on alternative methods for detecting cell kill in the proliferating and total tumor cell populations.

Antineoplastic Agents↗
PubMed1985

Effect of estrogens and antiestrogens on growth-regulatory enzymes in human breast cancer cells in tissue culture.

The effect of estrogens and antiestrogens is examined on three enzymes the activities of which are known to correlate with cell growth. Estrogen treatment increases thymidylate synthetase binding sites up to 4-fold over controls. The extent of induction is dependent on incubation time and the basal rate of cell growth in untreated cells. Amount of active enzyme generally shows a positive correlation with rates of DNA synthesis and cell growth. Thymidine kinase activity and the number of dihydrofolate reductase binding sites are similarly induced by estrogen treatment. Conversely, the effect of antiestrogens on MCF-7 cells is exceedingly complex in that responses in enzyme activities and several generally accepted indices of cell growth (cell number, protein content, rate of DNA synthesis) are dissimilar. Dose response, magnitude of response, and direction of response (increase or decrease) are distinct for each enzyme and for each measure of cell growth with each antiestrogen tested. These results suggest that specific cellular activities are modulated independently by estrogens and antiestrogens and that changes in ligand-receptor complex cannot be the sole explanation for the specificity of estrogen and antiestrogen action. Some degree of specificity and heterogeneity may reside at the level of receptor interaction with the various genes subject to estrogenic modulation.

Breast Neoplasms↗
PubMed1985

Effects of estrogen and tamoxifen on the regulation of dihydrofolate reductase gene expression in a human breast cancer cell line.

We have studied the effects of estrogen and the antiestrogen tamoxifen on the regulation of dihydrofolate reductase (DHFR) gene expression in a methotrexate-resistant (MTXR) human breast cancer cell line MCF-7, which contains a 50-fold increase in the level of DHFR enzyme and amplified DHFR gene sequences. Despite their selection for methotrexate resistance, the MTXR cells have retained many characteristics of the parental MCF-7 cell line. Concentrations of estrogen receptors as well as their binding affinity to estradiol are identical in both cell lines. MTXR MCF-7 cells remain sensitive to estrogen and respond to estradiol with an induction of progesterone receptors, as well as increases in the rate of DNA synthesis and cell growth. Incubation of MTXR MCF-7 cells with estradiol results in an additional 1.5- to 3.0-fold increase in their already elevated level of DHFR. The hormone-induced increases in DNA synthesis and DHFR levels are similar both with respect to the time course of inductions, as well as their dose response to estradiol. However, these two estrogen-induced effects are not coupled, since the induction of DHFR occurs even in the absence of concomitant DNA synthesis. Estradiol has no effect on DHFR enzyme stability; thus, the entire effect of estrogen on DHFR levels results from the increased synthesis of this housekeeping enzyme. In contrast, treatment of MTXR MCF-7 cells with the antiestrogen tamoxifen reduces the rate of DHFR enzyme synthesis, resulting in lower cellular levels of DHFR. These MTXR MCF-7 cells represent a useful model in which to study the mechanisms involved in the modulation of DHFR gene expression by estrogen and tamoxifen. Since the level of DHFR is a critical determinant of methotrexate cytotoxicity understanding, the regulation of DHFR gene expression may have clinical implications for the use of hormonal therapy in combination with chemotherapy for the treatment of breast cancer.

Breast Neoplasms↗