AFIRMA GSC: MOLECULAR ANALYSIS IN FOLLICULAR THYROID NODULES
Afirma GSC X Atlas represents the most advanced method currently available worldwide for the genomic analysis of indeterminate thyroid nodules (Thy 3a, Thy3f, Bethesda III (AUS/FLUS), Bethesda IV).
AFIRMA GSC AND AFIRMA GEC – DIFFERENCES
The Afirma method is applied in the differential diagnosis of indeterminate nodules (Thy 3a, Thy3f, Bethesda III (AUS/FLUS), Bethesda IV) to identify benign nodules and to avoid unnecessary diagnostic surgery. The first method of Afirma was Afirma – Gene Expression Classifier (GEC), a method with high sensitivity, which allowed to reclassify as less than half of the undetermined thyroid nodules as benign. Recent technological advances have allowed us to take another step forward with the development of Afirma – Genomic Sequencing Classifier (GSC) that allows us to reclassify as benign well over half of the undetermined thyroid nodules (Figure 1, Figure 2). Afirma GSC represents an advanced method available for the genomic analysis of indeterminate thyroid nodules (Thy 3a, Thy3f, Bethesda III (AUS/FLUS), Bethesda IV).
Afirma GSC combines the classic RNA sequencing with GEC microarray, with the RNA sequencing that allows to extract as much genomic information as possible from the thyroid aspirate sample and to identify genomic information previously not detectable with other methods.
Afirma GSC, in fact, is the only test currently available in the diagnosis of the thyroid nodule that combines the sequencing of new generation of RNA and analytical methods capable of detecting even small genomic alterations not detectable by traditional methods (such as DNA-based sequencing). Afirma GSC analyzes the RNA expression profiles of more than 1,000 genes. Therefore, with Afirma GSC, in addition to gene expression, the presence of DNA variants, fusions, copy number variants and other potentially predictive information of thyroid cancer is also evaluated.
Currently, by performing the Afirma GSC analysis, the following mutations are always analyzed: RET/PTC1, RET/PTC3, BRAFV600E (for papillary carcinoma), MTC (for medullary carcinoma) (Figure 3). This powerful combination of genomic evaluations is able to further increase the ability to distinguish between benign and malignant nodules. See available packages.
AFIRMA GSC – PAPILLARY THYROID CARCINOMA (BRAFV600E)
With GSC, the already powerful genomic testing tools available in Afirma, a new classifier was added (and validated): BRAF V600E. Using advanced generation sequencing technologies, it is possible to find BRAF V600E mutations that are not detectable by other previous techniques (such as PCR). It is known that the presence of the BRAF V600E mutation in the thyroid nodules confers the certainty of being in front of a tumor. Furthermore, since BRAF positive tumors tend to be more aggressive, determining the presence (or not) of BRAF may influence therapeutic options for the patient (eg a larger surgery rather than a lobectomy). Finally, the BRAF classifier of Afirma GSC generates few false positives.
AFIRMA GSC – MEDULLARY THYROID CARCINOMA (MTC)
The Afirma GSC method also employs a classifier for medullary carcinoma (MTC classifier), which allows a more correct therapeutic approach for patients with MTC.
AFIRMA GSC – HURTLE CARCINOMA
The Afirma GSC method also uses a classifier for Hürthle cells (Hürthle classifier). To improve Hürthle cell classification, Afirma GSC first determines whether the FNA sample contains Hürthle cells by analyzing more than 1400 genes. If successful, a second classifier examines additional criteria, including the loss of heterozygosity, specific to Hürthle cell neoplasms, to distinguish adenomas from Hürthle cell carcinomas. This second classifier uses over 2000 genes and nearly 190000 genomic variants. In this way Afirma GSC is able to differentiate Hürthle cell tumors from non-tumors. Finally, the latter, like any other undetermined sample, are then reevaluated by the global classifier to determine their benignity. This has meant that the Hürthle cell nodules that were previously considered suspect by GEC, with Afirma GSC will be classified as benign with a higher degree of precision (with a specificity that increases from 12% to 59%) (Figure 4), further increasing the number of patients with benign nodules able to avoid unnecessary surgical operations.
AFIRMA GSC – PARATHYROID
The parathyroids are located near the thyroid, and if enlarged, can sometimes be mistaken for thyroid nodules. Such enlarged formations (adenomas) if subjected to needle aspiration, could result, as indeterminate formations (THY3) as the cells that constitute them can often be exchanged with thyroid cells with atypical characteristics. Afirma GSC also provides a parathyroid classifier (Parathyroid Classifier) that allows to correctly identify these formations and avoid unnecessary surgeries.
AFIRMA GSC vs AFIRMA GEC
The comparison with GEC shows that the new method Afirma GSC not only maintains a high sensitivity (91%) but clearly improves its specificity (from 52 to 68%). This means that this latest generation test is able to correctly identify as benign about 30% more nodules than Afirma GEC, with a negative predictive value of up to 96% (Figure 3). These data confirm that Afirma GSC, even more than Afirma GEC, can help avoid diagnostic surgery.
AFIRMA GSC vs OTHER METHODS
The other existing genetic methods, on the other hand, only evaluate specific mutations, and only analyze a few genes or microRNAs and therefore have limited clinical utility as some of these mutations are only rarely present in indeterminate tumor nodules (Thy3). With Afirma GSC, on the other hand, RNA sequencing interrogates the entire genome and uses very large and complex genetic information with methods previously never used in health diagnostics.
AFIRMA GSC – OTHER INFORMATIONS
Afirma GSC is highly reproducible, but above all it has a very high analytical sensitivity with a limit of detection of 5%. This means that even if the sample taken from a malignant nodule was “diluted” or “contaminated” up to 95% from the normal adjacent tissue or benign nodular tissue, Afirma GSC is still able to classify it as malignant. This minimizes false negatives, even in the case of erroneous sampling that has accidentally affected the adjacent normal thyroid tissue. Finally, the performance of Afirma GSC is poorly influenced by the classical interferents commonly found in the samples of needle aspiration (such as blood and genomic DNA).
In conclusion, Afirma GSC and its further implementaion (Afirma GSC X Atlas) represent the most advanced method currently available worldwide for the genomic analysis of indeterminate thyroid nodules (Thy 3a, Thy3f, Bethesda III (AUS/FLUS), Bethesda IV).
Afirma, provides two possible results on indeterminate thyroid nodules: benign or suspicious for malignant. Afirma GSC, by reclassifying cytologically indeterminate thyroid nodules as benign, avoids unnecessary thyroid surgery and is useful in management of cytologically indeterminate thyroid nodules.
Afirma was developed in the United States and, from the first studies, now it is widespread to other countries.
The EndocrinologiaOggi Center in Rome (Italy), is the first center in Europe where Afirma is clinically and routinately used for stratification of indeterminate thyroid nodules. European patients that prefer a closer place to perform Afirma instead of San Francisco (United States), can do it in Rome (Italy) at a lower price (2400 €).
For more information about Afirma GEC, click here.
For more information about Afirma GSC X Atlas, click here.
For an appointment for Afirma GSC X Atlas, you can:
– write an email (firstname.lastname@example.org)
– call (0039 0686391386)
– book on line (click here).
Dr. Massimiliano Andrioli
MD, PhD, Endocrinologist
Centro EndocrinologiaOggi, Roma
viale Somalia 33A, Roma
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