Single Cell Profiling of Circulating Tumor Cells: Transcriptional Heterogeneity and Diversity from Breast Cancer Cell Lines

BACKGROUND: To improve cancer therapy, it is critical to target metastasizing cells. Circulating tumor cells (CTCs) are rare cells found in the blood of patients with solid tumors and may play a key role in cancer dissemination. Uncovering CTC phenotypes offers a potential avenue to inform treatment. However, CTC transcriptional profiling is limited by leukocyte contamination; an approach to surmount this problem is single cell analysis. Here we demonstrate feasibility of performing high dimensional single CTC profiling, providing early insight into CTC heterogeneity and allowing comparisons to breast cancer cell lines widely used for drug discovery. METHODOLOGY/PRINCIPAL FINDINGS: We purified CTCs using the MagSweeper, an immunomagnetic enrichment device that isolates live tumor cells from unfractionated blood. CTCs that met stringent criteria for further analysis were obtained from 70% (14/20) of primary and 70% (21/30) of metastatic breast cancer patients; none were captured from patients with non-epithelial cancer (n = 20) or healthy subjects (n = 25). Microfluidic-based single cell transcriptional profiling of 87 cancer-associated and reference genes showed heterogeneity among individual CTCs, separating them into two major subgroups, based on 31 highly expressed genes. In contrast, single cells from seven breast cancer cell lines were tightly clustered together by sample ID and ER status. CTC profiles were distinct from those of cancer cell lines, questioning the suitability of such lines for drug discovery efforts for late stage cancer therapy. CONCLUSIONS/SIGNIFICANCE: For the first time, we directly measured high dimensional gene expression in individual CTCs without the common practice of pooling such cells. Elevated transcript levels of genes associated with metastasis NPTN, S100A4, S100A9, and with epithelial mesenchymal transition: VIM, TGFß1, ZEB2, FOXC1, CXCR4, were striking compared to cell lines. Our findings demonstrate that profiling CTCs on a cell-by-cell basis is possible and may facilitate the application of 'liquid biopsies' to better model drug discovery

The outcome of peripheral t-cell lymphoma patients failing first-line therapy: A report from the prospective, international t-cell project

This analysis explored factors influencing survival of patients with primary refractory and relapsed peripheral T-cell lymphomas enrolled in the prospective International T-cell Project. We analyzed data from 1020 patients with newly diagnosed disease, enrolled between September 2006 and December 2015. Out of 937 patients who received first-line treatment, 436 (47%) were identified as refractory and 197 (21%) as relapsed. Median time from the end of treatment to relapse was 8 months (range 2-73). Overall, 75 patients (8%) were consolidated with bone marrow transplantation, including 12 refractory and 22 relapsed patients. After a median follow up of 38 months (range 1-96 months) from documentation of refractory/relapsed disease, 440 patients had died. The median overall survival (OS) was 5.8 months; 3-year overall survival rates were 21% and 28% for refractory and relapsed patients, respectively (P12 months, HR 0.57, P=0.001) and salvage therapy with transplantation (HR=0.36, P<0.001) were associated with a better OS. No difference was found in OS with respect to histology. This study accurately reflects outcomes for patients treated according to standards of care worldwide. Results confirm that peripheral T-cell lymphomas patients had dismal outcome after relapse or progression. Patients with chemotherapy sensitive disease who relapsed after more than 12 months might benefit from consolidation bone marrow transplantation. (Registered at clinicaltrials.gov identifier: 01142674)

Ipilimumab, Nivolumab and Brentuximab Vedotin in Patients with Relapsed Hodgkin Lymphoma: Phase 1 Results of a Multicenter Phase 1/2 Clinical Trial

Background: Recognising that the immune suppressive microenvironment promotes tumour growth in Hodgkin lymphoma, we hypothesised that activating immunity might augment the activity of targeted chemotherapy. We evaluated the safety and activity of combinations of brentuximab vedotin with nivolumab or ipilimumab, or both in patients with relapsed or refractory Hodgkin lymphoma. Methods: In this multicentre, open-label, phase 1/2 trial, patients with relapsed or refractory Hodgkin lymphoma aged 18 years or older who had relapsed after at least one line of therapy, with an Eastern Cooperative Oncology Group performance status of 2 or lower, and adequate organ and marrow function, with no pulmonary dysfunction were eligible for inclusion. Phase 1 primary objectives were to determine the maximum tolerated dose and dose limiting toxicities of brentuximab vedotin combined with ipilimumab (ipilimumab group), nivolumab (nivolumab group), or both (triplet therapy group) using a 3 + 3 dose escalation design with expansion cohorts. During the dose escalation phase, patients were enrolled sequentially into one of six cohorts: in the ipilimumab group fixed brentuximab vedotin 1·8 mg/kg with ipilimumab 1 mg/kg (cohort A) or 3 mg/kg (cohort B); in the nivolumab group fixed nivolumab 3 mg/kg with brentuximab vedotin 1·2 mg/kg (cohort D) or 1·8 mg/kg (cohort E); and in the triplet therapy group fixed nivolumab 3 mg/kg and ipilimumab 1 mg/kg with brentuximab vedotin 1·2 mg/kg (cohort G) or 1·8 mg/kg (cohort H). Additional patients were enrolled in the expansion phase at the same doses of cohorts B, E, and H. All drugs were given intravenously; brentuximab vedotin and nivolumab were given every 3 weeks, ipilimumab was given every 6 weeks in the ipilimumab group and every 12 weeks in the triplet therapy group. All eligible and treated patients were included in the analysis. This phase 1/2 study is registered with ClinicalTrials.gov, NCT01896999. The phase 2, randomised portion of the trial is still enrolling. Findings: Between March 7, 2014, and Dec 28, 2017, 64 patients were enrolled; two patients in the ipilimumab group and one patient in the nivolumab group were excluded due to ineligibility after enrolment and 61 were evaluable. A total of six dose limiting toxicities were reported in four patients, and the doses used in cohorts B, E, and H were established as maximum tolerated doses and patients were subsequently enrolled onto expansion cohorts (C, F, and I) with these schedules. There were ten (43%) grade 3-4 treatment related adverse events in the ipilimumab group, three (16%) in the nivolumab group, and 11 (50%) in the triplet therapy group including: eight (13%) of 64 patients reporting rash, and colitis, gastritis, pancreatitis and arthritis, and diabetic ketoacidosis each occurring in one (2%) patient. There were two (3%) treatment related deaths, one in the nivolumab group and one in the triplet therapy group. The overall response rate was 76% (95% CI 53-92) in the ipilimumab group, 89% (65-99) in the nivolumab group, and 82% (60-95) in the triplet therapy group, and the complete response rate was 57% (95% CI 34-78%) in the ipilimumab group, 61% (36-83%) in the nivolumab group, and 73% (50-89%) in the triplet therapy group. With a median follow-up of 2·6 years (IQR 1·8-2·9) in the ipilimumab group, 2·4 years (2·2-2·6) in the nivolumab group, and 1·7 years (1·6-1·9) in the triplet therapy group, median progression-free survival is 1·2 years (95% CI 1·7-not reached) in the ipilimumab group, but was not reached in the other two treatment groups. Median overall survival has not been reached in any of the groups. Interpretation: There are clear differences in activity and toxicity of the three combination regimens. The tolerability and preliminary activity for the two most active regimens, brentuximab vedotin with nivolumab and the triplet therapy, are being compared in a randomised phase 2 trial (NCT01896999)

Prognostic Significance of MYC Rearrangement and Translocation Partner in Diffuse Large B-Cell Lymphoma : A Study by the Lunenburg Lymphoma Biomarker Consortium

PURPOSE: MYC rearrangement (MYC-R) occurs in approximately 10% of diffuse large B-cell lymphomas (DLBCLs) and has been associated with poor prognosis in many studies. The impact of MYC-R on prognosis may be influenced by the MYC partner gene (immunoglobulin [IG] or a non-IG gene). We evaluated a large cohort of patients through the Lunenburg Lymphoma Biomarker Consortium to validate the prognostic significance of MYC-R (single-, double-, and triple-hit status) in DLBCL within the context of the MYC partner gene. METHODS: The study cohort included patients with histologically confirmed DLBCL morphology derived from large prospective trials and patient registries in Europe and North America who were uniformly treated with rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone therapy or the like. Fluorescence in situ hybridization for the MYC, BCL2, BCL6, and IG heavy and light chain loci was used, and results were correlated with clinical outcomes. RESULTS: A total of 5,117 patients were identified of whom 2,383 (47%) had biopsy material available to assess for MYC-R. MYC-R was present in 264 (11%) of 2,383 patients and was associated with a significantly shorter progression-free and overall survival, with a strong time-dependent effect within the first 24 months after diagnosis. The adverse prognostic impact of MYC-R was only evident in patients with a concurrent rearrangement of BCL2 and/or BCL6 and an IG partner (hazard ratio, 2.4; 95% CI, 1.6 to 3.6; P < .001). CONCLUSION: The negative prognostic impact of MYC-R in DLBCL is largely observed in patients with MYC double hit/triple-hit disease in which MYC is translocated to an IG partner, and this effect is restricted to the first 2 years after diagnosis. Our results suggest that diagnostic strategies should be adopted to identify this high-risk cohort, and risk-adjusted therapeutic approaches should be refined further

Targeting BTK with Ibrutinib in Relapsed or Refractory Mantle-Cell Lymphoma – Results of an International, Multicenter, Phase 2 Study of Ibrutinib (PCI-32765) – EHA Encore

Bruton's tyrosine kinase (BTK) is a central mediator of B-cell receptor (BCR) signaling essential for normal B-cell development. Ibrutinib is an oral BTK inhibitor that induces apoptosis and inhibits migration and adhesion of malignant B-cells. Updated results of this international, multicenter, phase 2 study of single agent ibrutinib in relapsed or refractory MCL will be presented.Ibrutinib 560mg PO QD was administered continuously until disease progression. Tumor response was assessed every 2 cycles (one cycle=28 days). The study enrolled 115 patients (65 bortezomib-naïve, 50 bortezomib-exposed); 111 patients were treated; 110 were evaluable for response. Baseline characteristics included: median age 68 years, time since diagnosis 42 months, number of prior treatments 3; bulky disease (>10cm) 13%, prior stem cell transplant 10%, high risk MIPI 49%.Median time on treatment was 9.2 months; 53% of patients remain on therapy. Median PFS was 13.9 months and DOR has not yet been reached. Responses increased with longer treatment: comparing to previous data described at ASH 2011, the CR rate increased from 16% to 39%, and the ORR increased from 69% to 75%

ACR Appropriateness Criteria® Recurrent Hodgkin Lymphoma

This topic addresses the management of recurrent Hodgkin lymphoma. While autologous stem cell transplantation may be appropriate for select cases of recurrent disease following comprehensive combined-modality therapy, other options exist for patients treated with lower-dose therapy for early-stage disease. Additionally, innovative targeted therapies provide newer salvage options to consider. The American College of Radiology Appropriateness Criteria® are evidence-based guidelines for specific clinical conditions that are reviewed annually by a multidisciplinary expert panel. The guideline development and revision include an extensive analysis of current medical literature from peer-reviewed journals and the application of well-established methodologies (RAND/UCLA Appropriateness Method and Grading of Recommendations Assessment, Development, and Evaluation, or GRADE) to rate the appropriateness of imaging and treatment procedures for specific clinical scenarios. In those instances where evidence is lacking or equivocal, expert opinion may supplement the available evidence to recommend imaging or treatment. By combining the most recent medical literature and expert opinion, this revised guideline can aid clinicians in the complex decisionmaking associated with the management of recurrent Hodgkin lymphoma

Integrative Analysis of Clinicopathological Features Defines Novel Prognostic Models for Mantle Cell Lymphoma in the Immunochemotherapy Era: A Report from The North American Mantle Cell Lymphoma Consortium

BACKGROUND: Patients with mantle cell lymphoma (MCL) exhibit a wide variation in clinical presentation and outcome. However, the commonly used prognostic models are outdated and inadequate to address the needs of the current multidisciplinary management of this disease. This study aims to investigate the clinical and pathological features of MCL in the immunochemotherapy era and improve the prognostic models for a more accurate prediction of patient outcomes. METHODS: The North American Mantle Cell Lymphoma Project is a multi-institutional collaboration of 23 institutions across North America to evaluate and refine prognosticators for front-line therapy. A total of 586 MCL cases diagnosed between 2000 and 2012 are included in this study. A comprehensive retrospective analysis was performed on the clinicopathological features, treatment approaches, and outcomes of these cases. The establishment of novel prognostic models was based on in-depth examination of baseline parameters, and subsequent validation in an independent cohort of MCL cases. RESULTS: In front-line strategies, the use of hematopoietic stem cell transplantation was the most significant parameter affecting outcomes, for both overall survival (OS, p \u3c 0.0001) and progression-free survival (PFS, p \u3c 0.0001). P53 positive expression was the most significant pathological parameter correlating with inferior outcomes (p \u3c 0.0001 for OS and p = 0.0021 for PFS). Based on the baseline risk factor profile, we developed a set of prognostic models incorporating clinical, laboratory, and pathological parameters that are specifically tailored for various applications. These models, when tested in the validation cohort, exhibited strong predictive power for survival and showed a stratification resembling the training cohort. CONCLUSIONS: The outcome of patients with MCL has markedly improved over the past two decades, and further enhancement is anticipated with the evolution of clinical management. The innovative prognostic models developed in this study would serve as a valuable tool to guide the selection of more suitable treatment strategies for patients with MCL

In Support of a Patient-Driven Initiative and Petition to Lower the High Price of Cancer Drugs

Comment in Lowering the High Cost of Cancer Drugs--III. [Mayo Clin Proc. 2016] Lowering the High Cost of Cancer Drugs--I. [Mayo Clin Proc. 2016] Lowering the High Cost of Cancer Drugs--IV. [Mayo Clin Proc. 2016] In Reply--Lowering the High Cost of Cancer Drugs. [Mayo Clin Proc. 2016] US oncologists call for government regulation to curb drug price rises. [BMJ. 2015