Response to Brentuximab Vedotin by CD30 Expression in Non-Hodgkin Lymphoma.

BACKGROUND: The safety and efficacy of brentuximab vedotin (BV), an antibody-drug conjugate directed to the CD30 antigen, has been assessed in several trials in patients with peripheral T-cell lymphoma (PTCL), cutaneous T-cell lymphoma (CTCL), or B-cell non-Hodgkin lymphoma (NHL). The objective of this research was to examine the relationship between CD30 expression level and clinical response to BV. PATIENTS AND METHODS: We analyzed response in patients treated with BV monotherapy in 5 prospective clinical studies in relapsed or refractory PTCL, CTCL, or B-cell NHL. CD30 expression was assessed by immunohistochemistry (IHC) using the Ber H2 antibody for 275 patients. RESULTS: Across all 5 studies, 140 (50.9%) patients had tumors with CD30 expression <10%, including 60 (21.8%) with undetectable CD30 by IHC. No significant differences were observed for any study in overall response rates between patients with CD30 expression ≥10% or <10%. Median duration of response was also similar in the CD30 ≥10% and <10% groups for all studies. CONCLUSIONS: In this analysis of studies across a range of CD30-expressing lymphomas, CD30 expression alone, as measured by standard IHC, does not predict clinical benefit from BV, making the determination of a threshold level of expression uncertain.

Tumor cell SYK expression modulates the tumor immune microenvironment composition in human cancer via TNF-α dependent signaling.

BACKGROUND: The expression of SYK in cancer cells has been associated with both tumor promoting and tumor suppressive effects. Despite being proposed as anticancer therapeutic target, the possible role of SYK in modulating local adaptive antitumor immune responses remains uncertain. Using detailed analysis of primary human tumors and in vitro models, we reveal the immunomodulatory effect of SYK protein in human solid cancer. METHODS: We spatially mapped SYK kinase in tumor cells, stromal cells and tumor-infiltrating leukocytes (TILs) in 808 primary non-small cell lung carcinomas (NSCLCs) from two cohorts and in 374 breast carcinomas (BCs) from two independent cohorts. We established the associations of localized SYK with clinicopathologic variables and outcomes. The immunomodulatory role of SYK on tumor cells was assessed using in vitro cytokine stimulation, transcriptomic analysis and selective SYK blockade using a small molecule inhibitor. Functional responses were assessed using cocultures of tumor cells with peripheral blood lymphocytes. T cell responses in baseline and post-treatment biopsies from patients with BC treated with a SYK inhibitor in a phase I clinical trial were also studied. RESULTS: Elevated tumor cell or leukocyte SYK expression was associated with high CD4+ and CD8+ TILs and better outcome in both NSCLC and BC. Tumor cell SYK was associated with oncogenic driver mutations in EGFR or KRAS in lung adenocarcinomas and with triple negative phenotype in BC. In cultured tumor cells, SYK was upregulated by TNFα and required for the TNFα-induced proinflammatory responses and T cell activation. SYK blockade after nivolumab in a phase I clinical trial including three patients with advanced triple negative BC reduced TILs and T cell proliferation. Our work establishes the proinflammatory function of tumor cell SYK in lung and breast cancer. SYK signaling in cultured tumor cells is required for T cell activation and SYK blockade limits adaptive antitumor immune responses and tumor rejection in patients with cancer. CONCLUSIONS: Together, our results establish the immunomodulatory role of SYK expression in human solid tumors. This information could be used to develop novel biomarkers and/or therapeutic strategies.

Response to brentuximab vedotin versus physician's choice by CD30 expression and large cell transformation status in patients with mycosis fungoides: An ALCANZA sub-analysis.

INTRODUCTION: Mycosis fungoides (MF), the most common type of cutaneous T-cell lymphoma, can lead to disfiguring lesions, debilitating pruritus and frequent skin infections. This study assessed response to brentuximab vedotin in patients with MF in the phase III ALCANZA study. METHODS: Baseline CD30 levels and large-cell transformation (LCT) status were centrally reviewed in patients with previously-treated CD30-positive MF using ≥2 skin biopsies obtained at screening; eligible patients required ≥1 biopsy with ≥10% CD30 expression. Patients were categorised as CD30min < 10% (≥1 biopsy with <10% CD30 expression), or CD30min ≥ 10% (all biopsies with ≥10% CD30 expression) and baseline LCT present or absent. Efficacy analyses were the proportion of patients with objective response lasting ≥4 months (ORR4) and progression-free survival (PFS). RESULTS: Clinical activity with brentuximab vedotin was observed across all CD30 expression levels in patients with ≥1 biopsy showing ≥10% CD30 expression. Superior ORR4 was observed with brentuximab vedotin versus physician's choice in patients: with CD30min < 10% (40.9% versus 9.5%), with CD30min ≥ 10% (57.1% versus 10.3%), with LCT (64.7% versus 17.6%) and without LCT (38.7% versus 6.5%). Brentuximab vedotin improved median PFS versus physician's choice in patients: with CD30min < 10% (16.7 versus 2.3 months), with CD30min ≥ 10% (15.5 versus 3.9 months), with LCT (15.5 versus 2.8 months) and without LCT (16.1 versus 3.5 months). Safety profiles were generally comparable across subgroups. CONCLUSION: These exploratory analyses demonstrated that brentuximab vedotin improved rates of ORR4 and PFS versus physician's choice in patients with CD30-positive MF and ≥1 biopsy showing ≥10% CD30 expression, regardless of LCT status. CLINICAL TRIAL REGISTRATION: Clinicaltrials.gov, NCT01578499.

Consistent expression of guanylyl cyclase-C in primary and metastatic gastrointestinal cancers.

BACKGROUND: The transmembrane receptor guanylate cyclase-C (GCC) has been found to be expressed in colorectal cancers. However, limited data are available on GCC protein expression in non-colorectal gastrointestinal tumors and few studies have reported whether GCC protein expression was consistently preserved in synchronous primary and metastatic cancer tissues. METHODS: GCC protein status was assessed by immunohistochemistry in tumor specimens from individuals (n = 627) with gastrointestinal tumors, including esophageal (n = 130), gastric (n = 276), pancreatic (n = 136), and colorectal (n = 85) primary and metastatic tumors. Tissue specimens consisted of tissue microarrays containing esophageal, gastric, pancreatic tumors, and whole-slide tissue sections from colorectal cancer patients with matching primary and metastatic tumors. RESULT: Among the evaluated esophageal, gastric, and pancreatic tumors, the frequency of GCC positivity at the protein level ranged from 59% to 68%. GCC was consistently expressed in primary and matched/synchronous metastatic lesions of colorectal cancer tissues derived from the same patients. CONCLUSION: This observational study demonstrated the protein expression of GCC across various gastrointestinal malignancies. In all cancer histotypes, GCC protein localization was observed predominantly in the cytoplasm compared to the membrane region of tumor cells. Consistent immunohistochemistry detection of GCC protein expression in primary colorectal cancers and in their matched liver metastases suggests that the expression of GCC is maintained throughout the process of tumor progression and formation of metastatic disease.

Brentuximab vedotin or physician's choice in CD30-positive cutaneous T-cell lymphoma (ALCANZA): an international, open-label, randomised, phase 3, multicentre trial.

BACKGROUND: Cutaneous T-cell lymphomas are rare, generally incurable, and associated with reduced quality of life. Present systemic therapies rarely provide reliable and durable responses. We aimed to assess efficacy and safety of brentuximab vedotin versus conventional therapy for previously treated patients with CD30-positive cutaneous T-cell lymphomas. METHODS: In this international, open-label, randomised, phase 3, multicentre trial, we enrolled adult patients with CD30-positive mycosis fungoides or primary cutaneous anaplastic large-cell lymphoma who had been previously treated. Patients were enrolled across 52 centres in 13 countries. Patients were randomly assigned (1:1) centrally by an interactive voice and web response system to receive intravenous brentuximab vedotin 1·8 mg/kg once every 3 weeks, for up to 16 3-week cycles, or physician's choice (oral methotrexate 5-50 mg once per week or oral bexarotene 300 mg/m2 once per day) for up to 48 weeks. The primary endpoint was the proportion of patients in the intention-to-treat population achieving an objective global response lasting at least 4 months per independent review facility. Safety analyses were done in all patients who received at least one dose of study drug. This trial was registered with ClinicalTrials.gov, number NCT01578499. FINDINGS: Between Aug 13, 2012, and July 31, 2015, 131 patients were enrolled and randomly assigned to a group (66 to brentuximab vedotin and 65 to physician's choice), with 128 analysed in the intention-to-treat population (64 in each group). At a median follow-up of 22·9 months (95% CI 18·4-26·1), the proportion of patients achieving an objective global response lasting at least 4 months was 56·3% (36 of 64 patients) with brentuximab vedotin versus 12·5% (eight of 64) with physician's choice, resulting in a between-group difference of 43·8% (95% CI 29·1-58·4; p<0·0001). Grade 3-4 adverse events were reported in 27 (41%) of 66 patients in the brentuximab vedotin group and 29 (47%) of 62 patients in the physician's choice group. Peripheral neuropathy was seen in 44 (67%) of 66 patients in the brentuximab vedotin group (n=21 grade 2, n=6 grade 3) and four (6%) of 62 patients in the physician's choice group. One of the four on-treatment deaths was deemed by the investigator to be treatment-related in the brentuximab vedotin group; no on-treatment deaths were reported in the physician's choice group. INTERPRETATION: Significant improvement in objective response lasting at least 4 months was seen with brentuximab vedotin versus physician's choice of methotrexate or bexarotene. FUNDING: Millennium Pharmaceuticals Inc (a wholly owned subsidiary of Takeda Pharmaceutical Company Ltd), Seattle Genetics Inc.

Use of the Total Cancer Care System to Enrich Screening for CD30-Positive Solid Tumors for Patient Enrollment Into a Brentuximab Vedotin Clinical Trial: A Pilot Study to Evaluate Feasibility.

BACKGROUND: One approach to identify patients who meet specific eligibility criteria for target-based clinical trials is to use patient and tumor registries to prescreen patient populations. OBJECTIVE: Here we demonstrate that the Total Cancer Care (TCC) Protocol, an ongoing, observational study, may provide a solution for rapidly identifying patients with CD30-positive tumors eligible for CD30-targeted therapies such as brentuximab vedotin. METHODS: The TCC patient gene expression profiling database was retrospectively screened for CD30 gene expression determined using HuRSTA-2a520709 Affymetrix arrays (GPL15048). Banked tumor tissue samples were used to determine CD30 protein expression by semiquantitative immunohistochemistry. Statistical comparisons of Z- and H-scores were performed using R statistical software (The R Foundation), and the predictive value, accuracy, sensitivity, and specificity of CD30 gene expression versus protein expression was estimated. RESULTS: As of March 2015, 120,887 patients have consented to the institutional review board-approved TCC Protocol. A total of 39,157 fresh frozen tumor specimens have been collected, from which over 14,000 samples have gene expression data available. CD30 RNA was expressed in a number of solid tumors; the highest median CD30 RNA expression was observed in primary tumors from lymph node, soft tissue (many sarcomas), lung, skin, and esophagus (median Z-scores 1.011, 0.399, 0.202, 0.152, and 1.011, respectively). High level CD30 gene expression significantly enriches for CD30-positive protein expression in breast, lung, skin, and ovarian cancer; accuracy ranged from 72% to 79%, sensitivity from 75% to 100%, specificity from 70% to 76%, positive predictive value from 20% to 40%, and negative predictive value from 95% to 100%. CONCLUSIONS: The TCC gene expression profiling database guided tissue selection that enriched for CD30 protein expression in a number of solid tumor types. Such an approach may improve screening efficiency for enrolling patients into biomarker-based clinical trials.

Understanding the Supersensitive Anti-Drug Antibody Assay: Unexpected High Anti-Drug Antibody Incidence and Its Clinical Relevance.

Numbers of biotherapeutic products in development have increased over past decade. Despite providing significant benefits to patients with unmet needs, almost all protein-based biotherapeutics could induce unwanted immunogenicity, which result in a loss of efficacy and/or increase the risk of adverse reactions, such as infusion reactions, anaphylaxis, and even life-threatening response to endogenous proteins. Recognizing these possibilities, regulatory agencies request that immunogenicity be assessed as part of the approval process for biotherapeutics. Great efforts have been made to reduce drug immunogenicity through protein engineering. Accordingly the immunogenicity incidence has been reduced from around 80% in murine derived products to 0-10% in fully human products. However, recent improvements in immunogenicity assays have led to unexpectedly high immunogenicity rates, even in fully human products, leading to new challenges in assessing immunogenicity and its clinical relevance. These new immunogenicity assays are becoming supersensitive and able to detect more of anti-drug antibodies (ADA) than with earlier assays. This paper intends to review and discuss our understanding of the supersensitive ADA assay and the unexpected high ADA incidence and its potential clinical relevance.

Development of a Drug-Response Modeling Framework to Identify Cell Line Derived Translational Biomarkers That Can Predict Treatment Outcome to Erlotinib or Sorafenib.

Development of drug responsive biomarkers from pre-clinical data is a critical step in drug discovery, as it enables patient stratification in clinical trial design. Such translational biomarkers can be validated in early clinical trial phases and utilized as a patient inclusion parameter in later stage trials. Here we present a study on building accurate and selective drug sensitivity models for Erlotinib or Sorafenib from pre-clinical in vitro data, followed by validation of individual models on corresponding treatment arms from patient data generated in the BATTLE clinical trial. A Partial Least Squares Regression (PLSR) based modeling framework was designed and implemented, using a special splitting strategy and canonical pathways to capture robust information for model building. Erlotinib and Sorafenib predictive models could be used to identify a sub-group of patients that respond better to the corresponding treatment, and these models are specific to the corresponding drugs. The model derived signature genes reflect each drug's known mechanism of action. Also, the models predict each drug's potential cancer indications consistent with clinical trial results from a selection of globally normalized GEO expression datasets.

Mutation of NRAS but not KRAS significantly reduces myeloma sensitivity to single-agent bortezomib therapy.

Various translocations and mutations have been identified in myeloma, and certain aberrations, such as t(4;14) and del17, are linked with disease prognosis. To investigate mutational prevalence in myeloma and associations between mutations and patient outcomes, we tested a panel of 41 known oncogenes and tumor suppressor genes in tumor samples from 133 relapsed myeloma patients participating in phase 2 or 3 clinical trials of bortezomib. DNA mutations were identified in 14 genes. BRAF as well as RAS genes were mutated in a large proportion of cases (45.9%) and these mutations were mutually exclusive. New recurrent mutations were also identified, including in the PDGFRA and JAK3 genes. NRAS mutations were associated with a significantly lower response rate to single-agent bortezomib (7% vs 53% in patients with mutant vs wild-type NRAS, P = .00116, Bonferroni-corrected P = .016), as well as shorter time to progression in bortezomib-treated patients (P = .0058, Bonferroni-corrected P = .012). However, NRAS mutation did not impact outcome in patients treated with high-dose dexamethasone. KRAS mutation did not reduce sensitivity to bortezomib or dexamethasone. These findings identify a significant clinical impact of NRAS mutation in myeloma and demonstrate a clear example of functional differences between the KRAS and NRAS oncogenes.

Sequence analysis of ß-subunit genes of the 20S proteasome in patients with relapsed multiple myeloma treated with bortezomib or dexamethasone.

Variations within proteasome ß (PSMB) genes, which encode the ß subunits of the 20S proteasome, may affect proteasome function, assembly, and/or binding of proteasome inhibitors. To investigate the potential association between PSMB gene variants and treatment-emergent resistance to bortezomib and/or long-term outcomes, in the present study, PSMB gene sequence variation was characterized in tumor DNA samples from patients who participated in the phase 3 Assessment of Proteasome Inhibition for Extending Remissions (APEX) study of bortezomib versus high-dose dexamethasone for treatment of relapsed multiple myeloma. Twelve new PSMB variants were identified. No associations were found between PSMB single nucleotide polymorphism genotype frequency and clinical response to bortezomib or dexamethasone treatment or between PSMB single nucleotide polymorphism allelic frequency and pooled overall survival or time to progression. Although specific PSMB5 variants have been identified previously in preclinical models of bortezomib resistance, these variants were not detected in patient tumor samples collected after clinical relapse from bortezomib, which suggests that alternative mechanisms underlie bortezomib insensitivity.

Validation of a flow cytometry based G(2)M delay cell cycle assay for use in evaluating the pharmacodynamic response to Aurora A inhibition.

Pharmacodynamic assays are important aspects for understanding molecularly targeted anticancer agents to investigate the relationship between drug concentration (pharmacokinetics) and drug "effect" or biological activity. As new drug entities are developed that affect DNA cell cycle, a pharmacodynamic assay which measures cell cycle perturbation would be a valuable clinical trial tool. During recent years, flow cytometry has established itself as a useful method to determine the relative nuclear DNA content and percentage of cycling cells of biological specimens. However to date, the analytical validation of cytometry based assays is limited and there is no suitable guidance for method validation of flow cytometry based cell cycle assays. Here we report the validation of a flow cytometry based cell cycle G(2)/M delay assay for use in evaluating the effect of investigational drug MLN8237, a small molecule inhibitor of a mitotic kinase Aurora A, for clinical trial use. The assay method was validated by examining assay robustness, repeatability, reproducibility, precision, and determining the cutoff for a true drug effect based on biostatistical analysis models. Experimental results show that the intra-assay repeatability was less than 20% with an intra-donor variability of less than 40%. The robustness of the assay was less than 30%. Since this is an ex-vivo stimulation assay, variability parameters were expected to be higher. Based on biostatistical modeling, an absolute change in %G(2)M of 5.2% (95% CI) was needed in order to detect a true drug effect. Overall, the assay demonstrated acceptable variability to warrant further in vivo testing.

Potential biomarkers of bortezomib activity in mantle cell lymphoma from the phase 2 PINNACLE trial.

Immunohistochemical analyses of archival tumor specimens were used for pre-planned exploratory analyses of protocol-specified candidate biomarkers of bortezomib activity in 73 patients with relapsed/refractory mantle cell lymphoma in the phase 2 PINNACLE study. Consistent with other studies, elevated Ki-67 was a marker of poor prognosis, demonstrating significant associations with shorter time to progression and overall survival. Elevated NF-kappaB p65 and low PSMA5 expression demonstrated a trend for better response and were significantly associated with longer time to progression; elevated NF-kappaB p65 demonstrated a trend toward longer overall survival. This is consistent with myeloma clinical genomics research, suggesting biomarker relevance across tumor types. Elevated p27 was significantly associated with longer overall survival. Overall survival analyses by International Prognostic Index and Mantle Cell Lymphoma International Prognostic Index confirmed differential prognosis by both scores. These biomarkers data begin to illuminate bortezomib's mechanism of action in lymphoma.

A randomized phase 2 study of erlotinib alone and in combination with bortezomib in previously treated advanced non-small cell lung cancer.

INTRODUCTION: This phase 2 study was conducted to determine the efficacy and safety of erlotinib alone and with bortezomib in patients with non-small cell lung cancer (NSCLC). METHODS: Patients with histologically or cytologically confirmed relapsed or refractory stage IIIb/IV NSCLC were randomized (1:1; stratified by baseline histology, smoking history, sex) to receive erlotinib 150 mg/d alone (arm A; n = 25) or in combination with bortezomib 1.6 mg/m2, days 1 and 8 (arm B; n = 25) in 21-day cycles. Responses were assessed using Response Evaluation Criteria in Solid Tumors. Tumor samples were evaluated for mutations predicting response. Six additional patients received the combination in a prior dose deescalation stage and were included in safety analyses. RESULTS: Response rates were 16% in arm A and 9% in arm B; disease control rates were 52 and 45%, respectively. The study was halted at the planned interim analysis due to insufficient clinical activity in arm B. Median progression-free survival and overall survival were 2.7 and 7.3 months in arm A, and 1.3 and 8.5 months in arm B. Six-month survival rates were 56.0% in both arms; 12-month rates were 40 and 30% in arms A and B, respectively. Response rate to erlotinib+/-bortezomib was significantly higher in patients with epidermal growth factor receptor mutations (50 versus 9% for wild type). The most common treatment-related grade > or =3 adverse event was skin rash (three patients in each treatment group). CONCLUSION: Insufficient activity was seen with erlotinib plus bortezomib in patients with relapsed/refractory advanced NSCLC to warrant a phase 3 study of the combination.

Effect of the CYP3A inhibitor ketoconazole on the pharmacokinetics and pharmacodynamics of bortezomib in patients with advanced solid tumors: a prospective, multicenter, open-label, randomized, two-way crossover drug-drug interaction study.

BACKGROUND: The proteasome inhibitor bortezomib undergoes oxidative biotransformation via multiple cytochrome P450 (CYP) enzymes, with CYP3A4 identified as a partial, yet potentially important, contributor based on in vitro drug metabolism studies. OBJECTIVE: The aim of this study was to assess the effect of concomitant administration of ketoconazole on the pharmacokinetics (PK) and pharmacodynamics (PD) of bortezomib. METHODS: This was a prospective, multicenter, open-label, randomized, multiple-dose, 2-way crossover study in patients with advanced solid tumors. All patients received bortezomib 1.0 mg/m(2) IV (on days 1, 4, 8, and 11 of two 21-day cycles) and were randomized to receive concomitant ketoconazole 400 mg on days 6, 7, 8, and 9 of cycle 1 or 2. Serial blood samples were collected over the day-8 dosing interval (immediately prior to bortezomib administration, and from 5 minutes to 72 hours after administration) in cycles 1 and 2 for measurement of plasma bortezomib concentrations for noncompartmental PK analysis and blood 20S proteasome inhibition for PD analysis. All adverse events (AEs) were recorded during each cycle including serious AEs and all neurotoxicity events for up to 30 days after the last dose of bortezomib. RESULTS: Twenty-one patients (median age, 57 years; sex, 67% male; race, 86% white; median body surface area, 2.01 m(2)) were randomized to treatment. Twelve patients completed the protocol-specified dosing and PK sampling in both cycles 1 and 2. Assessment of the effect of ketoconazole on bortezomib PK and PD was based on data in these 12 PK-evaluable patients. The ratio of geometric mean bortezomib AUC(0-tlast)(AUC from time 0 to last quantifiable concentration) for bortezomib plus ketoconazole versus bortezomib alone was 1.352 (90% CI, 1.032-1.772). Consistent with this observed mean increase in bortezomib exposure, concomitant administration of ketoconazole was associated with a corresponding increase (24%-46%) in the blood proteasome inhibitory effect. CONCLUSION: Concomitant administration of the CYP3A inhibitor ketoconazole with bortezomib resulted in a mean increase of 35% in bortezomib exposure. ClinicalTrials.gov identifier: NCT00129207.

Molecular profile of peripheral blood mononuclear cells from patients with rheumatoid arthritis.

Rheumatoid arthritis (RA) is a chronic inflammatory arthritis. Currently, diagnosis of RA may take several weeks, and factors used to predict a poor prognosis are not always reliable. Gene expression in RA may consist of a unique signature. Gene expression analysis has been applied to synovial tissue to define molecularly distinct forms of RA; however, expression analysis of tissue taken from a synovial joint is invasive and clinically impractical. Recent studies have demonstrated that unique gene expression changes can be identified in peripheral blood mononuclear cells (PBMCs) from patients with cancer, multiple sclerosis, and lupus. To identify RA disease-related genes, we performed a global gene expression analysis. RNA from PBMCs of 9 RA patients and 13 normal volunteers was analyzed on an oligonucleotide array. Compared with normal PBMCs, 330 transcripts were differentially expressed in RA. The differentially regulated genes belong to diverse functional classes and include genes involved in calcium binding, chaperones, cytokines, transcription, translation, signal transduction, extracellular matrix, integral to plasma membrane, integral to intracellular membrane, mitochondrial, ribosomal, structural, enzymes, and proteases. A k-nearest neighbor analysis identified 29 transcripts that were preferentially expressed in RA. Ten genes with increased expression in RA PBMCs compared with controls mapped to a RA susceptibility locus, 6p21.3. These results suggest that analysis of RA PBMCs at the molecular level may provide a set of candidate genes that could yield an easily accessible gene signature to aid in early diagnosis and treatment.

Gene expression profiling and correlation with outcome in clinical trials of the proteasome inhibitor bortezomib.

The aims of this study were to assess the feasibility of prospective pharmacogenomics research in multicenter international clinical trials of bortezomib in multiple myeloma and to develop predictive classifiers of response and survival with bortezomib. Patients with relapsed myeloma enrolled in phase 2 and phase 3 clinical trials of bortezomib and consented to genomic analyses of pretreatment tumor samples. Bone marrow aspirates were subject to a negative-selection procedure to enrich for tumor cells, and these samples were used for gene expression profiling using DNA microarrays. Data quality and correlations with trial outcomes were assessed by multiple groups. Gene expression in this dataset was consistent with data published from a single-center study of newly diagnosed multiple myeloma. Response and survival classifiers were developed and shown to be significantly associated with outcome via testing on independent data. The survival classifier improved on the risk stratification provided by the International Staging System. Predictive models and biologic correlates of response show some specificity for bortezomib rather than dexamethasone. Informative gene expression data and genomic classifiers that predict clinical outcome can be derived from prospective clinical trials of new anticancer agents.

Exploiting genotypic differences to identify genes important for EAE development.

Experimental autoimmune encephalomyelitis (EAE) is an animal model of the human autoimmune disease multiple sclerosis (MS) and is primarily driven by T helper type 1 (Th1) cells. Interleukin (IL)-12 and interferon (IFN)-gamma are important cytokines involved in the differentiation and amplification of Th1 cells, however mice deficient in either IFN-gamma or IL-12 still develop EAE. We have used microarray analysis of EAE-affected CNS tissues in wild-type, IFN-gamma -/- and IL-12 -/- animals to identify genes critical for development of EAE. Over 500 genes were regulated in at least one genotype and over 94 genes were regulated in all three. Of those, 17 were also upregulated in spleen during the disease. We show that a majority of the genes regulated in EAE are also regulated in diseased regions of human MS tissues. The genes in the pool of 94 are more likely to be found regulated in MS patients than the genes regulated in only one or two of the mouse strains suggesting that analyzing gene expression under these multiple genetic conditions may lead to better identification of the genes critical for disease development.

A single administration of recombinant human interleukin-12 is associated with increased expression levels of interferon-gamma and signal transducer and activator of transcription in healthy subjects.

The objectives of this study were to assess the safety and tolerability of single doses of 1, 4, and 8 mug of recombinant human interleukin-12 (rhIL-12) administered subcutaneously to healthy subjects. The pharmacokinetics, pharmacodynamics, and pharmacogenomics of rhIL-12 were evaluated. Recombinant human IL-12 was well tolerated in these healthy male and female subjects. The most frequently reported adverse events were flu-like symptoms, which exhibited a dose-response relationship. Pharmacokinetic analysis suggested that serum IL-12 levels increased with dose. Analysis of serum levels indicated that interferon-gamma increased with the dose of rhIL-12, whereas IL-6 levels showed no changes with rhIL-12 treatment. The messenger ribonucleic acid expression of signal transducer and activator of transcription was significantly increased 24 hours after the administration of rhIL-12 for all dose groups versus placebo, and results indicated that the magnitude of increase may be dose dependent. This study suggests that interferon-gamma and signal transducer and activator of transcription are biomarkers of rhIL-12 activity.

Transcriptional profiles in peripheral blood mononuclear cells prognostic of clinical outcomes in patients with advanced renal cell carcinoma.

PURPOSE: Given their accessibility, surrogate tissues, such as peripheral blood mononuclear cells (PBMC), may provide potential predictive biomarkers in clinical pharmacogenomic studies. In leukemias and lymphomas, the prognostic value of peripheral blast expression profiles is clear; however, it is unclear whether circulating mononuclear cells of patients with solid tumors might yield profiles with similar prognostic associations. EXPERIMENTAL DESIGN: In this study, we evaluated the association of expression profiles in PBMCs with clinical outcomes in patients with advanced renal cell cancer. Transcriptional patterns in PBMCs of 45 renal cell cancer patients were compared with clinical outcome data at the conclusion of a phase II study of the mTOR kinase inhibitor CCI-779 to determine whether pretreatment transcriptional patterns in PBMCs were correlated with eventual patient outcomes. RESULTS: Unsupervised hierarchical clustering of the PBMC profiles using all expressed genes identified clusters of patients with significant differences in survival. Cox proportional hazards modeling showed that the expression levels of many PBMC transcripts were predictors for the patient outcomes of time to progression and overall survival (time to death). Supervised class prediction approaches identified multivariate expression patterns in PBMCs capable of assigning favorable outcomes of time to death and time to progression in a test set of renal cancer patients, with overall performance accuracies of 72% and 85%, respectively. CONCLUSIONS: The present study provides the first example of gene expression profiling in peripheral blood, a clinically accessible surrogate tissue, for identifying patterns of gene expression associated with higher likelihoods of positive outcome in patients with a solid tumor.

Clinical pharmacogenomics and transcriptional profiling in early phase oncology clinical trials.

Microarray-based expression profiling studies in the field of oncology have demonstrated encouraging correlations between tumor transcriptional profiles and eventual patient outcomes. These findings have fueled great interest in the application of transcriptional profiling to samples available from real-time clinical trials, and clinical pharmacogenomic objectives utilizing transcriptional profiling strategies are becoming increasingly incorporated into clinical trial study designs. Over the last few years several retrospective studies based on the profiling of archival tumor tissues suggest that transcriptional analysis of oncology samples may provide general prognosis measures, and in some cases may even predict response to specific therapies. Recently the FDA released a voluntary genomic data guidance meant to assist both regulatory agencies and pharmaceutical companies alike in evaluating the potential benefit of implementing expression profiling studies during the preclinical and clinical phases of drug development. Despite the great promise afforded by this technology, the ultimate benefit of applying transcriptional profiling in prospective clinical trials has yet to be realized because a number of practical impediments to this process exist. The multi-fold purpose of the current review is to highlight the increasing evidence from studies that have identified transcriptional signatures in archived tumors prognostic of patient outcome, to describe some of the drivers for the implementation of transcriptional profiling strategies in real-time drug development, to discuss the use of transcriptional profiling in the context of increasingly complex translational medicine strategies, and to highlight the practical issues and potential approaches involved in the successful application of clinical pharmacogenomic objectives during real-time clinical trials. Strategic implementation of transcriptional profiling in early oncology clinical trials can provide an opportunity to identify predictive markers of clinical response and eventually provide a substantial step forward towards the era of personalized medicine.