Although the clinical and pathologic diagnosis of some melanomas is clear-cut, there are many histopathologic simulators of melanoma that pose problems. Over-diagnosis of melanoma can lead to inappropriate therapy and psychologic burdens, whereas under-diagnosis can lead to inadequate treatment of a deadly cancer. We used existing data on DNA copy number alterations in melanoma to assemble panels of fluorescence in situ hybridization (FISH) probes suitable for the analysis of paraffin-embedded tissue. Using FISH data from a training set of 301 tumors, we established a discriminatory algorithm and validated it on an independent set of 169 unequivocal nevi and melanomas as well as 27 cases with ambiguous pathology, for which we had long-term follow-up data. An algorithm-using signal counts from a combination of 4 probes targeting chromosome 6p25, 6 centromere, 6q23, and 11q13 provided the highest diagnostic discrimination. This algorithm correctly classified melanoma with 86.7% sensitivity and 95.4% specificity in the validation cohort. The test also correctly identified as melanoma all 6 of 6 cases with ambiguous pathology that later metastasized. There was a significant difference in the metastasis free survival between test-positive and negative cases with ambiguous pathology (P=0.003). Sufficient chromosomal alterations are present in melanoma that a limited panel of FISH probes can distinguish most melanomas from most nevi, providing useful diagnostic information in cases that cannot be classified reliably by current methods. As a diagnostic aid to traditional histologic evaluation, this assay can have significant clinical impact and improve classification of melanocytic neoplasms with conflicting morphologic criteria.

The diagnosis of certain melanocytic proliferations remains one of the most challenging areas in pathology. In recent times, fluorescence in situ hybridization (FISH) has emerged as a promising diagnostic aid to conventional microscopy. We previously showed that a 4-probe FISH assay targeting 6p25 (RREB1), 6q23 (MYB), Cep6 (centromere 6), and 11q13 (CCND1) could discriminate between histologically unequivocal melanomas and benign nevi with a sensitivity of 86.7% and specificity of 95.4%. However, the sensitivity of the assay is approximately 70% in melanomas with spitzoid morphology. Furthermore, differentiating true gains from tetraploidy may cause difficulties in interpretation by inexperienced examiners. Here we refine the current probe set to better target spitzoid melanomas and more easily distinguish cells with imbalanced copy number aberrations from tetraploid cells. Using FISH data from 3 training sets of 322 tumors, including 152 melanomas and 170 nevi, we identified 9p21, 6p25, 11q13, and 8q24 as a probe set with improved discriminatory power in differentiating melanomas from nevi. In a validation set of 51 melanomas and 51 nevi this probe set had a sensitivity of 94% and specificity of 98%, compared with the original probe set that had a sensitivity of 75% and specificity of 96% in the same validation cohort. We propose that by incorporating 9p21 into the 4-probe FISH assay, with a new diagnostic algorithm, this new probe set would have improved discriminatory power in melanocytic neoplasms and improved sensitivity for detecting spitzoid melanomas, as demonstrated by our previous studies.

The frequency of ROS1 rearrangements in clinically selected patients is higher than that reported for unselected patients, suggesting that ROS1 rearrangement is a druggable target in East-Asian never smokers with lung adenocarcinoma. Given the different treatment outcomes to conventional therapies and availability of ROS1 inhibitors, identification of ROS1 rearrangement can lead to successful treatment in ROS1-rearranged lung adenocarcinomas.

Relating specific genetic alterations to prognosis may help improve prognostication in melanoma, may identify key oncogenic drivers in cancer, and may assist in developing targeted therapies. Characteristic genetic alterations in melanoma include chromosomal copy number aberrations. We evaluated 97 melanomas (55 metastasizing and 42 nonmetastasizing) after a minimum 5-year follow-up in a case-control study using fluorescence in situ hybridization, targeting commonly altered chromosomal loci in melanoma. Eight probes arranged in two panels were used, and 11 parameters were evaluated. Parameters showing a statistically significant difference between the metastasizing and nonmetastasizing groups were evaluated with multivariate logistic regression analysis to compare their prognostic potential with other traditional prognostic markers used by the American Joint Committee on Cancer. Four of 11 parameters evaluated, including CCND1 (alias Bcl-1) gain, CCND1 r-gain, MYC (alias c-myc) gain, and MYC r-gain, had a statistically significant difference in the metastasizing versus nonmetastasizing group. All four parameters maintained statistical significance when evaluated in separate multivariate logistic regression analyses that included the seven currently used American Joint Commission on Cancer prognosticators in melanoma. In multivariate analyses, these four parameters were second only to ulceration in their prognostic potential. Copy number changes at 11q13 and 8q34 harboring CCND1 and MYC, respectively, are highly associated with prognosis. Fluorescence in situ hybridization targeting these loci may be a useful standardized prognostic marker in melanoma skin cancer.

To investigate the frequency and the prognostic impact of fibroblast growth factor receptor 1 (FGFR1) gene amplification in 526 curatively resected esophageal squamous cell carcinoma (ESCC). Using fluorescent in situ hybridization, high amplification was defined by an FGFR1/centromer 8 ratio is ≥ 2.0, or average number of FGFR1 signals/tumor cell nucleus ≥ 6.0, or percentage of tumor cells containing ≥ 15 FGFR1 signals or large cluster in ≥ 10%. Low amplification was defined by ≥ 5 FGFR1 signals in ≥ 50%. FGFR2 and FGFR3 mutations were assessed by direct sequencing in 388 cases and no mutation was detected. High and low amplification were detected in 8.6% and 1.1%, respectively. High FGFR1 amplification had significantly shorter disease-free survival (34.0 vs 158.5 months P=0.019) and overall survival (52.2 vs not reached P=0.022) than low/no amplification group. After adjusting for sex, smoking, stage, histology, and adjuvant treatment, high FGFR1 amplification had a greater risk of recurrence (adjusted hazard ratio [AHR], 1.6; P=0.029) and death (AHR, 1.53; P=0.050). High amplification was significantly higher in current smokers than former and never-smokers (Ptrend<0.001) and increased proportional to smoking dosage. High FGFR1 amplification is a frequent oncogenic alteration and an independent poor prognostic factor in resected ESCC.