Extreme lymphoplasmacytosis and hepatic failure associated with sulfasalazine hypersensitivity reaction and a concurrent EBV infection--case report and review of the literature.

We present an unusual case of a patient with extreme lymphoplasmacytosis and hepatic failure in association with a reaction to sulfasalazine and a concurrent Epstein-Barr virus (EBV) infection. Sulfa drugs can cause a wide range of allergic and hypersensitivity reactions and occasionally can lead to a fulminant illness. In the case under discussion the patient had hepatotoxicity, skin rash, fever, and peripheral blood atypical lymphocytosis. Initial impressions suggested the possibility of a malignant lymphoproliferative disorder. Flow cytometry of peripheral blood and a bone marrow biopsy provided clear evidence for a reactive, polyclonal process as opposed to a malignant disorder. Cessation of the offending drug and administration of steroids led to dramatic improvement. This case illustrates that drug hypersensitivity reactions can be manifested by an extreme lymphocytoid leukemoid reaction.

Attitudes and practices among pediatric oncologists regarding end-of-life care: results of the 1998 American Society of Clinical Oncology survey.

PURPOSE: In 1998, the American Society of Clinical Oncology (ASCO) surveyed its membership to assess the attitudes, practices, and challenges associated with end-of-life care of patients with cancer. In this report, we summarize the responses of pediatric oncologists and the implications for care of children dying from cancer. METHODS: The survey consisted of 118 questions, covering eight categories. All ASCO members in the United States, Canada, and the United Kingdom were mailed a survey, which was completed by 228 pediatric oncologists. Predictors of particular attitudes and practices were identified using stepwise logistic regression analysis. Potential predictors were age, sex, religious affiliation, importance of religious beliefs, recent death of a relative, specialty, type of practice (rural or urban, academic or nonacademic), amount of time spent in patient care, number of new patients in the past 6 months, and number of patients who died in the past year. RESULTS: Pediatric oncologists reported a lack of formal courses in pediatric palliative care, a strikingly high reliance on trial and error in learning to care for dying children, and a need for strong role models in this area. The lack of an accessible palliative care team or pain service was often identified as a barrier to good care. Communication difficulties exist between parents and oncologists, especially regarding the shift to end-of-life care and adequate pain control. CONCLUSION: Pediatric oncologists are working to integrate symptom control, psychosocial support, and palliative care into the routine care of the seriously ill child, although barriers exist that make such comprehensive care a challenge.

Attitudes and practices of U.S. oncologists regarding euthanasia and physician-assisted suicide.

BACKGROUND: The practices of euthanasia and physician-assisted suicide remain controversial. OBJECTIVE: To achieve better understanding of attitudes and practices regarding euthanasia and physician-assisted suicide in the context of end-of-life care. DESIGN: Cohort study. SETTING: United States. PARTICIPANTS: 3299 oncologists who are members of the American Society of Clinical Oncology. MEASUREMENTS: Responses to survey questions on attitudes toward euthanasia and physician-assisted suicide for a terminally ill patient with prostate cancer who has unremitting pain, requests for and performance of euthanasia and physician-assisted suicide, and sociodemographic characteristics. RESULTS: Of U.S. oncologists surveyed, 22.5% supported the use of physician-assisted suicide for a terminally ill patient with unremitting pain and 6.5% supported euthanasia. Oncologists who were reluctant to increase the dose of intravenous morphine for terminally ill patients in excruciating pain (odds ratio [OR], 0.61 [95% CI, 0.48 to 0.77]) and had sufficient time to talk to dying patients about end-of-life care issues (OR, 0.79 [CI, 0.71 to 0.87]) were less likely to support euthanasia or physician-assisted suicide. During their career, 3.7% of surveyed oncologists had performed euthanasia and 10.8% had performed physician-assisted suicide. Oncologists who were reluctant to increase the morphine dose for patients in excruciating pain (OR, 0.58 [CI, 0.43 to 0.79]) and those who believed that they had received adequate training in end-of-life care (OR, 0.86 [CI, 0.79 to 0.95]) were less likely to have performed euthanasia or physician-assisted suicide. Oncologists who reported not being able to obtain all the care that a dying patient needed were more likely to have performed euthanasia (P = 0.001). CONCLUSIONS: Requests for euthanasia and physician-assisted suicide are likely to decrease as training in end-of-life care improves and the ability of physicians to provide this care to their patients is enhanced.

SMARCAD1, a novel human helicase family-defining member associated with genetic instability: cloning, expression, and mapping to 4q22-q23, a band rich in breakpoints and deletion mutants involved in several human diseases.

Members of the DEAD/H box-containing helicase superfamily include proteins essential to genome replication, repair, and expression. We report here the cloning and initial characterization of a novel human member of this protein family, designated hHel1 (human helicase 1), now designated SMARCAD1 by HUGO. This DEAD/H box-containing molecule has seven highly conserved sequence regions that allow us to place it in the SNF2 family of the helicase superfamily. Uniquely, though, hHel1 contains two DEAD/H box motifs, a property not reported to be shared by any other SNF2 family members. This defines a new subfamily consisting of hHel1 and its homologues. In addition to these DEAD/H box/ATP-binding motifs, hHel1 has a putative nuclear localization signal and several regions that may mediate protein-protein interactions. Expression analysis indicates that hHel1 transcripts are ubiquitous, with particularly high levels in endocrine tissue. We have mapped the gene for hHel1 to human chromosome 4q22-q23; this region is rich in breakpoints and deletion mutants of genes involved in several human diseases, notably soft tissue leiomyosarcoma, hepatocellular carcinoma, and hematologic malignancies. Our observation that human Hel1 gene overexpression is present in an E1A-expressing cell line with increased capacity for gene reactivation events by genomic rearrangement suggests that human Hel1 may play a role in genetic instability development.

Dose-intensive therapy for limited-stage small-cell lung cancer: long-term outcome.

PURPOSE: To determine progression-free survival (PFS) and overall long-term survival for limited-stage small-cell lung cancer (SCLC) patients aged 60 years or younger who respond to first-line chemotherapy followed by high-dose combination alkylating agents (cyclophosphamide 5,625 mg/m(2), cisplatin 165 mg/m(2), and carmustine 480 mg/m(2)) with hematologic stem-cell support and chest and prophylactic cranial radiotherapy. PATIENTS AND METHODS: Patients were selected on the basis of their continued response to first-line therapy, their relative lack of significant comorbidity, and their ability to obtain financial clearance. RESULTS: Of 36 patients with stage III SCLC, nine patients (25%) had achieved a complete response (CR), 20 had achieved a near-CR, and seven had achieved a partial response before undergoing high-dose therapy. Toxicity included three deaths (8%). For all patients, the median PFS was 21 months. The 2- and 5-year survival rates after dose intensification were 53% (95% confidence interval [CI], 39% to 72%), and 41% (95% CI, 28% to 61%). Of the 29 patients who were in or near CR before undergoing high-dose therapy, 14 remain continuously progression-free a median of 61 months (range, 40 to 139 months) after high-dose therapy. Actuarial 2- and 5-year PFS rates were 57% (95% CI, 41% to 79%) and 53% (95% CI, 38% to 76%). By multivariate analysis, short intensive induction chemotherapy was associated with favorable outcome (P <.05). CONCLUSION: Use of high-dose systemic therapy with intensive local-regional radiotherapy was associated with manageable treatment-related morbidity and mortality. Patients who were in or near CR before intensification are enjoying an unmaintained 5-year PFS rate of 53%. Late complications were infrequent, and most patients returned to full-time work and activity. A randomized comparison of this approach and conventional-dose therapy should define the use of dose intensification with hematopoietic support in patients with responding limited-stage SCLC.

Genetic analysis of adenovirus E1A: induction of genetic instability and altered cell morphologic and growth characteristics are segregatable functions.

Single multifunctional oncoproteins contribute to genomic instability development, but relationships between one or more oncoprotein-associated activities and genetic changes accompanying tumor cell progression are uncertain. Using NIH 3T3 derivative EN/NIH 2-20 containing transcriptionally silent neomycin phosphotransferase gene (neo) integrants with undetectable spontaneous reactivations, we studied wild-type (WT) and mutant adenovirus E1A-induced neo reactivation by neo-allelic rearrangement. WT E1A expression, yielding differential splice transcripts 12S and 13S and resulting in altered cell morphologic and growth characteristics, produced neo reactivations in 9 of 21 subclones (median rate per cell, 35 x 10(-6); range, 0.33 x 10(-6) to 936 x 10(-6)). Only 3 of 17 cell lines expressing CTdl976, a '12S' functional equivalent inducing altered cell morphologic and growth characteristics while lacking the 13S trans activation domain, yielded neo reactivations (range, 0.33 x 10(-6) to 0.67 x 10(-6)). One of 21 subclones expressing NTdl646, an E1A mutant retaining the trans domain but lacking p300 binding activity and the ability to alter cell morphologic and growth characteristics, produced neo reactivations (8.7 x 10(-6)). Other E1A mutants, all lacking the ability to alter cell morphologic and growth characteristics while binding pRb but variously lacking the trans domain and binding for p107 and/or p300, displayed undetectable neo-reactivations. 98 EN/NIH 2-20 derivatives coexpressing complementary mutant E1As exhibited altered morphologic and growth features, but only 10 of these produced neo reactivations, and maximum rates (14 x 10(-6)) were substantially lower than those in comparably derived, morphologically altered E1AWT-expressing counterparts (497 x 10(-6)). These findings suggest that maximum rates of gene reactivations by genomic rearrangement require the collective activities of functional domains assembled in single multifunctional proteins (or complexes) while altered cell morphologic and growth characteristics may arise through comparable sets of functional domains distributed across more than one protein (or complex).

Neutropenic enterocolitis as a complication of high dose chemotherapy with stem cell rescue in patients with solid tumors: a case series with a review of the literature.

BACKGROUND: This case series with an accompanying review of the literature describes neutropenic enterocolitis as a complication of high dose chemotherapy with stem cell rescue in patients with solid tumors. METHODS: Neutropenic enterocolitis is documented in two patients undergoing autologous stem cell transplantation. A review of MEDLINE from 1970 to the present was performed to delineate the prior related disease settings in which this condition has been described. The nature of the clinical syndrome and controversies regarding management are discussed. RESULTS: The authors report two patients who each received autologous stem cell transplantation for the treatment of a solid tumor. Both patients presented with progressive abdominal pain, signs of peritoneal irritation on examination, and pneumatosis intestinalis. In the first case, the syndrome developed rapidly over several hours, requiring emergent surgical intervention during the period of postchemotherapy nadir. In the second case, the patient developed a slowly progressive clinical picture that was managed successfully with medical therapy alone. A review of the literature suggests that neutropenic enterocolitis arises due to drug-induced bowel wall mucosal injury followed by superinfection with colonic and opportunistic organisms. Surgical intervention potentially is beneficial in patients who develop an acute syndrome that is unresponsive to antibiotic and supportive therapy. Those patients who can be stabilized with medical management often will recover once reengraftment occurs. CONCLUSIONS: Neutropenic enterocolitis is described in two patients undergoing autologous stem cell transplantation for solid tumors. The management of this life-threatening complication is controversial and governed by the nature of the clinical presentation.

Dose escalation of the hypoxic cell sensitizer etanidazole combined with ifosfamide, carboplatin, etoposide, and autologous hematopoietic stem cell support.

Multiple mechanisms of drug resistance contribute to treatment failure. Although high-dose therapy attempts to overwhelm these defenses pharmacologically, this approach is only successful in a fraction of treated patients. Many drug resistance mechanisms are shared between malignant and normal cells, but the expression of various drug resistance mechanisms associated with hypoxia is largely confined to tumor tissue. Thus, reversal of this mechanism is likely to provide a therapeutic advantage to the host. This study was designed to define the dose-limiting toxicities and maximum tolerated dose of etanidazole when it is given concurrently with high-dose ifosfamide, carboplatin, and etoposide (ICE), with hematopoietic stem cell support. The maximum tolerated doses of high-dose ICE were administered concurrently with dose escalations of etanidazole, a hypoxic cell sensitizer. All agents were given by 96-h continuous i.v. infusion beginning on day -7. Mesna uroprotection was provided. Autologous marrow and cytokine mobilized peripheral blood progenitor cells were reinfused on day 0. Granulocyte colony-stimulating factor was administered following reinfusion until the granulocytes recovered to > 1000/microliter. Fifty-five adults with advanced malignancies were enrolled in cohorts of five to nine patients. Four dose levels of etanidazole between 3 and 5.5 g/m2/day (12, 16, 20, and 22 g/m2 total doses) and two doses of carboplatin (1600 and 1800 mg/m2 total doses) were evaluated. Seven patients died of organ toxicity (13%); two each from veno-occlusive disease of liver and sepsis; and one each from sudden death, renal failure, and refractory thrombocytopenic hemorrhage. Five deaths occurred at the top dose level. One additional patient suffered a witnessed cardiorespiratory arrest from ventricular fibrillation and was resuscitated. Dose-dependent and largely reversible peripheral neuropathy was observed consisting of two syndromes: severe cramping myalgic/neuralgic pain, predominantly in stocking glove distribution, occurring between day -3 and day 0, and a sensory peripheral neuropathy with similar distribution peaking around day +60. The maximal achievable dose of etanidazole (16 g/m2 dose level) resulted in a mean serum level of 38 micrograms/ml (25-55 micrograms/ml). Etanidazole significantly enhanced host toxicity of high-dose ICE. Effective modulatory doses of etanidazole could not be given with acceptable toxicity using this schedule.

High-dose multimodality therapy with autologous stem-cell support for stage IIIB breast carcinoma.

PURPOSE: Women with locally unresectable and inflammatory breast carcinoma (IBC) have an approximately 30% 5-year disease-free survival (DFS) rate with conventional multimodality therapy. A short but dose-intensive multimodality phase II trial was designed in an attempt to improve outcome in stage IIIB disease. Mastectomy was performed after high-dose therapy to evaluate pathologic response to treatment. METHODS: Women with newly diagnosed disease received four 2-week cycles of doxorubicin 90 mg/m2 with granulocyte colony-stimulating factor (G-CSF), followed by cyclophosphamide 6,000 mg/m2, thiotepa 500 mg/m2, and carboplatin 800 mg/m2 (CTCb) with marrow and peripheral-blood progenitor cell (PBPC) support. Local therapy consisted of mastectomy and radiotherapy. Tamoxifen (5 years) was begun if the patient was estrogen receptor-positive (ER+). RESULTS: Fifty women (46 stage IIIB [91% IBC], four stage IIIA) entered the study and 47 are assessable. Ten had mastectomy before any systemic therapy (seven with pathologic IBC, three with residual tumor after mastectomy). Eighty percent received full-dose doxorubicin with 60% on schedule. Clinical response rates to induction were 15% complete response (CR), 5% very good partial response (VGPR), 59% partial response (PR), and 21% minor response (MR)/stable disease (SD). Mastectomy after CTCb in 37 patients showed a 14% pathologic CR rate, 29% microscopic foci in breast and/or axilla, and 57% gross tumor. Fifteen (32%) patients have relapsed (median, 17 months post-CTCb). The 30-month DFS is estimated at 64%. For those in pathologic CR, with microscopic, or with gross disease remaining after CTCb, the 30-month DFS is estimated at 100%, 70%, and 38%, respectively. Those with zero, one to three, or > or = four positive nodes at axillary dissection had a median DFS of 31, 18, and 13 months, respectively. CONCLUSION: This short but dose-intensive multimodality approach for stage IIIB breast carcinoma is feasible with encouraging results to date.

The American Society of Clinical Oncology program to improve end-of-life care.

Too few cancer patients have optimal care at the end of life, as measured by unrelieved pain, death in a setting other than home, and uncoordinated care. The American Society of Clinical Oncology (ASCO), as the largest professional society whose members care for most cancer patients in the United States, has begun an initiative to improve end-of-life care. Educational programs for ASCO members have been started at the national level. A Task Force on End of Life Care was created to study the current state of end-of-life care, barriers to improved end-of-life care, and what ASCO can do to improve end-of-life care. A member-wide survey has been completed to assess, from the professional's perspective, the current state of and barriers to end-of-life care. Specific sections of the Task Force will address hospice care, physician- assisted suicide, clinical barriers, economic barriers, research initiatives, educational deficiencies, and quality of care at the end of life. Improvements in end-of-life care can be made if the current deficiencies are noted, barriers ascertained, and specific solutions found. Professional societies have both the interest and responsibility to improve end-of-life care.

High-dose cyclophosphamide, carmustine, and etoposide with autologous transplantation in Hodgkin's disease: a prognostic model for treatment outcomes.

PURPOSE: To identify clinical factors predictive of treatment outcome after high-dose chemotherapy (HDC) for Hodgkin's disease and to develop a prognostic model for progression-free and overall survival. PATIENTS AND METHODS: 102 patients with relapsed or refractory Hodgkin's disease were treated with high-dose cyclophosphamide, carmustine, and etoposide and autologous marrow and/or peripheral blood progenitor cell support. Median follow-up of survivors is 4.1 years (1.8-7.5 years). Factors potentially important for treatment outcome were examined in univariate analysis, and Cox regression with forward selection was performed. A prognostic model was developed. RESULTS: Poorer progression-free and overall survival were associated with nodular sclerosis histology, abnormal performance status, progressive disease at HDC, more than one extranodal site of disease, and shorter time from initial diagnosis to HDC. These factors and the presence of B symptoms at relapse also predicted for decreased overall survival. Progressive disease immediately prior to HDC, more than one extranodal disease site, and abnormal performance status retained significance for both progression-free and overall survival in multivariate analysis. Progression-free and overall survival are 42% (95% confidence interval, CI, 34 to 53) and 65% (95% CI 54 to 73) at three years. A model based on number of risk factors present divides patients into low, intermediate, and high risk groups with three-year actuarial survival of 82%, 56%, and 19% respectively. Treatment outcome for patients treated with HDC at first chemotherapy relapse was not significantly different from that of the group overall (p > 0.3). CONCLUSIONS: Asymptomatic patients with Hodgkin's disease involving at most one extranodal site whose disease is controlled by conventional dose chemotherapy or radiation therapy at the time of HDC have good outcomes after this therapy. Presence of increasing numbers of risk factors are associated with poorer outcomes. Results of HDC compare favorably to those of standard dose salvage therapy. These data can be used to estimate likely outcomes in patients undergoing HDC for Hodgkin's disease, to identify potential candidates for innovative therapies, and to evaluate strategies for the optimal use of HDC in Hodgkin's disease.

Cloning and characterization of the 5'-flanking sequence for the human DNA topoisomerase II beta gene.

Mammalian cells express two isoforms of type II DNA topoisomerase which are the intracellular targets of many structurally diverse antineoplastic agents. The levels of topoisomerase II isozymes are important determinants for the sensitivity of cells to the cytotoxicity of drugs that target topoisomerase II. To investigate whether the expression of topoisomerase II isoforms is coordinated and the mechanisms governing their expression in the context of drug resistance, the 5'-flanking sequence for the gene of human topoisomerase IIbeta isoform was cloned and characterized. The 5'-flanking region has a very high GC content and contains no canonical TATA box element. Two separate transcriptional start sites are located to an adenine and a guanine, 193 and 89 nucleotides, respectively, upstream from the ATG translation initiation codon. Except for a small region immediately upstream of the translation initiation codon, there is no obvious sequence homology between the 5'-flanking sequences of human topoisomerase IIbeta gene and the previously described alpha gene. Transient expression assays with different 5'- and 3'-deletions of the 5'-flanking region of the topoisomerase IIbeta gene have delineated regions important for transcriptional regulation of the gene. Interestingly, sequences within the first intron also contribute to the promoter activity. Gel mobility shift studies demonstrate that protein factors from the nuclear extracts can bind specifically to the downstream elements and may participate in transcriptional regulation.

Trials of 9-amino-20(S)-camptothecin in Boston.

9-Amino-20(S)-camptothecin (9-AC) is an analog of camptothecin with limited water solubility which has shown significant preclinical activity in a variety of human solid tumor xenografts. A Phase I trial using a soluble formulation of 9-AC, given as a 72-hour continuous infusion, has been completed. Thirty-one patients with resistant cancers received 5-60 micrograms/M2/h at three week intervals. The Maximum Tolerated Dose (MTD) was 45 micrograms/M2/hour. Neutropenia was the dose limiting toxicity, with few significant non-myelosuppressive toxicities. Minor responses were seen in 3/31 patients. Pharmacokinetic studies of 9-AC lactone (closed ring) showed substantial interpatient variability with a predicted half-life of 36 hours. A phase I/II trial of the same formulation of 9-AC is ongoing in refractory leukemia. Stomatitis and diarrhea are the non-myelosuppressive dose limiting toxicities. Evidence of antineoplastic activity has been seen in 3/15 patients. A Phase II trial in previously untreated metastatic breast cancer is also underway. A Phase I trial of a colloidal dispersion formulation, not yet completed, is better tolerated with a MTD > 45 micrograms/M2/h as a 72-hour continuous infusion. Evidence of antineoplastic activity has also been demonstrated.

Double dose-intensive chemotherapy with autologous stem-cell support for metastatic breast cancer: no improvement in progression-free survival by the sequence of high-dose melphalan followed by cyclophosphamide, thiotepa, and carboplatin.

PURPOSE: Twenty-one percent of responding metastatic breast cancer patients remain progression-free a median 50 months following one intensification cycle of cyclophosphamide (6,000 mg/m2), thiotepa (500 mg/ m2), and carboplatin (800 mg/m2) (CTCb) with autologous bone marrow transplantation (ABMT). This trial studied whether the sequence of high-dose melphalan followed by CTCb resulted in improved disease response and duration. METHODS: Women with at least partial responses (PRS) to induction received melphalan (140 or 180 mg/ m2) with peripheral-blood progenitor cell (PBPC) and granulocyte colony-stimulating factor (G-CSF) support. They were monitored as outpatients. After recovery, patients were hospitalized for CTCb with marrow, PBPC, and G-CSF support. RESULTS: Data on 67 women, at a median of 25 months from CTCb, were examined. After melphalan, 49 (73%) required admission for fever (89%), mucositis (35%), or infection (15%) (median stay, 8 days). All received CTCb. For the first 33 patients, the median days from start of melphalan to CTCb was 24. After liver toxicity (one death from venoocclusive disease [VOD]) developed in 11 patients during CTCb, the interval between intensifications was increased to 35 days without incident. Twenty-three patients (34%) are progression-free a median of 16 months post-CTCb. The median progression-free survival (PFS) and survival times for the whole group are estimated at 11 and 20 months, respectively. CONCLUSION: Treatment with this sequence of high-dose melphalan followed by CTCb has not resulted in superior PFS to date, when compared with single-intensification CTCb. This report discusses factors related to patient selection, the role of induced drug resistance, and the schedule of administration of alkylating agenting that may adversely influence outcome.

Passage to nonselective media transiently alters growth of mycophenolic acid-resistant mammalian cells expressing the escherichia coli xanthine-guanine phosphoribosyltransferase gene: implications for sequential selection strategies.

The Escherichia coli xanthine-guanine phosphoribosyltransferase gene (Ecogpt) rescues mammalian cells from inhibition of purine nucleotide biosynthesis by mycophenolic acid (MPA). We used Ecogpt and other selectable markers to obtain subclones of NIH 3T3 derivatives (EN/NIH) stably expressing transfected genes of interest. In their respective selective mediums, growth of MPA-resistant (MPA(R)) isolates was indistinguishable from that of aminoglycoside-resistant counterparts expressing selectable marker genes conferring resistance to protein synthesis inhibitors hygromycin B, puromycin, and G418. Growth of aminoglycoside-resistant isolates remained unaltered on passage to nonselective media. In contrast, MPA(R) cells transferred from MPA complete media to nonselective media displayed morphologic changes with static growth. These findings resolved completely by third passage in nonselective media and were independent of the gene of interest cis-linked to the selectable marker. Sequential selection strategies involving cell culture conditions resulting in these altered growth characteristics significantly impaired detection (by selection in G418) of genomic events associated with reactivation of enhancerless, transcriptionally silent neointegrants present in MPA(R) EN/NIH isolates. We explored the cause of these cell culture findings and defined transfection and sequential selection strategies for MPA(R) derivatives that successfully circumvented these effects.

DNA topoisomerase II alpha expression is associated with alkylating agent resistance.

Increased expression of DNA topoisomerase II alpha has been associated with resistance to certain DNA-damaging alkylating agents, but no causal relationship or mechanism has been established. To investigate this observation, we developed a model of topoisomerase II overexpression by transfecting a full-length Chinese hamster ovary topoisomerase II alpha into EMT6 mouse mammary carcinoma. Topoisomerase II alpha-transfected cell lines demonstrated continued topoisomerase II alpha mRNA and protein expression, which were undetectable in vector-only lines, in stationary phase (G0-G1). The topoisomerase II transfectants were approximately 5-10-fold resistant to the alkylating agents cisplatin and mechlorethamine. Upon release from G0-G1, the topoisomerase II transfectants demonstrated more rapid thymidine incorporation and shorter cell-doubling times than control cells. Purified topoisomerase II and nuclear extracts with topoisomerase II-decatenating activity bound to cisplatin-treated DNA with significantly greater affinity than to untreated DNA in a cisplatin concentration-dependent manner. These observations suggest that expression of topoisomerase II alpha may have a role in cellular resistance to antineoplastic alkylating agents. The mechanism for this may involve increased binding of topoisomerase II alpha to alkylating agent-damaged DNA.

Molecular cloning and characterization of the promoter for the Chinese hamster DNA topoisomerase II alpha gene.

To investigate the mechanisms governing the expression of DNA topoisomerase II alpha, the Chinese hamster topoisomerase II alpha gene has been cloned and the promoter region analyzed. There are several transcriptional start sites clustered in a region of 30 base pairs, with the major one being 102 nucleotides upstream from the ATG translation initiation site. Sequencing data reveal one GC box and a total of five inverted CCAAT elements (ICEs) within a region of 530 base pairs upstream from the major transcription start site. Sequence comparison between the human and Chinese hamster topoisomerase II alpha gene promoter regions shows a high degree of homology centered at the ICEs and GC box. In vitro DNase I footprinting results indicate protection by binding proteins at and around each ICE on both DNA strands. However, no obvious protection was observed for the GC box. Competition gel mobility shift assays with oligonucleotides containing either the wild-type or mutated ICE sequences suggest that identical or similar proteins specifically bind at each ICE, although with different affinities for individual ICE sequences. Chloramphenicol acetyltransferase assays employing nested 5'-deletions of the 5'-flanking sequence of the gene demonstrate that the sequence between -186 and +102, which contains three proximal ICEs, is sufficient for near wild-type level of promoter activity. When these three ICEs were gradually replaced with sequences which do not interact with the binding proteins, reducing promoter activity of the resulted constructs was observed. In conjunction with results from footprinting and gel mobility shift studies, the transient gene expression finding suggests that the ICEs are functionally important for the transcriptional regulation of the topoisomerase II alpha gene.

Prognostic factors for prolonged progression-free survival with high-dose chemotherapy with autologous stem-cell support for advanced breast cancer.

PURPOSE: With a median observation time of 50 months from transplant, 13 (22%) of 62 women with metastatic breast cancer treated with high-dose chemotherapy at the Dana-Farber Cancer Institute (DFCI)/Beth Israel Hospital (BIH) remain progression-free. This study determined factors prognostic for prolonged progression-free survival (PFS). METHODS: From June 1988 to January 1992, women who responded to standard chemotherapy received high-dose cyclophosphamide, thiotepa, and carboplatin with autotransplantation. Data encompassing initial breast cancer diagnosis, metastatic presentation, and response to induction treatment were examined for correlations with improved PFS. RESULTS: The 5-year PFS rate for the entire group is estimated to be 21% (95% confidence interval [CI], 10% to 32%). For those patients who attained a complete response (CR) to induction therapy, the 5-year PFS rate is estimated to be 31% (95% CI, 0% to 63%). In univariate analyses, a single metastatic site, CR to induction therapy, prolonged interval from primary diagnosis to first metastases, estrogen receptor (ER)-negative tumors, and older age (> or = 40 years) were associated with prolonged PFS. In multivariate analyses, single metastatic site (P = .002) and attainment of a CR to induction chemotherapy (P = .04) were the most significant predictors for PFS, with a strong trend observed for an interval from primary diagnosis to onset of metastatic disease of 24+ months (P = .066). CONCLUSION: We and others have shown that 10% to 25% of women with metastatic breast cancer are progression-free after high-dose chemotherapy with autotransplantation. Those with chemosensitive disease, minimal tumor bulk, and a prolonged disease-free interval appear to benefit most. Emphasis should continue to focus on the development of more effective cytotoxic regimens and biologic approaches to increase the percentage of patients who may benefit from this approach.