The HDL mimetic CER-001 remodels plasma lipoproteins and reduces kidney lipid deposits in inherited lecithin:cholesterol acyltransferase deficiency.

BACKGROUND: Kidney failure is the major cause of morbidity and mortality in familial lecithin:cholesterol acyltransferase deficiency (FLD), a rare inherited lipid disorder with no cure. Lipoprotein X (LpX), an abnormal lipoprotein, is primarily accountable for nephrotoxicity. METHODS: CER-001 was tested in an FLD patient with dramatic kidney disease for 12 weeks. RESULTS: Infusions of CER-001 normalized the lipoprotein profile, with a disappearance of the abnormal LpX in favour of normal-sized LDL. The worsening of kidney function was slowed by the treatment, and kidney biopsy showed a slight reduction of lipid deposits and a stabilization of the disease. In vitro experiments demonstrate that CER-001 progressively reverts lipid accumulation in podocytes by a dual effect: remodelling plasma lipoproteins and removing LpX-induced lipid deposit. CONCLUSION: This study demonstrates that CER-001 may represent a therapeutic option in FLD patients. It also has the potential to be beneficial in other renal diseases characterized by kidney lipid deposits.

Non-small cell lung cancer harboring EGFR G724S mutation and exon 19 deletion responded to afatinib monotherapy after multiple lines of target therapies.

Epidermal growth factor receptor (EGFR) G724S mutation represents a resistance mechanism to first- and third-generation EGFR tyrosine kinase inhibitors. Limited data are available regarding the efficacy of afatinib in patients with non-small cell lung cancer (NSCLC) harboring G724S mutation, particularly after osimertinib. A patient diagnosed with advanced EGFR-mutated (exon 19 deletion) NSCLC after several lines of treatment - gefitinib, osimertinib, heat shock protein inhibitors and chemotherapy-developed EGFR G724S mutation retaining the exon 19 deletion. She was then treated successfully with afatinib leading to a progression free survival of 9 months (and counting). This is the first report of the emergence of G724S mutation, together with ex19del, after three subsequent lines of therapy following progressive disease to Osimertinib, and we report for the first time the activity of afatinib against EGFR exon 18 G724S mutation in this setting.

Real-world data on treatment outcomes in EGFR-mutant non-small-cell lung cancer patients receiving osimertinib in second or further lines.

Aims: This study describes real-world outcomes of pretreated EGFR T790M-positive (T790M+) advanced non-small-cell lung cancer patients progressing after first- or second-generation tyrosine kinase inhibitors and receiving osimertinib, compared with T790M-negative (T790M-) patients. We have also described progression patterns and treatment sequences. Patients & methods: This is a retrospective multicenter Italian observational study including consecutive Caucasian patients referred between 2014 and 2018. Results: 167 patients were included. Median progression-free survival was 9.8 months (95% CI: 8.3-13.3) for T790M+ and 6.0 months (95% CI: 4.9-7.2) for T790M- patients, respectively. Median overall survival was 20.7 months (95% CI: 18.9-28.4) for T790M+ and 10.6 months (95% CI: 8.6-23.6) for T790M- patients, respectively. The T790M mutation correlated with absence of new sites of disease. After progression, most T790M+ patients continued osimertinib, whereas most T790M- patients received a different treatment line. Conclusion: Better outcomes were shown in patients receiving osimertinib. A more limited progression pattern for T790M+ was suggested.

Lay abstract Osimertinib is an oral drug that inhibits the growth of non-small-cell lung cancer (NSCLC) tumors with a specific mutation in EGFR. Osimertinib is given to patients with advanced EGFR-mutant NSCLC as initial therapy or after the failure of prior first- or second-generation tyrosine kinase inhibitors in patients who develop the EGFR T790M resistance mutation. Real-world data about the efficacy of EGFR-mutant NSCLC patients receiving osimertinib are needed to confirm the findings of large randomized clinical trials. Most real-world studies have investigated outcomes in Asian populations. This study aims to describe outcomes in EGFR T790M-positive patients receiving osimertinib after the failure of first- or second-generation tyrosine kinase inhibitors, compared with T790M-negative patients receiving a systemic treatment, in a Caucasian population. In addition, the study aims to describe how the disease spreads once it starts progressing again and any subsequent treatment lines. 167 patients were included. The results of this study suggest that EGFR T790M-positive patients receiving osimertinib as second- or further-line treatment had better outcomes and a more limited progression compared with T790M-negative cases.

Current prognostic and predictive biomarkers for gastrointestinal tumors in clinical practice.

The pathologist emerged in the personalized medicine era as a central actor in the definition of the most adequate diagnostic and therapeutic algorithms. In the last decade, gastrointestinal oncology has seen a significantly increased clinical request for the integration of novel prognostic and predictive biomarkers in histopathological reports. This request couples with the significant contraction of invasive sampling of the disease, thus conferring to the pathologist the role of governor for both proper pathologic characterization and customized processing of the biospecimens. This overview will focus on the most commonly adopted immunohistochemical and molecular biomarkers in the routine clinical characterization of gastrointestinal neoplasms referring to the most recent published recommendations, guidelines and expert opinions.

Validated clinico-pathologic nomogram in the prediction of HER2 status in gastro-oesophageal cancer.

BACKGROUND: HER2 is the only validated predictive biomarker in gastro-oesophageal carcinoma (GOC). However, several factors, such as heterogeneity in protein expression, shortage of evaluable tumour tissue and need for quick target assessment, underline the usefulness of a pre-screening tool in order to anticipate HER2 status. METHODS: Data from 723 consecutive GOC analysed for HER2 at four Italian Institutions were collected. HER2 positivity was defined as 3+ by immunohistochemistry (IHC) or 2+ with gene amplification by in situ hybridisation (ISH). A multivariate logistic regression model was built using data from 413 cases, whereas 310 patients served as validation cohort. C-index, visual inspection of the calibration plot, Brier score and Spiegelhalter z-test were used to assess the performance of the nomogram. RESULTS: HER2 positive rate was 17.4%. Four variables were retained after adjustment in the final model: grading, Lauren's histotype, pathologic material analysed (surgical specimen/biopsy) and site of tissue collection (primary tumour/metastases). Visual inspection of the calibration plot revealed a very good overlap between predicted and observed probabilities, with a Brier score of 0.101 and a non-significant Spiegelhalter z-test (P = 0.319). C-index resulted in 0.827 (95%CI 0.741-0.913). CONCLUSION: A simple nomogram based on always-available pathologic information accurately predicts the probability of HER2 positivity in GOC.

CK7 and consensus molecular subtypes as major prognosticators in V600EBRAF mutated metastatic colorectal cancer.

BACKGROUND: V600EBRAF mutated metastatic colorectal cancer (mCRC) is a subtype (10%) with overall poor prognosis, but the clinical experience suggests a great heterogeneity in survival. It is still unexplored the real distribution of traditional and innovative biomarkers among V600EBRAF mutated mCRC and which is their role in the improvement of clinical prediction of survival outcomes. METHODS: Data and tissue specimens from 155 V600EBRAF mutated mCRC patients treated at eight Italian Units of Oncology were collected. Specimens were analysed by means of immunohistochemistry profiling performed on tissue microarrays. Primary endpoint was overall survival (OS). RESULTS: CDX2 loss conferred worse OS (HR = 1.72, 95%CI 1.03-2.86, p = 0.036), as well as high CK7 expression (HR = 2.17, 95%CI 1.10-4.29, p = 0.026). According to Consensus Molecular Subtypes (CMS), CMS1 patients had better OS compared to CMS2-3/CMS4 (HR = 0.37, 95%CI 0.19-0.71, p = 0.003). Samples showing less TILs had worse OS (HR = 1.72, 95%CI 1.16-2.56, p = 0.007). Progression-free survival analyses led to similar results. At multivariate analysis, CK7 and CMS subgrouping retained their significant correlation with OS. CONCLUSION: The present study provides new evidence on how several well-established biomarkers perform in a homogenousV600EBRAF mutated mCRC population, with important and independent information added to standard clinical prognosticators. These data could be useful to inform further translational research, for patients' stratification in clinical trials and in routine clinical practice to better estimate patients' prognosis.

Effects of KEAP1 Silencing on the Regulation of NRF2 Activity in Neuroendocrine Lung Tumors.

BACKGROUND: The KEAP1/NRF2 pathway has been widely investigated in tumors since it was implicated in cancer cells survival and therapies resistance. In lung tumors the deregulation of this pathway is mainly related to point mutations of KEAP1 and NFE2L2 genes and KEAP1 promoter hypermethylation, but these two genes have been rarely investigated in low/intermediate grade neuroendocrine tumors of the lung. METHODS: The effects of KEAP1 silencing on NRF2 activity was investigated in H720 and H727 carcinoid cell lines and results were compared with those obtained by molecular profiling of KEAP1 and NFE2L2 in a collection of 47 lung carcinoids. The correlation between methylation and transcript levels was assessed by 5-aza-dC treatment. RESULTS: We demonstrated that in carcinoid cell lines, the KEAP1 silencing induces an upregulation of NRF2 and some of its targets and that there is a direct correlation between KEAP1 methylation and its mRNA levels. A KEAP1 hypermethylation and Loss of Heterozygosity at KEAP1 gene locus was also observed in nearly half of lung carcinoids. CONCLUSIONS: This is the first study that has described the effects of KEAP1 silencing on the regulation of NRF2 activity in lung carcinoids cells. The epigenetic deregulation of the KEAP1/NRF2 by a KEAP1 promoter hypermethylation system appears to be a frequent event in lung carcinoids.

Defining a prognostic score based on O6-methylguanine-DNA methyltransferase cut-off methylation level determined by pyrosequencing in patients with glioblastoma multiforme.

PURPOSE: Epigenetic variations in the O6-methylguanine-methyltransferase gene had been widely associated with a favorable impact on survival in patients affected by glioblastoma multiforme (GBM). Aim of this study is to explore a scoring system based on the gene promoter methylation in order to predict patients' prognosis. METHODS: A series of 128 patients with GBM was retrospectively analyzed. A training set and a validations set were then generated. The methylation level of CpGi from 74 to 83 was determined by pyrosequencing. In accordance to previous literature, each island was assigned with 1 point if the corresponding methylation level was higher than 9%. The sum consisted in a score that went from 0 (all CpGi < 9%) to 10 (all CpGi ≥ 9%). A threshold capable to detect a favorable outcome (overall survival, OS > 24 months) was identified by ROC analysis. RESULTS: Median OS and follow-up were 14 and 32.6 months respectively. Among the total population, 35% of the pts had a score of 0, while 29% had a score of 10. A score ≥ 6 was associated with a favorable prognosis also when corrected for age (> 70 vs. ≤ 70 years) and ECOG performance status (0-1 vs. 2-3). Similar results were observed also in terms of PFS. Results were consistent in the training and in the validation set. CONCLUSIONS: The present manuscript explored a novel scoring system capable to take into consideration the methylation status of each single CpGi, capable to better predict prognosis in GBM patients.

MGMT pyrosequencing-based cut-off methylation level and clinical outcome in patients with glioblastoma multiforme.

AIM: MGMT promoter methylation has been associated with improved survival in glioblastoma multiforme treated with temozolomide. However, there is no consensus on specific cut-off levels of methylation. The aims of the study were to explore the prognostic impact of MGMT methylation status and to analyze the role of specific cut-off values. MATERIALS & METHODS: We analyzed 108 glioblastoma multiforme patients treated between 2008 and 2013 stratified according to three pyrosequencing-based quantitative methylation in: unmethylated (methylation <9%), intermediate (9-29%) and highly methylated (>29%). RESULTS: The three-class stratification has a prognostic impact (median progression-free survival: 7.97, 11.6 and 15 months respectively; p = 0.004; median OS: 13.2, 15.8 and 19.5 months, respectively; p = 0.0002), especially in patients exposed to temozolomide. CONCLUSION: Our study confirmed that the independent prognostic role of MGMT methylation status. An average level of methylation between all investigated CpGs of 9% may help discriminating between methylated and unmethylated tumors.

Panitumumab use in metastatic colorectal cancer and patterns of RAS testing: results from a Europe-wide physician survey and medical records review.

BACKGROUND: In Europe, treatment of metastatic colorectal cancer (mCRC) with panitumumab requires prior confirmation of RAS wild-type mutation status. Two studies - a physician survey and a medical records review (MRR) - were conducted to evaluate the use of panitumumab and awareness among prescribing oncologists of the associated RAS testing requirements in clinical practice. METHODS: Both studies enrolled participants from nine European countries and were carried out in three consecutive rounds. Rounds 1 and 2 (2012-2013) examined KRAS (exon 2) testing only; the results have been published in full previously. Round 3 (2014-2015) examined full RAS testing (exons 2, 3, 4 of KRAS and NRAS) and was initiated following a change in prescribing guidelines, from requiring KRAS alone to requiring full RAS testing. For the physician survey, telephone interviews were conducted with oncologists who had prescribed panitumumab to patients with mCRC in the previous 6 months. For the MRR, oncologists were asked to provide anonymised clinical information, extracted from their patients' records. RESULTS: In Round 3, 152 oncologists and 131 patients' records were included in the physician survey and MRR, respectively. In Round 3 of the physician survey, 95.4% (n = 145) of participants correctly identified that panitumumab should only be prescribed in RAS wild-type mCRC compared with 99.0% (n = 298) of 301 participants in Rounds 1 and 2, responding to the same question about KRAS testing. In Round 3 of the MRR, 100% (n = 131) of patients included in the study had confirmed KRAS or RAS wild-type status prior to initiation of panitumumab compared with 97.7% (n = 299) of 306 patients in Rounds 1 and 2 (KRAS only). Of those patients in Round 3, 83.2% (n = 109) had been tested for RAS status and 16.8% (n = 22) had been tested for KRAS status only. CONCLUSIONS: Physicians' adherence to prescribing guidelines has remained high over time in Europe, despite the change in indication for panitumumab treatment, from KRAS to RAS wild-type mCRC. Additionally, this study demonstrates the uptake of full RAS testing among the majority of oncologists and pathologists.

Comparison of the molecular profile of brain metastases from colorectal cancer and corresponding primary tumors.

AIM: Little is known about molecular biology of brain metastasis (BM) from colorectal cancer and its concordance with matched primary tumors. MATERIALS & METHODS: We identified 56 consecutive colorectal cancer patients who underwent neurosurgical resection of BM. Tumor samples were tested for KRAS, NRAS, BRAF and PIK3CA. The molecular profile of the brain lesion was compared with the corresponding primary tumor. RESULTS: The molecular profile concordance rate was 95.1%. Median survival after neurosurgery was 5.5 months (95% CI: 4.7-6.3); median overall survival was 24.0 months (95% CI: 15.6-32.4). CONCLUSION: In this cohort, we report a high frequency of KRAS mutations and a very high concordance rate between the molecular status of BM and that of matched primary tumors.

Clinico-pathological nomogram for predicting BRAF mutational status of metastatic colorectal cancer.

BACKGROUND: In metastatic colorectal cancer (mCRC), BRAFV600E mutation has been variously associated to specific clinico-pathological features. METHODS: Two large retrospective series of mCRC patients from two Italian Institutions were used as training-set (TS) and validation-set (VS) for developing a nomogram predictive of BRAFV600E status. The model was internally and externally validated. RESULTS: In the TS, data from 596 mCRC patients were gathered (RAS wild-type (wt) 281 (47.1%); BRAFV600E mutated 54 (9.1%)); RAS and BRAFV600E mutations were mutually exclusive. In the RAS-wt population, right-sided primary (odds ratio (OR): 7.80, 95% confidence interval (CI) 3.05-19.92), female gender (OR: 2.90, 95% CI 1.14-7.37) and mucinous histology (OR: 4.95, 95% CI 1.90-12.90) were independent predictors of BRAFV600E mutation, with high replication at internal validation (100%, 93% and 98%, respectively). A predictive nomogram was calculated: patients with the highest score (right-sided primary, female and mucinous) had a 81% chance to bear a BRAFV600E-mutant tumour; accuracy measures: AUC=0.812, SE:0.034, sensitivity:81.2%; specificity:72.1%. In the VS (508 pts, RAS wt: 262 (51.6%), BRAFV600E mutated: 49 (9.6%)), right-sided primary, female gender and mucinous histology were confirmed as independent predictors of BRAFV600E mutation with high accuracy. CONCLUSIONS: Three simple and easy-to-collect characteristics define a useful nomogram for predicting BRAF status in mCRC with high specificity and sensitivity.

Lipophyllodes of the breast. A reappraisal of fat-rich tumors of the breast based on 22 cases integrated by immunohistochemical study, molecular pathology insights, and clinical follow-up.

We have studied 22 cases of mammary lipophyllodes tumors (LPT), analyzing their clinicopathologic features along with available follow-up. All cases were tested for cytokeratins, S100 protein, and MDM2, and in selected cases for estrogen receptor, smooth muscle actin, bcl2, desmin, and myogenin. Patients were women aged 21 to 69 years (average, 45 years), and LPT size ranged from 1.6 to 30 cm (average, 9.7 cm). Microscopically, LPT segregated as follows: atypical lipoma-like tumor/well-differentiated liposarcoma (ALT/WDL), 8 cases; myxoid, 6; and pleomorphic/poorly differentiated/round cell, 8, including a case of dedifferentiated liposarcoma. Immunohistochemistry studies showed focal positive staining for S100 and CD34 in most ALT/WDL, and desmin and myogenin in 2 cases with evidence of rhabdomyoblastic differentiation. MDM2 positivity was focally seen in 1 case. Follow-up was available in 8 cases. Multiple recurrent tumors were seen in 2 patients, and metastatic disease to the lung was seen in 2 patients. In 4 patients with a follow-up between 2 and 15 years there was no evidence of recurrent or metastatic disease. Patients with ALT/WDL (2/2) were alive with no evidence of disease; 2 of 4 patients with myxoid liposarcoma component experienced tumor recurrence, whereas pleomorphic liposarcoma LPT pursued a less favorable course although only 1 patient died of the condition. Absence of MDM2 reactivity in most cases seems not as meaningful as in fatty tumors of somatic soft parts.

A 16q deletion involving FOXF1 enhancer is associated to pulmonary capillary hemangiomatosis.

BACKGROUND: Pulmonary capillary hemangiomatosis (PCH) is an uncommon pulmonary disorder, with variable clinical features depending on which lung structure is affected, and it is usually linked to pulmonary arterial hypertension. Congenital PCH has been very rarely described and, so far, the only causative gene identified is EIF2AK4, which encodes for a translation initiation factor. However, not all PCH cases might carry a mutation in this gene. CASE PRESENTATION: We report the clinical and cytogenetic characterization of a patient (male, newborn, first child of healthy non-consanguineous parents) died after three days of life with severe neonatal pulmonary hypertension, due to diffuse capillary hemangiomatosis diagnosed post mortem. Conventional karyotyping, Microarray-Based Comparative Genomic Hydridization (CGHa) and quantitative PCR were performed. CGHa revealed a heterozygous chromosome 16q23.3q24.1 interstitial deletion, spanning about 2.6 Mb and involving a FOXF1 gene enhancer. Quantitative PCR showed that the proband's deletion was de novo. Microsatellite analysis demonstrate that the deletion occurred in the maternal chromosome 16. CONCLUSION: FOXF1 loss of function mutation have been so far identified in alveolar capillary dysplasia with misalignment of pulmonary veins (ACD/MPV), a lung disease different from PCH. Our data suggest the hypothesis that disruption of the FOXF1 gene enhancer could be a genetic determinant of PCH. Moreover, our findings support the idea that FOXF1 is a paternally imprinted gene.

Glioma-associated stem cells: a novel class of tumor-supporting cells able to predict prognosis of human low-grade gliomas.

BACKGROUND: Translational medicine aims at transferring advances in basic science research into new approaches for diagnosis and treatment of diseases. Low-grade gliomas (LGG) have a heterogeneous clinical behavior that can be only partially predicted employing current state-of-the-art markers, hindering the decision-making process. To deepen our comprehension on tumor heterogeneity, we dissected the mechanism of interaction between tumor cells and relevant components of the neoplastic environment, isolating, from LGG and high-grade gliomas (HGG), proliferating stem cell lines from both the glioma stroma and, where possible, the neoplasm. METHODS AND FINDINGS: We isolated glioma-associated stem cells (GASC) from LGG (n=40) and HGG (n=73). GASC showed stem cell features, anchorage-independent growth, and supported the malignant properties of both A172 cells and human glioma-stem cells, mainly through the release of exosomes. Finally, starting from GASC obtained from HGG (n=13) and LGG (n=12) we defined a score, based on the expression of 9 GASC surface markers, whose prognostic value was assayed on 40 subsequent LGG-patients. At the multivariate Cox analysis, the GASC-based score was the only independent predictor of overall survival and malignant progression free-survival. CONCLUSIONS: The microenvironment of both LGG and HGG hosts non-tumorigenic multipotent stem cells that can increase in vitro the biological aggressiveness of glioma-initiating cells through the release of exosomes. The clinical importance of this finding is supported by the strong prognostic value associated with the characteristics of GASC. This patient-based approach can provide a groundbreaking method to predict prognosis and to exploit novel strategies that target the tumor stroma.

MGMT promoter methylation status in brain metastases from colorectal cancer and corresponding primary tumors.

BACKGROUND: Brain metastases (BM) from colorectal cancer are usually associated with poor prognosis. The aim of this retrospective study is to evaluate MGMT promoter methylation in BM and their corresponding primary colorectal cancer tumors. MATERIALS & METHODS: MGMT promoter methylation status was assessed by pyrosequencing in 53 consecutive patients resected for BM. A concordance analysis between BM and matched primary tumor was performed in 39 cases. RESULTS: MGMT methylation was found in 34 (64.2%) BM and in 25 corresponding primary tumors (64.1%). Median survival after neurosurgery was independent from MGMT promoter methylation (163 days for those with methylated MGMT versus 193 days for the unmethylated). CONCLUSION: Epigenetic MGMT promoter methylation was common and the concordance between primary and secondary lesions was high.

Prognostic role of KRAS, NRAS, BRAF and PIK3CA mutations in advanced colorectal cancer.

AIM: To explore the prognostic value of extended mutational profiling for metastatic colorectal cancer (mCRC). MATERIALS & METHODS: We retrospectively reviewed survival results of 194 mCRC patients that were assigned to four molecular subgroups: BRAF mutated; KRAS mutated codons 12-13 only; any of KRAS codons 61-146, PIK3CA or NRAS mutations and all wild-type. Point mutations were investigated by pyrosequencing. RESULTS: BRAF (5.2%) and KRAS 12-13 (31.9%) mutations were associated with poorer survival (HR 2.8 and 1.76, respectively). Presenting with right-sided colon cancer, not resected primary tumor, WBC >10 × 10(9)/l, receiving less chemotherapy or no bevacizumab were all associated with inferior outcome. The all-wild-type subgroup (39.2%) reported the longest survival. CONCLUSION: Extended mutational profile combined with clinical factors may impact on survival in mCRC.

A new germline VHL gene mutation in three patients with apparently sporadic pheochromocytoma.

CONTEXT: Germline mutations in four genes (RET, VHL, SDHB and SDHD) are detected in about 17% of patients with apparently sporadic pheochromocytoma. Thus, genetic screening of all patients with this disease is suggested for a rational diagnostic approach and management. OBJECTIVE: To report the clinical, biochemical and genetic analysis of three unrelated patients affected by pheochromocytoma. DESIGN AND PATIENTS: All the coding regions and exon-intron boundaries of RET, VHL, SDHB and SDHD genes were sequenced in three unrelated patients with intra-adrenal pheochromocytoma: a 17-year-old girl, a 15-year-old boy and a 73-year-old man. The family history of all three cases was negative for von Hippel-Lindau lesions or other types of endocrine tumours. Structural modelling of the VHL protein was then performed. RESULTS: We identified a novel germline VHL gene point mutation, a G to A nucleotide substitution in exon 3, leading to an aspartate to asparagine amino acid change in codon 197 (D197N). No mutations were found in RET, SDHB and SDHD genes. Structural modelling of the VHL protein suggests that the D197N mutation could have a functional role. CONCLUSIONS: Our study expands the number of VHL gene known mutations and indicates the usefulness of performing the genetic analysis in all patients with apparently sporadic pheochromocytoma.

HER-2 Expression in Brain Metastases from Colorectal Cancer and Corresponding Primary Tumors: A Case Cohort Series.

Brain metastases (BM) from colorectal cancer (CRC) are a rare but increasing event. Surgical resection of oligometastatic disease, including BM, may produce a survival benefit in selected patients. Previous studies described the HER-2 expression patterns in CRC patients, but its prognostic role still remains controversial. Information on the HER-2 expression in BM from CRC is currently lacking. Among the over 500 patients treated at our Department of Neurosurgery in the last 13 years (1999-2012), we identified a cohort of 50 consecutive CRC patients resected for BM. Clinical data were retrospectively reviewed using electronic hospital charts and surgical notes. Formalin-fixed, paraffin-embedded tissue samples were retrieved and histologically reviewed. HER-2 status was assessed on 4-µm sections by HerceptTest™, and scored by two pathologists according to gastric cancer HER-2 status guidelines. In score 2+ cases HER-2 gene copy number was analyzed by FISH, performed using the PathVysion HER-2 DNA Probe Kit. Median age at time of BM resection was 65 years (35-82); most patients were males (60%) with a good performance status. The majority of the BM were single (74%) and sited in the supratentorial area (64%); 2-4 lesions were diagnosed in 9 patients (18%), and >4 in 3 patients (6%). The rate of HER-2 positivity (defined as IHC score 3+ or IHC score 2+ and FISH gene amplification) was 8.1% for the primary CRC tumors and 12% for their corresponding BM. The concordance rate between primary tumors and matched BM was 89%. Median overall survival after neurosurgery was 6.5 months for HER-2 IHC score 0 vs. 4.6 months for HER-2 IHC score 1+/2+/3+; the difference was statistically significant (p = 0.01, Log-rank test). HER-2 positivity of our case cohort was low but comparable to literature. Concordance rate of HER-2 expression between BM and corresponding primary tumors is high and similar to those reported for breast and gastric cancers. Our data suggest a potential negative prognostic value of HER-2 expression in brain lesions from CRC.

FACILITATE: A real-world, multicenter, prospective study investigating the utility of a rapid, fully automated real-time PCR assay versus local reference methods for detecting epidermal growth factor receptor variants in NSCLC.

Accurate testing for epidermal growth factor receptor (EGFR) variants is essential for informing treatment decisions in non-small cell lung cancer (NSCLC). Automated diagnostic workflows may allow more streamlined initiation of targeted treatments, where appropriate, while comprehensive variant analysis is ongoing. FACILITATE, a real-world, prospective, multicenter, European study, evaluated performance and analytical turnaround time of the Idylla™ EGFR Mutation Test compared with local reference methods. Sixteen sites obtained formalin-fixed paraffin-embedded biopsy samples with ≥ 10% neoplastic cells from patients with NSCLC. Consecutive 5 µm sections from patient samples were tested for clinically relevant NSCLC-associated EGFR variants using the Idylla™ EGFR Mutation Test and local reference methods; performance (concordance) and analytical turnaround time were compared. Between January 2019 and November 2020, 1,474 parallel analyses were conducted. Overall percentage agreement was 97.7% [n = 1,418; 95% confidence interval (CI): 96.8-98.3], positive agreement, 87.4% (n = 182; 95% CI: 81.8-91.4) and negative agreement, 99.2% (n = 1,236; 95% CI: 98.5-99.6). There were 38 (2.6%) discordant cases. Ninety percent of results were returned with an analytical turnaround time of within 1 week using the Idylla™ EGFR Mutation Test versus ∼22 days using reference methods. The Idylla™ EGFR Mutation Test performed well versus local methods and had shorter analytical turnaround time. The Idylla™ EGFR Mutation Test can thus support application of personalized medicine in NSCLC.