Introduction: The onset of precision medicine has led to the integration of traditional morphologic tissues evaluation with biochemical and molecular data for a more appropriate pathological diagnosis. The preanalytic phase and, particularly, timing of cold ischemia are crucial to guarantee high-quality biorepositories of formalin-fixed paraffin-embedded (FFPE) tissues for patients' needs and scientific research. However, delayed fixation using the gold-standard and carcinogenic fixative neutral-buffered formalin (NBF) can be a significant limitation to diagnosis and biopathological characterization. HistoCold (patented; Bio-Optica Milano S.p.A., Milano, Italy) is a nontoxic, stable, and refrigerated preservative solution for tissue handling. This study examined HistoCold's potential role in improving the preanalytic phase of the pathological diagnostic process.Materials and Methods: Breast, lung, or colorectal cancers (20, 25, and 10 cases, respectively) that were to be surgically resected were recruited between 2019 and 2021. Once specimens were surgically removed, three residual samples for each patient were first promptly immersed into HistoCold for 24, 48, and 72 hours and then FFPE. These were compared with routine specimens regarding morphologic features (hematoxylin and eosin) and tissue antigenicity (immunohistochemical stains).Results: Good concordance regarding both the morphologic characteristics of the neoplasms and their proteins expression between the routine and HistoCold handled tissues were found. The tissue handling with the solution never affected the histopathological diagnosis.Conclusions: The use of HistoCold for samples transporting is easy, allows for improving the management of cold ischemia time, and monitoring the fixation times in NBF, resulting in good quality tissue blocks for biobanking. Moreover, it could be a candidate to eliminate formalin from operating theaters. HistoCold looks very promising for the preanalytic phase of human tissues handling in the era of precision medicine, to provide the best service to patients, and to scientific research.

Purpose Decades of quality control efforts have raised the standards of immunohistochemistry (IHC), the principle method used for biomarker testing in breast cancer; however, computational pathology and reverse transcription quantitative PCR (RT-qPCR) may also hold promise for additional substantial improvements. Methods Herein, we investigated discrepancies in the assessment of estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2) and marker of proliferation Ki67 comparing routinely obtained IHC (and FISH) data (ORI) with the results of manual (REV) and semi-automated (DIA) re-evaluation of the original IHC slides and then with RNA expression data from the same tissue block using the MammaTyper® (MT) gene expression assay. Results Correlation for ER and PR was high between ORI IHC and the other three study methods (REV, DIA and RT-qPCR). For HER2, 10 out of 96 discrepant cases can be detected between ORI and REV that involved at least one call in the equivocal category (except for one case). For Ki67, 22 (29.1%) cases were categorized differently by either REV alone (n = 17), DIA alone (n = 15) or both (n = 10) and 28 cases (29.2%) for RT-qPCR. Most of the discrepant Ki67 cases changed from low to high between the original and following assessment and belonged to the intermediate Ki67 expression range (between 9 and 30%). Conclusions Determination of the breast cancer biomarkers ER, PR, HER2 and Ki67 at the mRNA level shows high degree of correlation with IHC and compares well with correlations between original with subsequent independent manual or semi-automated IHC assessments. The use of methods with wider dynamic range and higher reproducibility such as RT-qPCR may offer more precise assessment of endocrine responsiveness, improve Ki67 standardization and help resolve HER2 cases that remain equivocal or ambiguous by IHC/FISH. In summary, our findings seem to configure RT-qPCR as a complementary method to be used in cases of either equivocal results or presenting, at the traditional determination assays, biomarkers expressions close to the cut-off values.

Analysis of breast cancer prognostic and predictive factors is still nowadays poorly accurate and standardized. The advent of multi-gene expression profiles (MGEPs) has improved the prediction of breast cancer outcome, particularly regarding early luminal breast cancers (LBCs). The availability in our Institute of EndoPredict® (EP), a last-generation prognostic gene signature assay, has prompted us to study a series of LBCs, firstly verifying its reproducibility on six routine representative cases, either presenting non-optimal preanalytical conditions or different tumor samples from the same patient; secondly, correlating EP results on 8 retrospectively recruited samples with patients’ follow-up; thirdly, applying prospectively EP on 100 routinely diagnosed cases, assessing the oncologists’ and pathologists’ attitude toward it. The complete reproducibility of EP on all the samples investigated in the first phase allowed to state that EP overcomes the detrimental effects of an inaccurate pre-analytic phase, determining the most appropriate prognostic and predictive parameters of breast cancer. The second phase confirmed EP as a fundamental tool in guiding therapeutic decision, improving the classical bio-pathological characterization and recovering 38% patients’ inadequately managed. Finally, the study disclosed how oncologists sometimes inadequately requested EP, but also how it allows a better stratification of breast cancer otherwise considered poorly aggressive and not requiring an EP test, such as G1 neoplasms or tubular histotype. In conclusion, the introduction of EP test in an Anatomic Pathology Department emerges as a useful tool in routine breast cancer diagnosis, both for the characterization of individual cases and, as a result, for more appropriate therapeutic choices.

Myoepithelial carcinoma ex pleomorphic adenoma is defined as a malignant epithelial neoplasm arising from a primary or recurrent benign pleomorphic adenoma. This type of tumor comprises 3.6% of all salivary gland tumors and 12% of malignant ones. Clinically, it most commonly presents as a firm mass in the parotid gland. The development of this neoplasm in the sinonasal and nasopharyngeal regions is extremely rare and only few cases are reported in the literature. The prognosis of myoepithelial carcinoma is variable. Marked cellular pleomorphism, high mitotic rate, and high proliferative activity correspond to a poor prognosis. In this article, the authors report the histopathological features of a clinical case of a 64-years-old patient with a large median maxillary neoplasm diagnosed as myoepithelial carcinoma/ex-pleomorphic adenoma. The tumor was resected and subjected to secondary reconstruction using a revascularized free fibula flap. The myoepithelial derivation of neoplastic cells was demonstrated by immunohistochemical positivity for S-100 protein (strong and diffuse), cytokeratin 14 (strong and diffuse), and GFAP (focal).

ALK receptor tyrosine kinase gene (ALK) rearrangements identify a subset of molecularly defined NSCLCs that display exquisite sensitivity to ALK receptor tyrosine kinase tyrosine kinase inhibitors.1Ricciuti B. De Giglio A. Mecca C. et al.Precision medicine against ALK-positive non-small cell lung cancer: beyond crizotinib.Med Oncol. 2018; 35: 72Google Scholar High-grade neuroendocrine carcinomas (HGNECs) have anecdotally been associated with ALK rearrangements, which makes the predictive role of such driver mutations in these patients still largely unknown.2Omachi N. Shimizu S. Kawaguchi T. et al.A case of large-cell neuroendocrine carcinoma harboring an EML4-ALK rearrangement with resistance to the ALK inhibitor crizotinib.J Thorac Oncol. 2014; 9: e40-e42Abstract Full Text Full Text PDF PubMed Scopus (42) Google Scholar, 3Hayashi N. Fujita A. Saikai T. et al.Large cell neuroendocrine carcinoma harboring an anaplastic lymphoma kinase (ALK) rearrangement with response to alectinib.Intern Med. 2018; 57: 713-716Crossref PubMed Scopus (29) Google Scholar Here we report what, to the best of our knowledge, is the first case of ALK-rearranged and MNNG HOS Transforming gene (MET)-mutant large cell neuroendocrine carcinoma (LCNEC) of the lung in which the patient experienced a Lazarus response to crizotinib. A 58-year-old female never-smoker presented in December 2017 with neurological symptoms. The diagnosis was advanced LCNEC (Ki67 level 70%) of the lung with multiple brain, liver, and lymph node metastases (cT2N2M1b). She received dexamethasone and whole brain radiotherapy and started taking first-line platinum-etoposide chemotherapy with extracranial disease progression. Her clinical condition rapidly worsened because of acute-onset of type I respiratory failure and liver failure, making any second-line cytotoxic treatment unfeasible. Fortunately, in light of the patient's never-smoker status, a molecular work-up for potential actionable mutations was performed and revealed the presence of ALK rearrangement on the basis of immunohistochemistry (score 2+); this rearrangement was confirmed by fluorescent in situ hybridization (Fig. 1A–D) and next-generation sequencing for fusion transcripts. Of note, next-generation sequencing also revealed coexistence of an echinoderm microtubule associated protein like 4 gene (EML4)-ALK transcript of a MET exon 14 R988C mutation (Fig. 1E). Immediately upon documentation of ALK rearrangement, the patient started taking crizotinib, 250 mg twice daily, with marked improvement of her clinical condition within a few days, tapering of oxygen therapy until suspension, and progressive normalization of her liver function within 60 days (Fig. 2A). A positron emission tomography/computed tomography scan performed 28 days after the patient had started taking crizotinib showed a dramatic response to treatment (Fig. 2B). Currently, the patient is still receiving crizotinib with no evidence of disease progression. To the best of our knowledge, this is the first report of a patient with LCNEC of the lung coharboring an ALK-rearrangement and MET exon 14 R988C mutation. Chemotherapy still represents the standard of care of HGNEC, albeit with still unsatisfying survival rates. Because of the extremely low likelihood of finding a targetable genetic alteration, patients with HGNEC are not candidates for molecular testing according to current guidelines unless they present with unusual clinical characteristics that can hide an oncogene addiction, such as never-smoker status and young age. In fact, HGNECs are widely recognized to be strictly related with a heavy smoking habit and to share molecular features, including high expression of Ki-67, Bcl-2, p21, and telomerase activity, as well as abnormal p53 level and absent Rb.4Rekhtman N. Pietanza M.C. Hellmann M.D. et al.Next-generation sequencing of pulmonary large cell neuroendocrine carcinoma reveals small cell carcinoma-like and non-small cell carcinoma-like subsets.Clin Cancer Res. 2016; 22: 3618Crossref PubMed Scopus (298) Google Scholar Moreover, in roughly 1% to 2 % of cases, oncogene-addicted NSCLCs undergo histological transformation to HGNECs whereas de novo presentation of HGNECs with actionable molecular targets is exceedingly rare.5Oser M.G. Niederst M.J. Sequist L.V. Engelman J.A. Transformation from non-small-cell lung cancer to small-cell lung cancer: molecular drivers and cells of origin.Lancet Oncol. 2015; 16: e165-e172Abstract Full Text Full Text PDF PubMed Scopus (560) Google Scholar Importantly, our patient experienced a Lazarus-like response to crizotinib that reversed a life-threatening disease progression, suggesting that oncogene addiction rather than histologic type is predictive for response in such cases. Moreover, along with ALK gene rearrangement, our patient also carried a MET exon 14 R988C mutation, the predictive role of which for in vivo sensitivity to crizotinib (which also works as a MET inhibitor) is still debated. Our findings underline the need for testing for actionable genetic alterations in those patients with HGNEC of the lung who harbor clinical characteristics suggestive of an underlying targetable genetic mutation.

The presence of interfering substances like free hemoglobin, bilirubin or lipids compromises sample quality and potentially affects laboratory analysis and test results. Recently, the use of serum indices for objectively assessing sample quality has become commonplace and many preanalytical platforms, as well as clinical chemistry and coagulation analyzers, are now equipped with this analytical feature. Nevertheless, some important drawbacks remain in this practice, mainly pertaining the measurement procedure, the approach for reporting interference data, the definition of objective thresholds of interference after which test results may be biased, and the lack of harmonized practices for describing how interference cut-offs have been identified. Therefore, this document aims to discuss these important caveats and propose some reliable solutions that may be adopted by manufacturers for increasing worldwide harmonization of serum indices.

We present the case of a middle-aged male with synchronous Clear Cell Papillary Renal Cell Carcinoma and a Capsular Leiomyoma in a kidney. The former constitutes 1% of renal neoplasms while the latter is found with a 4.2% frequency at autopsies. Analysis of both tumours focused on their histological and immunohistochemical features and similarities. Few cases of synchronous, histologically different, renal tumours have been reported and this is the first observation of a new association: a Clear Cell Papillary Renal Cell Carcinoma and a Capsular Leiomyoma.