Perls’ Stain Guidelines from the French-Speaking Cellular Hematology Group (GFHC)

In order to standardize cellular hematology practices, the French-speaking Cellular Hematology Group (Groupe Francophone d’Hématologie Cellulaire, GFHC) focused on Perls’ stain. A national survey was carried out, leading to the proposal of recommendations on insoluble iron detection and quantification in bone marrow. The criteria presented here met with a “strong professional agreement” and follow the suggestions of the World Health Organization’s classification of hematological malignancies

Base-Position Error Rate Analysis of Next-Generation Sequencing Applied to Circulating Tumor DNA in Non-Small Cell Lung Cancer: A Prospective Study

International audienceBACKGROUND:Circulating tumor DNA (ctDNA) is an approved noninvasive biomarker to test for the presence of EGFR mutations at diagnosis or recurrence of lung cancer. However, studies evaluating ctDNA as a noninvasive "real-time" biomarker to provide prognostic and predictive information in treatment monitoring have given inconsistent results, mainly due to methodological differences. We have recently validated a next-generation sequencing (NGS) approach to detect ctDNA. Using this new approach, we evaluated the clinical usefulness of ctDNA monitoring in a prospective observational series of patients with non-small cell lung cancer (NSCLC).METHODS AND FINDINGS:We recruited 124 patients with newly diagnosed advanced NSCLC for ctDNA monitoring. The primary objective was to analyze the prognostic value of baseline ctDNA on overall survival. ctDNA was assessed by ultra-deep targeted NGS using our dedicated variant caller algorithm. Common mutations were validated by digital PCR. Out of the 109 patients with at least one follow-up marker mutation, plasma samples were contributive at baseline (n = 105), at first evaluation (n = 85), and at tumor progression (n = 66). We found that the presence of ctDNA at baseline was an independent marker of poor prognosis, with a median overall survival of 13.6 versus 21.5 mo (adjusted hazard ratio [HR] 1.82, 95% CI 1.01-3.55, p = 0.045) and a median progression-free survival of 4.9 versus 10.4 mo (adjusted HR 2.14, 95% CI 1.30-3.67, p = 0.002). It was also related to the presence of bone and liver metastasis. At first evaluation (E1) after treatment initiation, residual ctDNA was an early predictor of treatment benefit as judged by best radiological response and progression-free survival. Finally, negative ctDNA at E1 was associated with overall survival independently of Response Evaluation Criteria in Solid Tumors (RECIST) (HR 3.27, 95% CI 1.66-6.40, p < 0.001). Study population heterogeneity, over-representation of EGFR-mutated patients, and heterogeneous treatment types might limit the conclusions of this study, which require future validation in independent populations.CONCLUSIONS:In this study of patients with newly diagnosed NSCLC, we found that ctDNA detection using targeted NGS was associated with poor prognosis. The heterogeneity of lung cancer molecular alterations, particularly at time of progression, impairs the ability of individual gene testing to accurately detect ctDNA in unselected patients. Further investigations are needed to evaluate the clinical impact of earlier evaluation times at 1 or 2 wk. Supporting clinical decisions, such as early treatment switching based on ctDNA positivity at first evaluation, will require dedicated interventional studies

Effect of patient and tumor baseline characteristics on OS (n = 109).

<p>Effect of patient and tumor baseline characteristics on OS (n = 109).</p

Prognostic impact of positive baseline ctDNA.

<p>(A) PFS and (B) OS in patients with positive and negative ctDNA (<i>n</i> = 105).</p

Clinical characteristics associated with ctDNA concentration.

<p>Correlation between T0 ctDNA concentration tertiles and (A) tumor burden defined by the sum of the RECIST target lesions (Mann-Whitney test) and (B) presence of liver metastasis (Fisher’s exact test). (C) Correlation between positive ctDNA at baseline and Ki67 proliferative index expressed as a % of positive cells in a subset of tumors with available tissue (<i>n</i> = 19, Mann-Whitney test). *<i>p</i> < 0.05, **<i>p</i> < 0.005, ***<i>p</i> < 0.001.</p

Summary of baseline patient and tumor characteristics.

<p>Summary of baseline patient and tumor characteristics.</p

Study design.

<p>(A) Flowchart of the study population. (B) Time point of ctDNA follow-up. *Includes ALK fusion (<i>n</i> = 3) and ROS1 fusion (<i>n</i> = 1). **PIK3CA p.H1047L. ^†TP53 p.Gly244Cys. ^$KRAS p.G12V, TP53 p.C135Y, and TP53 p.R248W. No mutations were found for <i>AKT1</i>, <i>ERBB2</i>, <i>FBXW7</i>, <i>FGFR2</i>, <i>MET</i>, or <i>NOTCH1</i>.</p

Baseline ctDNA detection using NGS.

<p>On the left, the figure shows <i>EGFR</i>, <i>KRAS</i>, and <i>BRAF</i> mutation testing with dPCR and NGS. On the right, the figure shows the 105 patients eligible for baseline ctDNA NGS testing, among whom 74 were positive.</p

Mutations identified in tumor and in baseline plasma samples.

<p>Comparison of molecular alterations found in tumor tissue and/or in baseline (T0) plasma (<i>n</i> = 109 participants).</p

Plasma characteristics.

<p>Plasma characteristics.</p