The FACIT-AI, a new tool for assessing symptoms associated with malignant ascites.

OBJECTIVE: The objectives of this study are to assess the clinical relevance and validity of the Functional Assessment of Chronic Illness Therapy-Ascites Index (FACIT-AI) in women with ovarian cancer and malignant ascites, and to modify the instrument guided by qualitative feedback from patients with recurrent malignant ascites. METHODS: Fourteen adult female patients with recurrent symptomatic malignant ascites were enrolled from three centers. All completed an open-ended symptom list to identify their primary concerns regarding their condition. They then completed a draft 10-item FACIT-AI questionnaire created from expert input. Eleven patients provided comments regarding the FACIT-AI questionnaire using a written feedback format. Three patients participated in a "think-aloud" cognitive debriefing interview to ensure patient comprehension of questionnaire items. RESULTS: Of the first 11 patients surveyed, 7 believed that the draft FACIT-AI contained all important symptoms associated with malignant ascites. Responses from the remaining 4 patients revealed three symptoms that 2 or more patients nominated for inclusion: urinary frequency, constipation and emotional distress. These items were added to the original FACIT-AI to produce a 13-item index of symptoms associated with malignant ascites. CONCLUSIONS: The 13-item FACIT-AI has content validity among women with malignant ascites associated with ovarian cancer. It is available for use in clinical research or practice, with the expectation that more will be learned about its performance and interpretation over time.

Patient Preferences for Multi-Cancer Early Detection (MCED) Screening Tests.

BACKGROUND: Emerging blood-based multi-cancer early detection (MCED) tests can detect a variety of cancer types across stages with a range of sensitivity, specificity, and ability to predict the origin of the cancer signal. However, little is known about the general US population's preferences for MCED tests. OBJECTIVE: To quantify preferences for MCED tests among US adults aged 50-80 years using a discrete choice experiment (DCE). METHODS: To quantify preferences for attributes of blood-based MCED tests, an online DCE was conducted with five attributes (true positives, false negatives, false positives, likelihood of the cancer type unknown, number of cancer types detected), among the US population aged 50-80 years recruited via online panels and social media. Data were analyzed using latent class multinomial logit models and relative attribute importance was obtained. RESULTS: Participants (N = 1700) were 54% female, mean age 63.3 years. Latent class modeling identified three classes with distinct preferences for MCED tests. The rank order of attribute importance based on relative attribute importance varied by latent class, but across all latent classes, participants preferred higher accuracy (fewer false negatives and false positives, more true positives) and screenings that detected more cancer types and had a lower likelihood of cancer type unknown. Overall, 72% of participants preferred to receive an MCED test in addition to currently recommended cancer screenings. CONCLUSIONS: While there is significant heterogeneity in cancer screening preferences, the majority of participants preferred MCED screening and the accuracy of these tests is important. While the majority of participants preferred adding an MCED test to complement current cancer screenings, the latent class analyses identified a small (16%) and specific subset of individuals who value attributes differently, with particular concern regarding false-negative and false-positive test results, who are significantly less likely to opt-in.

Performance of a Cell-Free DNA-Based Multi-cancer Detection Test in Individuals Presenting With Symptoms Suspicious for Cancers.

PURPOSE: A multi-cancer detection test using a targeted methylation assay and machine learning classifiers was validated and optimized for screening in prospective, case-controlled Circulating Cell-free Genome Atlas (ClinicalTrials.gov identifier: NCT02889978) substudy 3. Here, we report test performance in a subgroup of participants with symptoms suspicious for cancer to assess the test's ability to potentially facilitate efficient diagnostic evaluation in symptomatic individuals. METHODS: We evaluated test performance (sensitivity, specificity, and accuracy of cancer signal origin [CSO] prediction accuracy) in participants with clinically presenting cancers (CPCs) and noncancer with underlying medical conditions and among two subgroups (65 years and older and GI cancers). Overall survival (OS) of participants who had a cancer signal detected/not detected was compared with SEER-based expected survival. RESULTS: A total of 2,036 cancer and 1,472 noncancer participants were included. Specificity was high in all noncancer participants (99.5% [95% CI, 98.4 to 99.8]). In participants with CPCs, the overall sensitivity was 64.3% (95% CI, 62.2 to 66.4) and the overall accuracy of CSO prediction in true positives was 90.3%. For GI cancers, the overall sensitivity was 84.1% (95% CI, 80.6 to 87.1). In participants 65 years and older, test performance was similar to that of all participants. Individuals with cancers not detected had a significantly better OS than that expected from SEER (P < .01). CONCLUSION: This test detected a cancer signal with high specificity and CSO prediction accuracy and moderate sensitivity in symptomatic individuals, with especially high performance in participants with GI cancers. The survival analysis implied that the cancers not detected were less clinically aggressive than cancers detected by the test, providing prognostic insights to physicians. This multi-cancer detection test could facilitate efficient workup and stratify cancer risk in symptomatic individuals.

Metastatic mucinous ovarian cancer and treatment decisions based on histology and molecular markers rather than the primary location.

Approximately 22,000 cases of ovarian cancer occur each year in the United States, and likely fewer than 2000 cases of mucinous ovarian cancers. Although 90% of patients with mucinous ovarian cancer present with stage I disease and have curative surgeries, advanced-stage disease is known to have a poor response to standard platinum- and taxane-based chemotherapy. Despite limited enthusiasm, standard chemotherapy is still recommended for most patients with advanced-stage mucinous malignancies of the ovary. This report presents an unusual case of a woman with HER2-positive metastatic mucinous carcinoma of the ovary treated with chemotherapy regimens typically used for colorectal malignancies, followed by epidermal growth factor receptor-targeted therapies.

Evaluation of cell-free DNA approaches for multi-cancer early detection.

In the Circulating Cell-free Genome Atlas (NCT02889978) substudy 1, we evaluate several approaches for a circulating cell-free DNA (cfDNA)-based multi-cancer early detection (MCED) test by defining clinical limit of detection (LOD) based on circulating tumor allele fraction (cTAF), enabling performance comparisons. Among 10 machine-learning classifiers trained on the same samples and independently validated, when evaluated at 98% specificity, those using whole-genome (WG) methylation, single nucleotide variants with paired white blood cell background removal, and combined scores from classifiers evaluated in this study show the highest cancer signal detection sensitivities. Compared with clinical stage and tumor type, cTAF is a more significant predictor of classifier performance and may more closely reflect tumor biology. Clinical LODs mirror relative sensitivities for all approaches. The WG methylation feature best predicts cancer signal origin. WG methylation is the most promising technology for MCED and informs development of a targeted methylation MCED test.

Immunologic and clinical effects of antibody blockade of cytotoxic T lymphocyte-associated antigen 4 in previously vaccinated cancer patients.

Cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) functions as a negative regulator of endogenous and vaccine-induced antitumor immunity. The administration of fully human anti-CTLA-4 blocking monoclonal antibodies to advanced-cancer patients increases immune-mediated tumor destruction in some subjects. Nonetheless, patients that respond also frequently manifest serious inflammatory pathologies, raising the possibility that the therapeutic and toxic effects of CTLA-4 blockade might be linked. Here we show that periodic infusions of anti-CTLA-4 antibodies after vaccination with irradiated, autologous tumor cells engineered to secrete GM-CSF (GVAX) generate clinically meaningful antitumor immunity without grade 3 or 4 toxicity in a majority of metastatic melanoma patients. The application of this sequential immunotherapy to advanced ovarian carcinoma patients also revealed that tumor destruction and severe inflammatory pathology could be dissociated, although further refinements are required to increase clinical responses and to minimize toxicity in this population. The extent of therapy-induced tumor necrosis was linearly related to the natural logarithm of the ratio of intratumoral CD8(+) effector T cells to FoxP3(+) regulatory T cells (Tregs) in posttreatment biopsies. Together, these findings help clarify the immunologic and clinical effects of CTLA-4 antibody blockade in previously vaccinated patients and raise the possibility that selective targeting of antitumor Tregs may constitute a complementary strategy for combination therapy.

Prognostic Significance of Blood-Based Multi-cancer Detection in Plasma Cell-Free DNA.

PURPOSE: We recently reported the development of a cell-free DNA (cfDNA) targeted methylation (TM)-based sequencing approach for a multi-cancer early detection (MCED) test that includes cancer signal origin prediction. Here, we evaluated the prognostic significance of cancer detection by the MCED test using longitudinal follow-up data. EXPERIMENTAL DESIGN: As part of a Circulating Cell-free Genome Atlas (CCGA) substudy, plasma cfDNA samples were sequenced using a TM approach, and machine learning classifiers predicted cancer status and cancer signal origin. Overall survival (OS) of cancer participants in the first 3 years of follow-up was evaluated in relation to cancer detection by the MCED test and clinical characteristics. RESULTS: Cancers not detected by the MCED test had significantly better OS (P < 0.0001) than cancers detected, even after accounting for other covariates, including clinical stage and method of clinical diagnosis (i.e., standard-of-care screening or clinical presentation with signs/symptoms). Additionally, cancers not detected by the MCED test had better OS than was expected when data were adjusted for age, stage, and cancer type from the Surveillance, Epidemiology, and End Results (SEER) program. In cancers with current screening options, the MCED test also differentiated more aggressive cancers from less aggressive cancers (P < 0.0001). CONCLUSIONS: Cancer detection by the MCED test was prognostic beyond clinical stage and method of diagnosis. Cancers not detected by the MCED test had better prognosis than cancers detected and SEER-based expected survival. Cancer detection and prognosis may be linked by the underlying biological factor of tumor fraction in cfDNA.

CD133 expression defines a tumor initiating cell population in primary human ovarian cancer.

Evidence is accumulating that solid tumors contain a rare phenotypically distinct population of cells, termed cancer stem cells (CSC), which give rise to and maintain the bulk of the tumor. These CSC are thought to be resistant to current chemotherapeutic strategies due to their intrinsic stem-like properties and thus may provide the principal driving force behind recurrent tumor growth. Given the high frequency of recurrent metastasis associated with human ovarian cancer, we sought to determine whether primary human ovarian tumors contain populations of cells with enhanced tumor-initiating capacity, a characteristic of CSC. Using an in vivo serial transplantation model, we show that primary uncultured human ovarian tumors can be reliably propagated in NOD/SCID mice, generating heterogeneous tumors that maintain the histological integrity of the parental tumor. The observed frequency of tumor engraftment suggests only certain subpopulations of ovarian tumor cells have the capacity to recapitulate tumor growth. Further profiling of human ovarian tumors for expression of candidate CSC surface markers indicated consistent expression of CD133. To determine whether CD133 expression could define a tumor-initiating cell population in primary human ovarian tumors, fluorescence-activated cell sorting (FACS) methods were employed. Injection of sorted CD133(+) and CD133(-) cell populations into NOD/SCID mice established that tumor-derived CD133(+) cells have an increased tumorigenic capacity and are capable of recapitulating the original heterogeneous tumor. Our data indicate that CD133 expression defines a NOD/SCID tumor initiating subpopulation of cells in human ovarian cancer that may be an important target for new chemotherapeutic strategies aimed at eliminating ovarian cancer.

Phase II trial of GM-CSF in women with asymptomatic recurrent müllerian tumors.

OBJECTIVE: GM-CSF is a recombinant human cytokine, which promotes the proliferation and differentiation of granulocytes and monocytes, and is associated with anti-tumor activity. The primary objective was to define the median time to treatment termination (TTT) with women with relapsed ovarian cancer treated with single agent GM-CSF delivered subcutaneously (SC). PATIENTS AND METHODS: Open label phase II study in asymptomatic patients with recurrent müllerian malignancy without an indication for immediate systemic chemotherapy. In the first cohort of 35 women, GM-CSF 250 microg/m(2) was administered SC on days 1-14 of a 28-day cycle, the second cohort received continuous GM-CSF 150 microg/m(2) given with dose escalation. RESULTS: Seventy-two women were enrolled. Best overall response included one complete response, and 20 patients with stable disease (23%), 4 of whom had stable disease for >6 months. Median TTT was 78 days. Toxicity in both cohorts was generally mild; however, four patients experienced excessive toxicity and withdrew consent. In the first cohort, CA-125 dropped in 70% of women from their baseline on study value (median change -23%, range -48 to +116%) after 14 days of GM-CSF. The magnitude of CA-125 drop during the first 2 weeks of therapy also showed a positive inverse correlation with day 15 white cell count for the whole group (p=0.038). CONCLUSION: GM-CSF is well tolerated and frequently associated with a decline in CA-125 that is correlated with leukocytosis. Although median TTT is modest, a subset of women had prolonged stable disease.

Risk stratification for desensitization of patients with carboplatin hypersensitivity: clinical presentation and management.

BACKGROUND: Women with ovarian cancer treated with chemotherapeutic platinum agents frequently develop hypersensitivity reactions (HSRs). How best to risk-stratify patients for desensitization is uncertain. OBJECTIVES: To evaluate skin test (ST) reactivity to carboplatin in patients with recent and remote histories of carboplatin HSR and to review the relationship between skin test reactivity and tolerance of subsequent carboplatin desensitization. METHODS: Thirty-eight women with carboplatin HSR were evaluated by ST to carboplatin. Thirty women subsequently underwent 106 desensitizations to carboplatin. RESULTS: Carboplatin ST was positive in 25 of 38 patients (66%). Of patients with recent HSR (<3 months), 20 of 24 (83%) tested positive, whereas 5 of 14 (36%) with remote HSR (>9 months) tested positive (P < .01). Nineteen carboplatin ST+ and 11 ST- patients underwent desensitization to carboplatin. Seven ST+ patients (37%) had mild HSR during desensitization but completed the desensitization with additional treatment or protocol modification. ST- patients with a recent history of HSR (n = 3) tolerated a rapid protocol without HSR and remained ST- with repeated testing. Six of 8 ST- patients (75%) with remote HSR reacted during desensitization. The HSRs were more severe and often associated with an elevated tryptase level. Five of 7 patients retested became ST+ before the second desensitization. Carboplatin desensitization was successfully completed in 105 of 106 (99%) treatment courses. CONCLUSIONS: The timing of carboplatin ST in relation to initial HSR is vital for risk stratification and subsequent desensitization. Initial ST- patients with a remote history of HSR are at high risk for conversion to ST+ and can develop more severe HSR.

Modulation of drug resistance in ovarian adenocarcinoma by enhancing intracellular ceramide using tamoxifen-loaded biodegradable polymeric nanoparticles.

PURPOSE: To modulate intracellular ceramide levels and lower the apoptotic threshold in multidrug-resistant ovarian adenocarcinoma, we have examined the efficacy and preliminary safety of tamoxifen coadministration with paclitaxel in biodegradable poly(ethylene oxide)-modified poly(epsilon-caprolactone) (PEO-PCL) nanoparticles. EXPERIMENTAL DESIGN: In vitro cytotoxicity and proapoptotic activity of paclitaxel and tamoxifen, either as single agent or in combination, was examined in wild-type (SKOV3) and MDR-1-positive (SKOV3TR) human ovarian adenocarcinoma cells. Subcutaneous SKOV3 and SKOV3TR xenografts were established in female nu/nu mice, and this model was used to evaluate the antitumor efficacy and preliminary safety. Paclitaxel (20 mg/kg) and tamoxifen (70 mg/kg) were administered i.v. either as a single agent or in combination in aqueous solution and in PEO-PCL nanoparticles. RESULTS: In vitro cytotoxicity results showed that administration of paclitaxel and tamoxifen in combination lowered the IC50 of paclitaxel by 10-fold in SKOV3 cells and by >3-fold in SKOV3TR cells. The combination paclitaxel/tamoxifen co-therapy showed even more pronounced effect when administered in nanoparticle formulations. Upon i.v. administration of paclitaxel/tamoxifen combination in PEO-PCL nanoparticle formulations, significant enhancement in antitumor efficacy was observed. Furthermore, the combination paclitaxel/tamoxifen therapy did not induce any acute toxicity as measured by body weight changes, blood cell counts, and hepatotoxicity. CONCLUSIONS: The results of this study show that combination of paclitaxel and tamoxifen in biodegradable PEO-PCL nanoparticles can serve as an effective clinically translatable strategy to overcome multidrug resistance in ovarian cancer.

Modulation of intracellular ceramide using polymeric nanoparticles to overcome multidrug resistance in cancer.

Although multidrug resistance (MDR) is known to develop through a variety of molecular mechanisms within the tumor cell, many tend to converge toward the alteration of apoptotic signaling. The enzyme glucosylceramide synthase (GCS), responsible for bioactivation of the proapoptotic mediator ceramide to a nonfunctional moiety glucosylceramide, is overexpressed in many MDR tumor types and has been implicated in cell survival in the presence of chemotherapy. The purpose of this study was to investigate the therapeutic strategy of coadministering ceramide with paclitaxel, a commonly used chemotherapeutic agent, in an attempt to restore apoptotic signaling and overcome MDR in the human ovarian cancer cell line SKOV3. Poly(ethylene oxide)-modified poly(epsilon-caprolactone) (PEO-PCL) nanoparticles were used to encapsulate and deliver the therapeutic agents for enhanced efficacy. Results show that indeed the cotherapy eradicates the complete population of MDR cancer cells when they are treated at their IC(50) dose of paclitaxel. More interestingly, when the cotherapy was combined with the properties of nanoparticle drug delivery, the MDR cells can be resensitized to a dose of paclitaxel near the IC(50) of non-MDR (drug sensitive) cells, indicating a 100-fold increase in chemosensitization via this approach. Molecular analysis of activity verified the hypothesis that the efficacy of this therapeutic approach is indeed due to a restoration in apoptotic signaling, although the beneficial properties of PEO-PCL nanoparticle delivery seemed to enhance the therapeutic success even further, showing the promising potential for the clinical use of this therapeutic strategy to overcome MDR.

Phase I clinical trial of STA-4783 in combination with paclitaxel in patients with refractory solid tumors.

PURPOSE: STA-4783 is a new compound that markedly enhances the therapeutic index of paclitaxel against human tumor xenograft models. A phase I clinical trial was undertaken to determine the maximum tolerated dose, toxicity profile, and pharmacokinetics of STA-4783 in combination with paclitaxel. EXPERIMENTAL DESIGN: Adults with refractory solid tumors concurrently received STA-4783 and paclitaxel as a 3-h i.v. infusion at starting doses of 44 and 135 mg/m(2), respectively. After increasing paclitaxel to 175 mg/m(2), the STA-4783 dose was escalated as permitted by dose-limiting toxicity during the first 21-day cycle. RESULTS: Thirty-five patients were treated with eight dose levels of STA-4783/paclitaxel. In patients receiving 175 mg/m(2) paclitaxel, the incidence of severe toxicity increased with escalation of the STA-4783 dose above 263 mg/m(2), and 438 mg/m(2) was the maximum tolerated dose. All toxicities were typical of paclitaxel, with neutropenia, mucositis, and myalgia/arthralgia being dose limiting. Partial responses were achieved in one patient with Kaposi's sarcoma and another with ovarian cancer that progressed during prior treatment with paclitaxel. STA-4783 exhibited linear pharmacokinetics characterized by rapid elimination from plasma (biological half-life, 1.06 +/- 0.24 h) and a low steady-state apparent volume of distribution (25.1 +/- 8.1 L/m(2)). The total body clearance of paclitaxel decreased significantly with escalation of the STA-4783 dose. CONCLUSIONS: The STA-4783/paclitaxel combination was well tolerated with a toxicity profile similar to single-agent paclitaxel. Enhanced systemic exposure to paclitaxel resulting from a dose-dependent interaction with STA-4783 was associated with increased toxicity. Objective responses in two heavily pretreated patients, both with taxane exposure, have encouraged further clinical evaluation of this regimen.

Pooling of case specimens to create standard serum sets for screening cancer biomarkers.

BACKGROUND: Multiple identical sets of sera from cancer cases and controls would facilitate standardized testing of biomarkers. We describe the creation and use of standard serum sets developed from healthy donors and pooled sera from ovarian, breast, and endometrial cancer cases. METHODS: Two hundred seventy-five 0.3-mL aliquots of sera were created for each of the 95 healthy women, and residual serum was pooled to create 275 identical sets of 20 0.3-mL aliquots. Aliquots (1.0-1.5 mL) from 441 women were combined to create 12 breast and pelvic disease pools with at least 115 0.3-mL aliquots. Sets were assembled to contain aliquots from individual controls, replicates, and disease pools. Cancer antigens (CA), CA 125, CA 19.9, and CA 15.3, and carcinoembryonic antigen were measured in one set and in 217 women comprising six of the pelvic disease pools. Use of a set was illustrated for mesothelin (soluble mesothelin-related protein). Statistical output included concentration differences between pooled cases and controls (z values for single analytes; Mahalanobis distances for pairs), correlation between z values and sensitivities, coefficient of variations, and standardized biases. RESULTS: Marker concentrations in the six pelvic disease pools were generally within 0.25 SD of the actual average, and z values correlated well with sensitivities. CA 125 remains the best single marker for nonmucinous ovarian cancer, complemented by CA 15.3 or soluble mesothelin-related protein. There is no comparable breast cancer biomarker among the current analytes tested. CONCLUSION: The potential value of standard serum sets for initial assessment of candidate biomarkers is illustrated. Sets are now available through the Early Detection Research Network to evaluate biomarkers for women's cancers.

Phase II study of intraperitoneal recombinant interleukin-12 (rhIL-12) in patients with peritoneal carcinomatosis (residual disease < 1 cm) associated with ovarian cancer or primary peritoneal carcinoma.

BACKGROUND: Pharmacokinetic advantages of intraperitoneal (IP) rhIL-12, tumor response to IP delivery of other cytokines as well as its potential anti-angiogenic effect provided the rationale for further evaluation of IPrhIL-12 in patients with persistent ovarian or peritoneal carcinoma. METHODS: A phase 2 multi-institutional trial (NCI Study #2251) of IP rIL-12 (300 nanogram/Kg weekly) was conducted in patients with ovarian carcinoma or primary peritoneal carcinoma. Patients treated with primary therapy for ovarian cancer who had no extraabdominal/parenchymal disease or bulky peritoneal disease were eligible. Four to 8 weeks from last chemotherapy, eligible patients underwent a laparotomy/laparoscopy. Patients with residual disease < or = 1 cm were registered for the treatment phase 2-5 weeks post surgery. The effect of IP rIL-12 on the expression of TNFalpha , INFalpha , IL-10, IP-10, IL-8, FGF, VEGF was also studied. RESULTS: Thirty-four patients were registered for the first screening phase of the study. Median age was 56.6 years (range: 31-71); 12 completed the second phase and were evaluable for response/toxicity. Performance scores of IL-12 treated patients were 0 (11 pts) and 1 (1 pt). There were no treatment related deaths, peritonitis or significant catheter related complications. Toxicities included grade 4 neutropenia (1), grade 3 fatigue (4), headache (2), myalgia (2), non-neutropenic fever (1), drug fever (1), back pain (1), and dizziness (1). The best response observed was SD. Two patients had SD and 9 had PD, and 1 was evaluable for toxicity only. Peritoneal fluid cytokine measurements demonstrated a > or = 3 fold relative increase post-rhIL-12: IFN-gamma, 5/5 pts; TNF-alpha , 1/5; IL-10, 4/5; IL-8, 5/5; and VEGF, 3/5. IP10 levels were increased in 5/5 patients. Cytokine response profiles suggest either NK or T-cell mediated effects of IP rhIL-12. Cytokine/chemokine results also suggest a pleiotropic response since proteins with potential for either anti-tumor (IFN-gamma , IP-10) or pro-tumor growth effects (VEGF, IL-8) were detected. CONCLUSION: IP IL-12 can safely be administered at this dose and schedule to patients after first line chemotherapy for ovarian/peritoneal carcinoma. The maximum response was stable disease. Future IP therapies with rhIL-12 will require better understanding and control of pleiotropic effects of IL-12.

SD-1029 inhibits signal transducer and activator of transcription 3 nuclear translocation.

PURPOSE: Signal transducer and activator of transcription 3 (Stat3) proteins have important roles in cancer cell survival and proliferation. Recent studies show that aberrant Stat3 activation promotes tumor growth and survival in several human cancers, and thus, presents an attractive pathway for the development of targeted anticancer therapy. Stat3 is a DNA-binding transcription factor, and thus, its function depends on cytoplasmic to nuclear translocation. To discover novel inhibitors of the Stat3 signaling pathway, we designed a cell-based screening assay capable of identifying compounds that inhibit Stat3 nuclear translocation and activity. EXPERIMENTAL DESIGN: Cell-based fluorescence microscope screening and quantitative measurement of enhanced green fluorescent protein-Stat3 nuclear translocation assays were used to identify novel Stat3 inhibitors. The effects of identified Stat3 inhibitors on Janus kinase (Jak), Stat3 expression, and activation were determined by Western blotting and kinase in vitro autophosphorylation assay. The effects of identified Stat3 inhibitors on cell growth was evaluated by cell proliferation assay and apoptosis assay. RESULTS: Among the National Cancer Institute Diversity set, a 2,000-member library of bioactive small molecules, we identified SD-1029 as a micromolar inhibitor of IL-6 or oncostatin-induced Stat3 nuclear translocation. Biochemical analysis shows that SD-1029 inhibits tyrosyl phosphorylation of Stat3 implicating SD-1029 as an inhibitor of Jak. Further analysis shows that this compound inhibits tyrosyl phosphorylation of the Jak2 isoenzyme. The antiapoptotic proteins Bcl-X(L) and survivin, target proteins of activated Stat3, are down-regulated by SD-1029 resulting in the induction of apoptosis in several human breast and ovarian cancer cell lines. SD-1029 also enhances apoptosis induced by paclitaxel in ovarian cancer cells. CONCLUSIONS: These results show that SD-1029 directly abrogates the Jak-Stat3 signaling pathway in human cancer cells expressing constitutively active Stat, and add to the growing literature that validates this pathway as a viable target for further drug development. Finally, SD-1029 may represent a suitable prototype for structural optimization and exploration as a therapeutic lead.

Signal transducers and activators of transcription 3 pathway activation in drug-resistant ovarian cancer.

PURPOSE: One of the major obstacles in the treatment of ovarian cancer is the development of multidrug resistance. Recent evidence shows that high-grade ovarian cancer often shows activation of the signal transducers and activators of transcription 3 (Stat3) pathway with subsequent transcription of genes that support tumor growth and survival. Less studied is the role of the Stat3 pathway in acquired drug resistance. There is no information on Stat3 expression in chemotherapy naïve ovarian cancer as compared with tumors collected later in the natural history of the disease. To further clarify the significance of Stat3 activation in ovarian cancer, here we investigated the Stat3 expression and activation in ovarian cancer and ovarian cancer multidrug resistance cell lines. EXPERIMENTAL DESIGN: Western blotting, electrophoretic mobility shift assay, luciferase assays, ELISA assay, and real-time reverse transcription-PCR determined interleukin-6 and Stat3 pathway expression and activation in cell lines. Stat3 expression in ovarian cancer tissue microarray was evaluated by immunohistochemistry. RESULTS: Activated (phosphorylated) Stat3 is overexpressed in most paclitaxel-resistant ovarian cancer cells. Inhibition of Stat3 activation results in significant decreases in paclitaxel resistance and enhanced apoptosis. Drug-resistant recurrent tumors have significantly greater phosphorylated Stat3 (pStat3) expression as compared with matched primary tumors. Tumors with associated inflammatory cell infiltrates also have a higher proportion of cells staining intensely for nuclear phosphorylated Stat3 as compared with tumors without inflammatory infiltrates, consistent with paracrine activation of the Stat3 pathway by immune-mediated cytokines. CONCLUSIONS: These data support the hypothesis that interruption of Stat3 signaling could reverse resistance to paclitaxel and perhaps other chemotherapy agents in human cancer.

Chemosensitization to cisplatin by inhibitors of the Fanconi anemia/BRCA pathway.

Cisplatin resistance occurs, at least in part, through the function of the Fanconi anemia (FA)/BRCA pathway, a DNA-damage response pathway required for repair of cisplatin cross-links. In the current study, we designed a cell-based screening strategy to identify small-molecule inhibitors of the FA/BRCA pathway with the hypothesis that such molecules could restore sensitivity to platinum agents. We identified four inhibitors, including three protein kinase inhibitors (wortmannin, H-9, and alsterpaullone) and one natural compound (curcumin) that inhibit the FA/BRCA pathway. We show that curcumin, a compound that is generally regarded as safe, inhibits the monoubiquitination of the FANCD2 protein as predicted by the screen and consequently sensitizes ovarian and breast tumor cell lines to cisplatin through apoptotic cell death. We believe that this study shows an efficient, high-throughput method for identifying new compounds that may sensitize cancer cells to DNA-damaging chemotherapy.

Multicenter, randomized phase II trial of oral CI-1033 for previously treated advanced ovarian cancer.

PURPOSE: To evaluate the antitumor activity and toxicity of two doses of CI-1033 in patients with platinum-refractory or recurrent ovarian cancer, and to determine baseline expression of epidermal growth factor receptor in tumor cells. PATIENTS AND METHODS: This phase II, open-label clinical trial evaluated CI-1033 in patients with ovarian cancer who failed prior platinum-based therapy. Two oral doses of CI-1033 were evaluated--a 50-mg and a 200--mg oral dose administered daily for 21 days in a 28-day cycle. Patients were evaluated for tumor response and toxicity; in addition, archival baseline tumor samples were analyzed by immunohistochemistry for erbB1 to erbB4 status. RESULTS: One hundred five eligible patients were treated. Baseline demographic characteristics were balanced in this heavily pretreated patient population. The median number of prior chemotherapy regimens received was four. The most commonly encountered drug-related adverse events for both dose arms were gastrointestinal (diarrhea, nausea, stomatitis) toxicity, asthenia, and rash. No responses were observed. Stable disease was confirmed in 34% and 26% of patients in the 200-mg and 50-mg arms, respectively, and 1-year survival rates were 38.5% and 37.7%, respectively. Baseline erbB3 and erbB4 revealed the highest frequencies of expression, while erbB2 was the lowest. CONCLUSION: CI-1033 did not show activity in unscreened patients with advanced ovarian cancer. At 50 mg/d, CI-1033 had a more favorable adverse events profile than at 200 mg/d. erbB3 and erbB4 receptors showed the highest expression in tumor samples while erbB2 revealed the least. There appears to be no association between baseline erbB expression and disease stability.

GBP1 overexpression is associated with a paclitaxel resistance phenotype.

In the search for novel genes involved in the paclitaxel resistance phenotype, prior studies of gene expression in paclitaxel-resistant cell lines and their paired drug-sensitive parental lines using high-density Affymetrix GeneChip arrays identified guanylate-binding protein 1 (GBP1) gene as an overexpressed transcript. The GBP1 gene encodes a large GTPase that is induced by interferon gamma (IFN-gamma) in a variety of eukaryotic cells. In this report we characterize GBP1 and demonstrate that GBP1 expression is consistently upregulated in 7 of 8 paclitaxel or doxorubicin-resistant human cancer cell lines as compared to its expression in the relevant drug-sensitive parental lines. Analysis of GBP1 expression using the Cancer Profiling Array showed that GBP1 is ubiquitously expressed with no significant difference in expression levels between normal and tumor tissue. Parallel analysis of the Cancer Cell Line Profiling Array determined that GBP1 expression in a majority of cell lines derived from human tumors of different tissue origin was induced to variable levels following exposure to multiple stress agents including paclitaxel and doxorubicin. Importantly, stable expression of a GBP1 transgene in the paclitaxel-sensitive ovarian cancer cell line OVCAR8 was sufficient to confer moderate paclitaxel resistance. Our data suggest that increased expression of the GBP1 gene may play an important role in the development of multi-drug resistance (MDR).