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E Dmitrovsky

Publications and source records attributed to E Dmitrovsky.

At least 55 records · Page 3Linked to original sources
PubMed1996

The epidermal growth factor receptor and its ligands as therapeutic targets in human tumors.

The epidermal growth factor receptor (EGFR) is detected on many non-haematopoietic tissues and is frequently overexpressed in human tumors. With its ligand, TGF-alpha, it forms a well-defined autocrine growth loop. Several clinical approaches, using EGFR as a therapeutic target, are being investigated, particularly monoclonal antibodies combined with chemotherapy, and pharmacological inhibition of downstream components of the EGFR signaling pathway.

Animals↗
PubMed1996

Derivation and characterization of retinoid-resistant human embryonal carcinoma cells.

The retinoids exert potent growth and differentiation effects on normal and neoplastic cells through two families of nuclear receptors. These are the retinoic acid receptors (RAR alpha, RAR beta, RAR gamma) and the retinoid-X receptors (RXR alpha, RXR beta, RXR gamma). All-trans retinoic acid (RA) induces terminal neuronal differentiation and represses tumorigenicity of the multipotent human embryonal carcinoma cell line NTERA-2 clone D1 (NT2/D1). Hexamethylene bisacetamide (HMBA) induces a phenotype distinct from RA-treated NT2/D1 cells. This study reports the derivation and characterization of RA- and HMBA-resistant NT2/D1 clones. Nine RA-resistant (NT2/D1-R1 through NT2/D1-R9) and one HMBA-resistant (NT2/D1-H1) clones were derived after mutagen treatment of NT2/D1 cells and selection in RA or HMBA. NT2/D1-R cells were cross-resistant to 9-cis retinoic acid (9-cis RA), a ligand activating the RAR and RXR pathways, but retained maturation response to HMBA. A representative RA-resistant clone, NT2/D1-R1, overcame the antitumorigenic actions of RA as assessed in athymic mice. NT2/D1-H1 cells were dually resistant to RA and 9-cis RA. All these retinoid resistant cells exhibit deregulated expression of RAR gamma but not RAR alpha or RAR beta. Southern analysis using RAR gamma probes shows no apparent structural differences in genomic DNA between NT2/D1 cells and the RA-resistant subclones. Pulsed-field gel electrophoresis (PFGE) with RAR gamma probes demonstrated an Mlu-I restriction fragment length polymorphism, but no other structural abnormalities in these cells or a panel of germ cell tumor (GCT) cell lines. Full-length RAR gamma 1 coding region cDNAs were cloned from NT2/D1 and NT2/D1-R1 cells and these sequences were identical, suggesting RA resistance in these cells is due to altered regulation of RAR gamma. These differentiation-resistant cells are useful to study RAR gamma target genes or mechanisms engaged by these differentiation inducing agents in human embryonal carcinomas.

Acetamides↗
PubMed1996

Expression of retinoic acid receptor beta in human renal cell carcinomas correlates with sensitivity to the antiproliferative effects of 13-cis-retinoic acid.

The differentiation and growth suppressive effects of retinoic acid are mediated through retinoic acid nuclear receptors (RARs and RXRs), which are ligand-activated transcription factors. Recent data suggest that both altered and regulated expression of RARs are linked to retinoic acid response in a cell context-dependent manner. This study examined the antiproliferative effects of 13-cis-retinoic acid (cRA) on 12 renal cancer cell lines and correlated these findings with the basal and induced expression of RAR-alpha, -beta and -gamma. Eleven of 12 renal cancers that were either resistant to or only minimally inhibited by cRA did not basally express RAR-beta as determined by Northern blot analysis. In these cells, cRA treatment did not induce RAR-beta expression. In contrast, 1 of 12 cell lines (SK-RC-06) was >90% inhibited by cRA and basally expressed RARbeta. Furthermore, RAR-beta mRNA in SK-RC-06 cells was up-regulated by cRA treatment. Amplification of cDNA using PCR and RAR-beta isoform-specific primer pairs revealed that only SK-RC-06 cells expressed the RAR-beta1 isoform. Expression of RAR-alpha transcripts was abundant in all 12 cell lines examined, whereas low levels of RAR-gamma transcripts were detectable in 6 of 10 renal cancers. Expression of RAR-alpha and RAR-gamma was not affected by cRA. These data showing that the majority of renal cancer cell lines are resistant to cRA suggest that: (a) resistance to the antiproliferative action of cRA correlates with repressed RAR-beta mRNA expression; and (b) the antiproliferative effects of cRA in renal cancer cells are mediated through RAR-beta1.

Antineoplastic Agents↗
PubMed1995

Growth suppression of transformed human bronchial epithelial cells by all-trans-retinoic acid occurs through specific retinoid receptors.

The retinoids are reported to chemoprevent second aerodigestive tract cancers in patients with prior lung or head and neck cancers. Since those retinoids already examined in clinical trials do not induce major clinical responses in lung cancers, it is hypothesized that the beneficial chemoprevention activity in lung neoplasias occurs within 'fields' of carcinogen-transformed epithelial cells. To begin to investigate this retinoid action during lung carcinogenesis, the BZR-T33 ras transformed human bronchial epithelial cell line that grows in an anchorage independent manner was examined. This study reports, as compared to controls, that all-trans-retinoic acid (RA)-treatment suppresses BZR-T33 proliferation in monolayer cultures and in anchorage independent and cloning efficiency growth assays. RA induces RAR-gamma 2 > RAR gamma 1 in BZR-T33 cells but expression at the total cellular RNA level of RAR alpha and RXR alpha is not augmented by RA-treatment. RAR beta mRNA expression is repressed before and after RA-treatment and is only detected using a reverse transcription polymerase chain reaction (RT-PCR) assay. To determine directly which of these expressed retinoid receptors signals growth suppression, each receptor was individually transfected into BZR-T33 cells using episomal vector in colony efficiency assays. RAR gamma over-expression in the presence or absence of RA-treatment did not suppress BZR-T33 growth more than controls. In contrast, over-expression of the other examined retinoid receptors inhibited BZR-T33 cellular cloning efficiency prior to RA-treatment and in this decreasing order after RA-treatment: RAR alpha > RAR beta > RXR alpha. The findings reported here reveal that RA suppresses proliferation and cloning efficiency in this transformed human bronchial epithelial cell through specific retinoid receptors. Further work is needed to evaluate the role of RA or its nuclear receptors in inhibiting even earlier steps in lung carcinogenesis.

Base Sequence↗
PubMed1995

Aberrant p53 expression predicts clinical resistance to cisplatin-based chemotherapy in locally advanced non-small cell lung cancer.

The development of cisplatin-based induction chemotherapy followed by surgical resection or radiation has improved the poor prognosis of stage III non-small cell lung cancer (NSCLC). In vitro studies indicate that p53 can modulate cisplatin-induced cytotoxicity, but the molecular genetic features determining response or resistance to cisplatin in vivo must be defined. For this reason, tumor specimens from 52 patients with stage IIIA NSCLC entered in a prospective clinical trial of cisplatin-based induction chemotherapy followed by surgical resection were examined for p53 expression by immunohistochemical staining before and after induction chemotherapy. p53 expression was correlated with clinical and pathological response using Fisher's exact test. No correlation was established between p53 expression and clinical response because 47 of the 52 patients studied had a major response. However, a significant association was observed between aberrant p53 expression and resistance to chemotherapy as assessed by pathological response. Only 3 of the 20 patients whose tumors exhibited a high level (+ + to + + + +) of p53 staining experienced a major (+ + + to + + + +) pathological response to chemotherapy. Only 7 of 52 cases examined before and after chemotherapy treatment exhibited a change in the level of p53 expression after cisplatin-based chemotherapy. These results indicate that cisplatin alters p53 expression infrequently and suggest a direct link between aberrant p53 expression and resistance to cisplatin-based chemotherapy in NSCLC.

Carcinoma, Non-Small-Cell Lung↗
PubMed1995

all-trans retinoic acid for treating germ cell tumors. In vitro activity and results of a phase II trial.

BACKGROUND: Germ cell tumors (GCTs) are characterized by a capacity to differentiate in vivo and in vitro. The authors' previous work highlighted the finding that retinoid-mediated GCT differentiation in vitro correlates with expression of the retinoic acid receptor-gamma. METHODS: Six human GCT cell lines were studied to determine whether their growth and differentiation were linked to the retinoic acid response pathway. The maturation and growth inhibitory responses to all-trans retinoic acid (RA) were studied as well as the expression of retinoic acid receptor-gamma (RAR-gamma). The clinical antitumor activity of RA was studied in a Phase II trial of RA in patients with chemotherapy-refractory GCTs. RESULTS: The RA response pathway was correlated with the control of growth and maturation in three of the six GCT cell lines studied. In one GCT cell line, RA induced RAR-gamma expression and terminal differentiation. In a second, there was high basal expression of RAR-gamma and poor basal proliferative potential. A third cell line showed a more mature basal phenotype than other cell lines studied and marked growth inhibition when treated with RA. Sixteen patients were treated with RA for an average of 7 weeks in the clinical trial. No complete or partial responses were noted. CONCLUSION: A Phase II clinical trial of RA failed to show significant clinical antitumor activity in patients with chemotherapy-refractory GCTs. However in vitro data suggest that the control of growth and maturation in some GCT cell lines involve the RA signaling pathway. Further studies are warranted to define the role that other retinoids with receptor selectivity or more favorable pharmacologic properties may play in the maturation or antitumor responses of GCTs.

Adolescent↗
PubMed1995

Growth suppression of acute promyelocytic leukemia cells having increased expression of the non-rearranged alleles: RAR alpha or PML.

The balanced t(15;17) rearrangement found in acute promyelocytic leukemia (APL) cells fuses PML on chromosome 15 to the retinoic acid receptor alpha (RAR alpha) on chromosome 17. PML/RAR alpha is expressed in APL cells with the non-rearranged alleles, PML and RAR alpha. Clinical remissions induced by all-trans-retinoic acid (RA) treatment of APL patients are linked to expression of PML/RAR apha, a transcription factor with reported dominant negative functions. The roles of PML and RAR alpha in the RA response of APL have not yet been fully explored. This study examines these roles by individually transfecting RAR alpha and PML into NB4 APL cells. NB4 is the sole APL cell line containing the t(15;17). RA treatment represses NB4 cell growth and induces a myeloid phentoype. Full length cDNAs for RAR alpha and PML were individually cloned into a CMV-driven expression vector containing the neomycin resistance gene. Surprisingly, none of the obtained stable transfectants expressed exogenous RAR alpha or PML mRNAs even when reverse transcription polymerase chain reaction (RT-PCR) detection assays were used. All clones expressed the neomycin resistance gene and were similar to parental NB4 cells in their growth and differentiation properties. An explanation explored for this lack of gene expression was that increased levels of RAR alpha or PML might suppress APL cell growth. To examine this possibility, transfection experiments were repeated using an episomal vector-based expression system containing an SV40 driven RAR alpha or PML cDNA and the hygromycin B resistance gene. A new selection strategy augmented expression of the desired cDNAs. A control episomal vector lacked a cDNA insert. Following electroporation and selection, exogenous RAR alpha expression was obtained. Compared to controls, the growth of these transfectants was markedly inhibited before and after RA-treatment and these cells more prominently induced myeloid maturation markers. In contrast, exogenous PML expression was transient since these transfectants did not appear to propagate in culture. These findings indicate: (1) a growth disadvantage for NB4 cells having increased expression of RAR alpha or PML and (2) increased RAR alpha expression augmented RA-mediated maturation of NB4 cells. This implicates a role for RAR alpha or PML in regulating the growth or differentiation of APL cells. It is hypothesized this occurs through antagonism of PML/RAR alpha actions in these leukemic cells.

Alleles↗
PubMed1995

Overexpression of the retinoic acid receptor gamma directly induces terminal differentiation of human embryonal carcinoma cells.

All-trans retinoic acid (RA) exerts profound effects on the growth and differentiation of normal, embryonic, and malignant cells. The effects of RA are mediated through multiple members of the retinoic acid receptor (RAR) and retinoid X receptor (RXR) families of nuclear transcription factors. The RARs and RXRs exhibit specific patterns of expression during development and in adult tissues suggesting tissue or cell-type specific functions. Using NTera2/clone D1 (NT2/D1) human embryonal carcinoma cells as a model, we report that the RA induced terminal differentiation of these cells into a neuronal phenotype is characterized by an increase in expression of RAR alpha, RAR beta, RAR gamma, and a slight induction of RXR alpha. To study the role of these receptors in the differentiation process we individually overexpressed RAR alpha, beta, gamma and RXR alpha in NT2/D1 cells by cDNA transfection. Using induced cDNA expression by episomal vector amplification we show that RAR gamma over-expression causes the terminal mesenchymal differentiation of these cells while over-expression of RAR alpha, beta and RXR alpha has no observed maturation or growth inhibitory effects. Over-expression of these receptors in the derived RA resistant subclone NT2/D1-R1 showed phenotypic changes characteristic of RA response in RAR gamma transfectants. These studies indicate that of the retinoid receptors expressed in RA-treated NT2/D1 cells, it is the upregulation of RAR gamma that specifically induces the terminal differentiation of these cells.

Base Sequence↗
PubMed1995

Aberrant expression of p53 or the epidermal growth factor receptor is frequent in early bronchial neoplasia and coexpression precedes squamous cell carcinoma development.

New strategies are needed for the detection and treatment of lung cancer and must derive from a fuller understanding of lung carcinogenesis. Frequent molecular genetic abnormalities occur in non-small cell lung cancer (NSCLC), but little is known about which of these precede an invasive carcinoma. We examined the expression of p53, epidermal growth factor receptor (EGFR), and transforming growth factor alpha, the most common molecular genetic abnormalities in NSCLC, in preneoplastic bronchial lesions. Primary NSCLC and associated bronchial lesions were identified by retrospective review of resected tumors at this center. Expression in the invasive carcinomas, the associated bronchial lesions, and normal lung were contrasted using immunohistochemistry. Thirty-four NSCLC associated with 62 bronchial lesions were identified. The invasive tumors included 15 squamous cell carcinomas (SCCs) and 19 non-SCCs. Bronchial lesions included areas of squamous metaplasia (n = 14), inflammatory atypia (n = 19), dysplasia (n = 17), and carcinoma in situ (n = 12). Nineteen (56%) NSCLC and 10 (16%) bronchial lesions exhibited aberrant p53 immunostaining, whereas 18 (53%) NSCLC and 30 (48%) bronchial lesions showed abnormal EGFR immunostaining. Positive staining for transforming growth factor alpha was seen in 16 (47%) NSCLC but occurred inconsistently in the bronchial lesions and in normal bronchial epithelium. Only bronchial lesions associated with squamous cell carcinomas exhibited staining for p53. Aberrant EGFR expression was not associated with a specific type of invasive carcinoma or with specific preneoplastic lesions, although there was a trend toward increased expression in dysplasia and carcinoma in situ relative to metaplasia and atypia. All but one of the NSCLC simultaneously showing aberrant p53 and EGFR staining were SCC. We conclude that: (a) transforming growth factor alpha is variably expressed in normal respiratory epithelium as well as reactive and preneoplastic bronchial lesions; (b) p53 expression is seen in preneoplastic bronchial lesions but is not present in reactive or metaplastic epithelium; (c) aberrant EGFR expression occurs in both reactive and preinvasive bronchial lesions and may be an early marker of neoplastic transformation; and (d) the simultaneous aberrant expression of EGFR and p53 occurs predominantly in SCC and their associated bronchial lesions. These findings indicate that aberrant expression of p53 or the EGFR is frequent in bronchial neoplasia, and coexpression may predispose to the development of squamous cell carcinomas of the lung.

Bronchial Neoplasms↗
PubMed1995

Molecular biologic features of non-small cell lung cancer. Clinical implications.

A fuller understanding of the fundamental mechanisms involved in tumor initiation, growth, and metastasis will enable us to develop innovative approaches to detection and treatment that will improve the poor survival of patients with lung cancer. Current information suggests that certain individuals may be predisposed to developing lung cancer and that lung cancers, like other solid tumors, are characterized by the activation of oncogenes, the expression of growth factor loops, and the inactivation of tumor suppressor genes. Within the next decade, it is likely that genetic abnormalities will be used to identify individuals at risk for lung cancer, to select patients for adjuvant therapy, and to develop novel forms of treatment.

Animals↗
PubMed1994

A ribozyme which discriminates in vitro between PML/RAR alpha, the t(15;17)-associated fusion RNA of acute promyelocytic leukemia, and PML and RAR alpha, the transcripts from the nonrearranged alleles.

Acute promyelocytic leukemia (FAB M3) is distinguished by the presence of the t(15;17) and clinical response to all-trans retinoic acid (RA) treatment. Acute promyelocytic leukemia is associated with a chromosomal translocation which results in the fusion of genes encoding a putative transcription factor (PML) and the retinoic acid receptor alpha (RAR alpha). It is suggested that the PML/RAR alpha fusion protein functions as an inhibitor of myeloid differentiation. The potential use of ribozymes as therapeutic agents has been investigated in the present study. Hammerhead ribozymes, which by hybridizing to both PML and RAR alpha sequences discriminate between the fusion transcript and the normal transcripts from the nonrearranged alleles, were designed and synthesized. Two hammerhead cleavage sites were targeted: site 1, an AUU located 4 nucleotides 3' to the fusion junction; and site 2, a UUC located 26 nucleotides 3' to the junction. Both sites are located in the RAR alpha portion of the fusion transcript. Using a full-length PML/RAR alpha RNA or an RNA corresponding to 788 nucleotides of the PML/RAR alpha mRNA and a full-length RAR alpha RNA or an RNA corresponding to 960 nucleotides of the RAR alpha mRNA as model substrates, the catalytic behavior of several ribozymes was studied. A modified hammerhead directed against site 2 displayed the highest degree of selectivity for PML/RAR alpha. It is hypothesized that ribozyme-mediated inactivation of PML/RAR alpha provides a new approach to study the role of PML/RAR alpha in the deregulated growth and RA response of acute promyelocytic leukemia.

Base Sequence↗
PubMed1994

Retinoic acid stimulates protein kinase A-associated G proteins during human teratocarcinoma differentiation.

Retinoic acid (RA) treatment of F9 murine teratocarcinoma (TC) cells reduces the expression of the protein kinase A (PKA)-associated G protein, G alpha i2. The present study reveals interactions between the RA and PKA pathways during differentiation of the multipotent human TC cell line NTERA-2 clone D1 (abbreviated NT2/D1) which differ from prior reports in F9 TC cells. Compared to untreated NT2/D1 cells, differentiated NT2/D1 cells expressed increased levels of G alpha s and G alpha i1,2 proteins as shown by both immunoblot analysis and cholera toxin- and pertussis toxin-induced ADP ribosylation. To further explore cooperation between these pathways during human TC differentiation, we examined the effects of cyclic adenosine monophosphate (cAMP) on RA-responsive genes and of RA treatment on the transcriptional activation of a cAMP response element (CRE). Compared to RA alone, combined treatment with RA and cAMP augmented the expression of the RA nuclear receptor-beta (RAR-beta). Also, transient transfection assays revealed that cAMP and RA cooperated to enhance CRE transcriptional activation. The cAMP-induced enhancement of RA actions in NT2/D1 cells extended to immunophenotypic changes typical of the neuronal differentiation program induced by RA. In contrast to these findings in NT2/D1 cells, prior work in F9 TC cells showed that cAMP inhibits the RA-mediated augmentation of RAR-beta expression and switches the differentiation program from visceral to parietal endoderm. Thus, unlike murine TC cells, in human NT2/D1 cells RA stimulates PKA-associated G proteins and PKA pathway activation enhances RA-mediated TC differentiation.

Cell Differentiation↗
PubMed1994

All-trans retinoic acid for acute promyelocytic leukemia. Results of the New York Study.

OBJECTIVE: To evaluate the safety and efficacy of all-trans retinoic acid to induce complete remission and to examine its effects on duration of remission and survival in patients with acute promyelocytic leukemia. DESIGN: Phase II evaluation and comparison with historical control patients. SETTING: Tertiary care cancer referral center. PATIENTS: Consecutive patients with morphologic diagnoses of acute promyelocytic leukemia were treated during a 2-year period with all-trans retinoic acid (daily oral dose, 45 mg/m2). Newly diagnosed patients discontinued the drug approximately 30 days after they achieved complete remission, at which time they received three courses of combination chemotherapy. Patients treated with previous cytotoxic chemotherapy who then relapsed were continued on all-trans retinoic acid as "maintenance" therapy until they relapsed again. RESULTS: 56 patients entered the study: 34 were newly diagnosed and 22 had relapsed from previous treatment. Fifty-one patients subsequently were found to have the PML/RAR-alpha gene rearrangement indicative of acute promyelocytic leukemia, and 44 of these patients achieved complete remission (86%; 95% Cl, 76% to 96%). A distinctive respiratory distress syndrome developed in 13 patients (23%) during treatment, and 5 patients (9%; Cl, 3% to 20%) died of this complication. The 5 patients who lacked PML/RAR-alpha rearrangements were withdrawn and given chemotherapy. The 13 patients given all-trans retinoic acid alone as maintenance therapy (10 of whom had relapsed from a chemotherapy-induced remission) had a median duration of remission of only 3.5 months (range, 1 to 23 months). Only 3 of 19 patients who relapsed from a remission induced by all-trans retinoic acid could be brought into remission again using this drug. The median survival time of all newly diagnosed patients has not been reached, but it now exceeds 31 months (range, 0.4 to 36+ months). No decrease in the early mortality rate was observed compared with a historical control group composed of 80 consecutive, newly diagnosed patients treated only with chemotherapy at this center; however, overall survival was superior. CONCLUSIONS: All-trans retinoic acid is an effective agent to induce remission in patients with a molecular diagnosis of acute promyelocytic leukemia, but remissions are short and resistance develops rapidly. Although the incidence of early death was not reduced, the use of all-trans retinoic acid to induce remission, followed by cytotoxic chemotherapy for "consolidation," was associated with longer survival times when compared with historical controls treated only with chemotherapy. Additional studies to prevent or mitigate consequences of the "retinoic acid syndrome" and to identify specific patients who might benefit from earlier intervention with chemotherapy are needed to maximize the advantages of this approach.

Adolescent↗
PubMed1994

All-trans-retinoic acid and hexamethylene bisacetamide (HMBA) regulate TGF-alpha and Hst-1/kFGF expression in differentiation sensitive but not in resistant human teratocarcinomas.

The multipotent human teratocarcinoma (TC) cell NTera-2 clone D1 (abbreviated NT2/D1) differentiates into a neuronal lineage after retinoic acid (RA) treatment and a distinct phenotype after hexamethylene bisacetamide (HMBA) treatment. We previously reported that RA treatment of NT2/D1 cells reduces cellular cloning efficiency and nude mouse tumorigenicity. This accompanied a loss of mRNA expression of transforming growth factor-alpha (TGF-alpha) and the fibroblast growth factor kFGF, also known as hst-1 (abbreviated hst-1/kFGF). This study extends prior work by reporting that the distinct phenotype induced by HMBA also decreases cloning efficiency, tumorigenicity, and TGF-alpha and hst-1/kFGF mRNA expression in NT2/D1 cells. These RNA findings were confirmed by measurements of growth factor protein in the conditioned media of inducer-treated and untreated NT2/D1 cells. In two established human TC lines refractory to the actions of RA, N2102ep and Tera-1, RA fails to decrease expression of either growth factor despite induction of its nuclear receptor, RAR-beta. However, HMBA induces morphologic maturation and down-regulation of these growth factors in N2102ep cells. This indicates that the loss of TGF-alpha and hst-1/kFGF expression serves as a new marker of differentiation in human TCs. To explore the effects of these growth factors on growth and differentiation of NT2/D1 cells, TGF-alpha or hst-1/kFGF protein was added following inducer treatment or no treatment. Neither growth factor blocked immunophenotypic differentiation, but both promoted the growth of uninduced NT2/D1 cells in cloning assays.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetamides↗
PubMed1994

Response and resistance to retinoic acid are mediated through the retinoic acid nuclear receptor gamma in human teratocarcinomas.

All-trans-retinoic acid (RA) treatment of the multipotent human teratocarcinoma (TC) cell line NTERA-2 clone D1 (abbreviated NT2/D1) induces a neuronal phenotype. Compared to untreated NT2/D1 cells, RA treated cells have reduced proliferative potential. To identify which retinoic acid receptor (RAR) is directly linked to RA response in these cells, nine RA resistant subclones were derived and characterized. Unlike parental NT2/D1 cells, all these subclones and a de novo RA resistant human TC cell line, N2102Ep clone 2AG (abbreviated N2102ep), exhibited reduced RAR gamma expression. The RAR gamma gene was studied within NT2/D1 cells and a representative RA resistant NT2/D1 subclone called NT2/D1-R1 by Southern analysis and by the transcriptional properties of cloned RAR gamma cDNAs. No structural or functional differences between these RAR gamma species were found suggesting that RA resistance is due to reduced levels of RAR gamma expressed in NT2/D1-R1 cells. To explore this possibility an RAR gamma cDNA was stably transfected into NT2/D1-R1 cells. Expression of this cDNA partially restored the response to RA in NT2/D1-R1 cells. The role of RAR gamma in parental NT2/D1 cells was studied in transient transfection assays using an FGF4 promoter-enhancer reporter construct that is transcriptionally active in undifferentiated but not in differentiated TC cells. The dose dependent co-transfection of an expressed RAR gamma cDNA with this reporter more efficiently repressed its transcriptional activity in the presence of RA than transfection of the reporter without expressed RAR gamma. Co-transfection also decreased reporter activity in the absence of exogenously added ligand. Together, these findings reveal that RAR gamma expression is tightly coupled to RA response and resistance in human TC cells. These data implicate an important role for RAR gamma in the RA-mediated differentiation of these TCs.

Base Sequence↗
PubMed1993

Suramin for germ cell tumors. In vitro growth inhibition and results of a phase II trial.

BACKGROUND: Several studies have indicated a role for specific growth factors in the differentiation or deregulated growth of germ cell cancers. One proposed mechanism of antitumor activity for suramin is the inhibition of these tumor-derived growth factors. METHODS: Based on these data, the effects of suramin were studied on the growth of a monolayer culture of two human teratocarcinoma cell lines, NT2/D1 and N2102ep. These represented one cell line that was sensitive (NT2/D1) and one that was resistant (N2102ep) to the differentiation agent retinoic acid. These studies were followed by a Phase II trial of suramin in patients with germ cell tumors. Patients were treated with a continuous infusion of suramin. Dosing was determined by a nomogram based on weekly serum concentrations. RESULTS: Suramin inhibited the growth of both cell types in a dose-dependent fashion at a concentration attainable in patient plasma. This provided the rationale for a clinical trial of suramin in patients with cisplatin-refractory germ cell tumors. Fourteen patients were treated, and no complete or partial responses were observed. One patient achieved a prolonged decline in serum alpha-fetoprotein level without change in the size of a bidimensionally measurable retroperitoneal mass for more than 11 months. CONCLUSIONS: Despite the results of the in vitro studies, a Phase II trial failed to demonstrate significant antitumor activity for suramin in patients with cisplatin-refractory germ cell tumors. Although the dosing of suramin by the nomogram was not associated with neurotoxicity, the time needed to achieve therapeutic levels was lengthy, and the levels, once achieved, were of short duration. Thus, an alternative means of administration, i.e., by intermittent bolus administration, is being investigated in trials for patients with prostatic carcinoma. The lack of definitive efficacy for suramin in this trial has led the authors to consider other agents in Phase II trials for cisplatin-refractory germ cell tumor.

Adult↗
PubMed1993

Retinoic acid stimulates the protein kinase C pathway before activation of its beta-nuclear receptor during human teratocarcinoma differentiation.

We previously reported that protein kinase C (PKC) stimulation through phorbol ester (TPA) treatment enhances the effects of all-trans retinoic acid (RA) on immunophenotypic differentiation and RA nuclear receptor (RAR) activation in the multipotential human teratocarcinoma (TC) cell line NTera-2/clone D1 (abbreviated NT2/D1). This study extends prior work in NT2/D1 cells by demonstrating that PKC pathway activation is an early effect of RA treatment which regulates RAR transcriptional activity. RA activated the PKC pathway prior to induction of RAR-beta expression at 6 h, which is an established early marker of RAR activation in NT2/D1 cells. RA caused a transient 1.3-fold increase in intracellular diacylglycerol (DG) at 2 min and a translocation of the gamma isozyme of PKC (PKC-gamma) within 5 min. Transient co-transfection studies provided evidence that PKC pathway activation plays a role in the regulation of RAR-beta expression. In these studies a constitutively active PKC-gamma augmented the RA-mediated transactivation of a luciferase reporter containing the native RAR-beta promoter which has a retinoic-acid-response element (RARE). These findings reveal that PKC pathway activation is an early step in RA-mediated human TC differentiation and that PKC-gamma can potentiate the effects of RA on RAR transcriptional activation.

Cell Differentiation↗