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Several platforms for noninvasive EGFR testing are currently used in the clinical setting with sensitivities ranging from 30% to 100%. Prospective studies evaluating agreement and sources for discordant results remain lacking. Herein, seven methodologies including two next-generation sequencing (NGS)-based methods, three high-sensitivity PCR-based platforms, and two FDA-approved methods were compared using 72 plasma samples, from EGFR-mutant non-small-cell lung cancer (NSCLC) patients progressing on a first-line tyrosine kinase inhibitor (TKI). NGS platforms as well as high-sensitivity PCR-based methodologies showed excellent agreement for EGFR-sensitizing mutations (K = 0.80–0.89) and substantial agreement for T790M testing (K = 0.77 and 0.68, respectively). Mutant allele frequencies (MAFs) obtained by different quantitative methods showed an excellent reproducibility (intraclass correlation coefficients 0.86–0.98). Among other technical factors, discordant calls mostly occurred at mutant allele frequencies (MAFs) ≤ 0.5%. Agreement significantly improved when discarding samples with MAF ≤ 0.5%. EGFR mutations were detected at significantly lower MAFs in patients with brain metastases, suggesting that these patients risk for a false-positive result. Our results support the use of liquid biopsies for noninvasive EGFR testing and highlight the need to systematically report MAFs.

Estrone is a powerful growth-inducing hormone that is present in milk, mainly in the form of fatty acid esters, at concentrations that promote growth in experimental animals. We present here a method useful for the measurement of this natural hormone in foods and applied it to several common dairy products. Samples were frozen, finely powdered, and lyophilized then extracted with trichloromethane/methanol; the dry extract was saponified with potassium hydroxide. The free estrone evolved was extracted with ethyl acetate and was used for the estimation of total estrone content through radioimmunoassay. Application of the method to dairy products showed high relative levels of total estrone (essentially acyl-estrone) in milk, in the range of 1 ¿M, which were halved in skimmed milk. Free estrone levels were much lower, in the nanomolar range. A large proportion of estrone esters was present in all other dairy products, fairly correlated with their fat content. The amount of estrone carried by milk is well within the range, where its intake may exert a physiological response in the sucklings for which it is provided. These growth-inducing and energy expenditure-lowering effects may affect humans ingesting significant amounts of dairy products.