Daily Practice Assessment of KRAS Status in NSCLC Patients: A New Challenge for the Thoracic Pathologist Is Right around the Corner.

KRAS mutations are among the most frequent genomic alterations identified in non-squamous non-small cell lung carcinomas (NS-NSCLC), notably in lung adenocarcinomas. In most cases, these mutations are mutually exclusive, with different genomic alterations currently known to be sensitive to therapies targeting EGFR, ALK, BRAF, ROS1, and NTRK. Recently, several promising clinical trials targeting KRAS mutations, particularly for KRAS G12C-mutated NSCLC, have established new hope for better treatment of patients. In parallel, other studies have shown that NSCLC harboring co-mutations in KRAS and STK11 or KEAP1 have demonstrated primary resistance to immune checkpoint inhibitors. Thus, the assessment of the KRAS status in advanced-stage NS-NSCLC has become essential to setting up an optimal therapeutic strategy in these patients. This stimulated the development of new algorithms for the management of NSCLC samples in pathology laboratories and conditioned reorganization of optimal health care of lung cancer patients by the thoracic pathologists. This review addresses the recent data concerning the detection of KRAS mutations in NSCLC and focuses on the new challenges facing pathologists in daily practice for KRAS status assessment.

Setting-Up a Rapid SARS-CoV-2 Genome Assessment by Next-Generation Sequencing in an Academic Hospital Center (LPCE, Louis Pasteur Hospital, Nice, France).

Introduction: Aside from the reverse transcription-PCR tests for the diagnosis of the COVID-19 in routine clinical care and population-scale screening, there is an urgent need to increase the number and the efficiency for full viral genome sequencing to detect the variants of SARS-CoV-2. SARS-CoV-2 variants assessment should be easily, rapidly, and routinely available in any academic hospital. Materials and Methods: SARS-CoV-2 full genome sequencing was performed retrospectively in a single laboratory (LPCE, Louis Pasteur Hospital, Nice, France) in 103 SARS-CoV-2 positive individuals. An automated workflow used the Ion Ampliseq SARS-CoV-2 panel on the Genexus Sequencer. The analyses were made from nasopharyngeal swab (NSP) (n = 64) and/or saliva (n = 39) samples. All samples were collected in the metropolitan area of the Nice city (France) from September 2020 to March 2021. Results: The mean turnaround time between RNA extraction and result reports was 30 h for each run of 15 samples. A strong correlation was noted for the results obtained between NSP and saliva paired samples, regardless of low viral load and high (>28) Ct values. After repeated sequencing runs, complete failure of obtaining a valid sequencing result was observed in 4% of samples. Besides the European strain (B.1.160), various variants were identified, including one variant of concern (B.1.1.7), and different variants under monitoring. Discussion: Our data highlight the current feasibility of developing the SARS-CoV-2 next-generation sequencing approach in a single hospital center. Moreover, these data showed that using the Ion Ampliseq SARS-CoV-2 Assay, the SARS-CoV-2 genome sequencing is rapid and efficient not only in NSP but also in saliva samples with a low viral load. The advantages and limitations of this setup are discussed.

High expression of TRF2, SOX10, and CD10 in circulating tumor microemboli detected in metastatic melanoma patients. A potential impact for the assessment of disease aggressiveness.

Circulating tumors cells (CTCs) can be detected in the blood of metastatic melanoma patients (MMPs) both as isolated circulating tumor cells (iCTCs) and circulating tumor microemboli (CTMs), but their clinical significance remains unknown. The aim of this work was to evaluate the prognostic impact in metastatic cutaneous melanoma of CTMs and iCTCs identified by a cytomorphological approach using the isolation by size of tumor cell (ISET) method. We characterized the phenotype of CTCs using anti-PS100, anti-SOX10, anti-CD10, and anti-TRF2 antibodies. 128 MMPs and 37 control healthy individuals with benign nevi were included in this study. Results were compared to the follow-up of patients. 109/128 (85%) MMPs showed CTCs, 44/128 (34%) with 2 to 6 CTMs and 65/128 (51%) with 4 to 9 iCTCs. PS100 expression was homogeneous in iCTCs and heterogeneous in CTMs. SOX10, CD10, and TRF2 were mainly expressed in CTMs. None of the control subjects demonstrated circulating malignant tumor cells. Overall survival was significantly decreased in patients with CTMs, independently of the therapeutic strategies. In conclusion, the presence of CTMs is an independent predictor of shorter survival from the time of diagnosis of MMPs.

Assessment of morphology, antigenicity, and nucleic acid integrity for diagnostic thyroid pathology using formalin substitute fixatives.

BACKGROUND: With the advent of the formaldehyde standard law in France, and because of the impact of new methods for diagnosis and prognosis in pathology, formalin replacement in surgical pathology laboratories is currently being discussed in France. However, a set of criteria must be assessed before introducing a formalin substitute fixative. The objective of this study was to compare formalin substitute fixation with formalin fixation and cryoconservation of tissues from several benign and malignant thyroid pathologies with respect to morphology, antigenicity, and nucleic acid (RNA, DNA, microRNA) integrity. METHODS: Calibrated specimens (200 mg, 1 cm(2) each) from four conventional papillary thyroid carcinomas, four follicular variant of papillary thyroid carcinomas, three minimally invasive follicular carcinomas, four thyroid adenomas, five thyroid nodular hyperplasias, and five normal thyroid tissues were fixed for 6, 12, or 24 hours, in different fixatives (formalin, Glyo-Fixx, FineFIX, ExcellPlus, RCL2) at room temperature or at 4 degrees C. Tissues were stained (hematoxylin-eosin, periodic acid Schiff, trichromic Masson, and Sweet-Gordon staining) and their antigenicity determined by immunohistochemistry (performed with HBME-1, galectin-3, CK19, vimentin, CD31, and KL1 antibodies). Evaluation by four pathologists was made blinded. The quantity and quality of DNA, RNA, and two representative microRNA extracted from deparaffinized sections of paraffin embedded specimen were compared with that of cryosections. RESULTS: The staining and morphology were not altered by the use of different fixatives. However, formalin, FineFIX, and RCL2 gave the best results for immunohistochemistry. Moreover, FineFIX and RCL2 gave the highest amount of nucleic acids and of the best quality. CONCLUSIONS: All the formalin substitute fixatives used in this study provided good histomorphologic quality for the different stained thyroid tissues, but individually, some fixatives performed better for immunohistochemical and molecular biological procedures for different thyroid pathologies.