Citations

Abstract A group of investigators met at a Specialized Programs of Research Excellence Workshop to discuss key issues in the translation of biomarker discovery to the development of useful laboratory tests for cancer care. Development and approval of several new markers and technologies have provided informative examples that include more specific markers for prostate cancer, more sensitive tests for ovarian cancer, more objective analysis of tissue architecture and an earlier indication of response to treatment in breast cancer. Although there is no clear paradigm for biomarker development, several principles are clear. Marker development should be driven by clinical needs, including early cancer detection, accurate pretreatment staging, and prediction of response to treatment, as well as monitoring disease progression and response to therapy. Development of a national repository that uses carefully preserved, well-annotated tissue specimens will facilitate new marker development. Reference standards will be an essential component of this process. Both hospital-based and commercial laboratories can play a role in developing biomarkers from discovery to test validation. Partnering of academe and industry should occur throughout the process of biomarker development. The National Cancer Institute is in a unique position to bring together academe, industry, and the Food and Drug Administration to (a) define clinical needs for biomarkers by tumor type, (b) establish analytic and clinical paradigms for biomarker development, (c) discuss ways in which markers from different companies might be evaluated in combination, (d) establish computational methods to combine data from multiple biomarkers, (e) share information regarding promising markers developed in National Cancer Institute–supported programs, and (f) exchange data regarding new platforms and techniques that can accelerate marker development.

AbstractPurpose: The B7 family molecules have been shown to regulate immune responses in both positive and negative fashions. Their roles in the progression of human cancers, however, are not well established. The aim of this study was to examine whether leukemic cells of acute myeloid leukemia express functional B7 family molecules and, if so, whether such expression has any clinical significance.Experimental Design: The expression of four B7 family molecules, B7.1, B7.2, B7-H1, and B7-H2, on leukemic cells from acute myeloid leukemia patients was analyzed by flow cytometry. The function of the expressed molecules was examined by the allogeneic mixed lymphocyte-leukemic cell reaction, and their relationship to the clinical data and survival was analyzed.Results: Although B7.1 and B7-H1 expressions were rare, the cells from a substantial number of acute myeloid leukemia cases expressed B7.2 and B7-H2 molecules [mean percentages of B7.2- and B7-H2-positive cells were 28.9% (n = 58) and 15.3% (n = 59), respectively]. Patients in whom >25% of leukemic cells expressed B7-H2 had significantly shorter survival, and this B7-H2 positivity had the strongest prognostic value when B7-H2 and other prognostic factors were analyzed together by multivariate analysis (P = 0.0108). Furthermore, B7.2 expression was associated with hyperleukocytosis (P = 0.026). Consistent with this finding, acute myeloid leukemia cells expressing B7.2 and B7-H2 induced allogeneic CD4+ T cells to proliferate and secrete interleukin-4 and interleukin-10 in vitro, effects that were partially blocked by antibodies against B7.2 and B7-H2.Conclusions: Our results indicate the expression of functional B7.2 and B7-H2 molecules, and these molecules may facilitate progression of acute myeloid leukemia.

Abstract Purpose: Flavopiridol, a cyclin-dependent kinase inhibitor, transcription inhibitor, and DNA-interacting agent, was combined with cisplatin or carboplatin to establish toxicities, evaluate pharmacokinetics, and examine its effects on patient cancers and levels of selected polypeptides in patient peripheral blood mononuclear cells (PBMC). Experimental Design: Therapy was given every 3 weeks. Stage I: cisplatin was fixed at 30 mg/m2 with escalating flavopiridol. Stage II: flavopiridol was fixed at the stage I maximum tolerated dose (MTD) with escalation of cisplatin. Stage III: flavopiridol was fixed at the stage I MTD with escalation of carboplatin. Results: Thirty-nine patients were treated with 136 cycles of chemotherapy. Neutropenia was seen in only 11% of patients. Grade 3 flavopiridol/CDDP toxicities were nausea (30%), vomiting (19%), diarrhea (15%), dehydration (15%), and neutropenia (10%). Flavopiridol combined with carboplatin resulted in unexpectedly high toxicities and one treatment-related death. Stable disease (>3 months) was seen in 34% of treated patients, but there were no objective responses. The stage II MTD was 60 mg/m2 cisplatin and 100 mg/m2/24 hours flavopiridol. As given, CDDP did not alter flavopiridol pharmacokinetics. Flavopiridol induced increased p53 and pSTAT3 levels in patient PBMCs but had no effects on cyclin D1, phosphoRNA polymerase II, or Mcl-1. Conclusions: Flavopiridol and cisplatin can be safely combined in the treatment of cancer patients. Unexpected toxicity in flavopiridol/carboplatin-treated patients attenuates enthusiasm for this alternative combination. Analysis of polypeptide levels in patient PBMCs suggests that flavopiridol may be affecting some, but not all, of its known in vitro molecular targets in vivo.

Abstract Purpose: The objective of this study was to use gene expression profiling to identify novel biomarkers that are predictive of aggressive behavior in clear cell renal cell carcinoma (CCRCC). Experimental Design: Candidate genes were discovered using Human Genome U133 Plus 2 Arrays and validated on independent samples by quantitative reverse transcription-PCR (RT-PCR). Both the discovery and the validation cohorts included nonaggressive primary CCRCC, aggressive primary CCRCC, metastatic CCRCC, and nonneoplastic kidney adjacent to tumor. Results: Aggressive primary and metastatic CCRCC displayed no significant differences in gene expression. In contrast, we identified significant differences in gene expression between nonaggressive and aggressive CCRCC (including metastatic CCRCC). Thirty-four of the 35 transcripts that displayed the most significant differential expression by microarray analysis also displayed significant differential expression in independent validation studies using quantitative RT-PCR (P < 0.001 for 31 candidates and P < 0.005 for the remaining three candidates). Hierarchical clustering of the quantitative RT-PCR data using our candidate markers accurately grouped 88% (23 of 26) of aggressive and metastatic CCRCC samples, 100% (14 of 14) of nonaggressive CCRCC samples, and 100% (15 of 15) of nonneoplastic samples into separate clusters. Finally, we evaluated the ability of protein expression levels of one of our candidate markers (survivin) to predict survival among a cohort of 183 CCRCC patients treated surgically at Mayo Clinic from 1990 to 1992. In multivariate analysis, expression of survivin (BIRC5) was inversely associated with cancer-specific survival (P = 0.017). Conclusion: We used a combination of genomic profiling and validation by quantitative PCR to identify a panel of candidate biomarkers for determining CCRCC aggressiveness. Our data also indicate that the gene expression alterations that result in aggressive behavior and metastatic potential can be identified in the primary tumor.

Abstract Past lung cancer screening trials in the United States with chest X-ray and sputum cytology were not able to show any decrease in lung cancer mortality; however, these trials are over 20 years old. Recent follow-up of the Mayo Lung Project showed a better survival from lung cancer in the screened arm, but no difference in overall mortality, suggesting an overdiagnosis of nonfatal cancers. Recent reports of low radiation dose spiral computed tomography (CT) chest screening for lung cancer have shown that CT screening detects cancers at a smaller size than chest X-rays. To date, there have been no randomized trials of CT versus observation or chest radiographs for screening purposes. All data available thus far on CT screening are from phase II proof-of-principle trials. The major limitations of CT screening, discussed here, include (a) a high rate of nodule detection: over 50% of participants will have at least one noncalcified nodule; (b) resulting follow-up CT scans, associated with increased costs; (c) cost and morbidity of biopsy or resection of benign noncalcified nodule (20-25% of such procedures in several trials); and (d) a small, but difficult to quantify, risk of cancer associated with multiple follow-up CT scans.

AbstractThe identification of novel markers and therapeutic targets in advanced cancer is critical for improving diagnosis and therapy. Six-transmembrane epithelial antigen of the prostate (STEAP) is expressed predominantly in human prostate tissue and in other common malignancies including prostate, bladder, colon, and ovarian carcinomas, and in Ewing's sarcoma, suggesting that it could function as an almost universal tumor antigen. We have used MHC peptide binding algorithms to predict potential STEAP sequences capable of stimulating in vitro naïve HLA-A2–restricted CTLs. Four of six peptides predicted by these algorithms were able to induce antigen-specific CTLs that killed peptide-pulsed HLA-A2 target cells. Two of these peptides, STEAP-292 (MIAVFLPIV) and a modification of this peptide STEAP-292.2L (MLAVFLPIV), were the most efficient in the induction of primary CTL responses. More importantly, these CTLs were able to respond to tumor cells that express HLA-A2 and STEAP (colon, bladder, prostate, Ewing's sarcoma, and melanoma). Our results provide strong evidence that STEAP-292 is naturally processed by many tumor types and is presented in the context of HLA-A2 in sufficient amounts to allow recognition by CTLs. Also because STEAP-292.2L is a more immunogenic peptide able to induce CTL recognition of these STEAP-containing tumors and may have potential as an antitumor peptide vaccine.

Abstract Purpose: We examined the feasibility of using molecular characterization of circulating tumor cells as a method for early detection of breast cancer. Research Design: Women without a prior history of cancer who had a breast abnormality detected on imaging followed by a breast biopsy were enrolled in this study. Density gradient centrifugation and immunomagnetic capture were used to enrich for epithelial cells from ∼20 mL of blood. Real-time reverse transcription-PCR was used to quantitate the expression levels of the highly breast-specific genes, mammaglobin, γ-aminobutyric acid type A receptor π subunit (GABA Aπ), B305D-C, and B726P in the epithelial cell–enriched samples. Results: The assay was technically feasible in 154 of 199 accrued patients. From their clinical assessment, 100 patients had benign breast disease, 10 patients had ductal carcinoma in situ, and 44 patients had invasive breast cancer. We constructed a diagnostic test that classified patients with mammaglobin levels of at least 32.2 copies/pg β-actin (units) in their circulating epithelial cells as positive for invasive breast cancer. This resulted in a sensitivity and specificity of 63.3% and 75.0%, respectively. A diagnostic test that classified patients as positive for invasive breast cancer when either mammaglobin levels were >46.3 units or B305D-C levels were >11.6 units increased the sensitivity and specificity to 70.5% and 81.0%, respectively. In the latter test, 12 of the 14 node-positive breast cancer patients were correctly identified. Including GABA Aπ and B726P in the test did not increase its diagnostic potential. Conclusions: These results suggest that molecular characterization of circulating epithelial cells using mammaglobin and B305D-C offers potential for early detection of invasive breast cancer.

Abstract Purpose: To define the toxicities, pharmacodynamics, and clinical activity of the proteasome inhibitor, PS-341 (bortezomib), in patients with advanced malignancies. Patients and Methods: Twenty-eight patients (14 male and 14 female) received PS-341 twice weekly for 4 of 6 weeks (schedule I). Because toxicity necessitated dose omissions on this schedule, 16 additional patients (12 male and 4 female) received PS-341 twice weekly for 2 of every 3 weeks (schedule II). A total of 73 courses of treatment was given (median, 2; range, 1-4). Inhibition of 20S proteasome activity in peripheral blood mononuclear cells (PBMC) and accumulation of proteasome-targeted polypeptides in tumor tissue were evaluated as pharmacodynamic markers of PS-341 activity. Results: The most common toxicity was thrombocytopenia, which was dose limiting at 1.7 mg/m2 (schedule I) and 1.6 mg/m2 (schedule II), respectively. Sensory neuropathy was dose-limiting in a patient in schedule I. Grade ≥3 toxicities for schedule I were constipation, fatigue, myalgia, and sensory neuropathy. Grade ≥3 toxicities for schedule II were dehydration resulting from diarrhea, nausea and vomiting, fatigue, hypoglycemia, and hypotension. The maximum tolerated dose was 1.5 mg/m2 for both schedules. Reversible dose-dependent decreases in 20S proteasome activity in PBMCs were observed, with 36% inhibition at 0.5 mg/m2, 52% at 0.9 mg/m2, and 75% at 1.25 mg/m2. Accumulation of proteasome-targeted polypeptides was detected in tumor samples after treatment with PS-341. A patient with multiple myeloma had a partial response. Conclusion: PS-341 given 1.5 mg/m2 twice weekly for 2 of every 3 weeks is well tolerated and should be further studied.

Abstract Purpose: YKL-40 is a secreted protein that has been reported to be overexpressed in epithelial cancers and gliomas, although its function is unknown. Previous data in a smaller sample set suggested that YKL-40 was a marker associated with a poorer clinical outcome and a genetically defined subgroup of glioblastoma. Here we test these findings in a larger series of patients with glioblastoma, and in particular, determine if tumor YKL-40 expression is associated with radiation response. Experimental Design: Patients (n = 147) with subtotal resections were studied for imaging-assessed changes in tumor size in serial studies following radiation therapy. An additional set (n = 140) of glioblastoma patients who underwent a gross-total resection was tested to validate the survival association and extend them to patients with minimal residual disease. Results: In the subtotal resection group, higher YKL-40 expression was significantly associated with poorer radiation response, shorter time to progression and shorter overall survival. The association of higher YKL-40 expression with poorer survival was validated in the gross-total resection group. In multivariate analysis with both groups combined (n = 287), YKL-40 was an independent predictor of survival after adjusting for patient age, performance status, and extent of resection. YKL-40 expression was also compared with genetically defined subsets of glioblastoma by assessing epidermal growth factor receptor amplification and loss at chromosome 10q, two of the common recurring aberrations in these tumors, using fluorescent in situ hybridization. YKL-40 was significantly associated with 10q loss. Conclusions: The findings implicate YKL-40 as an important marker of therapeutic response and genetic subtype in glioblastomas and suggest that it may play an oncogenic role in these tumors.