Genetic Profiling Uncovers Genome-Wide Loss of Heterozygosity and Provides Insight into Mechanisms of Sarcomatoid Transformation in Chromophobe Renal Cell Carcinoma.

Sarcomatoid transformation occurs in ∼8% of chromophobe renal cell carcinoma (chRCC) and is associated with aggressive clinical behavior. In recent years, several studies have identified genomic, transcriptomic, and epigenomic correlates of aggressive behavior in chRCC; however, the molecular mechanisms associated with sarcomatoid transformation remain incompletely understood. In this study, we analyzed paired conventional and sarcomatoid histologic components of individual chRCC to elucidate the genomic alterations that underlie sarcomatoid transformation in this tumor type. Massively parallel sequencing was performed on paired (conventional and sarcomatoid) components from 8 chRCCs. All cases harbored TP53 variants (87.5% showing TP53 variants in both components and 12.5% only in the sarcomatoid component). Intratumor comparisons revealed that TP53 variants were concordant in 71% and discordant in 29% of cases. Additional recurrent single-nucleotide variants were found in RB1 (37.5% of cases) and PTEN (25% of cases), with the remaining single-nucleotide variants detected in these tumors (PBRM1, NF1, and ASXL1) being nonrecurrent. Copy number variant analysis showed the characteristic pattern of chromosomal losses associated with chRCC (1, 2, 6, 10, 13, 17, and 21) in the conventional histologic components only. Interestingly, the sarcomatoid components of these tumors demonstrated widespread loss of heterozygosity but lacked the above chromosomal losses, likely as a consequence of whole-genome duplication/imbalanced chromosomal duplication events. Overall, the findings suggest that TP53 variants followed by whole-genome duplication/imbalanced chromosomal duplication events underlie sarcomatoid transformation in chRCC.

Genomic analysis of primary epithelial neoplasms of the seminal vesicle identifies a subset of mucinous cystic tumours driven by KRAS mutations.

BACKGROUND: Carcinomas of the seminal vesicle are exceedingly rare, with a limited number of cases described in the literature. Reported cases span a relatively wide morphological spectrum, and their genomic features remain unexplored. DESIGN: In this study, we interrogated five primary epithelial neoplasms of the seminal vesicle using a targeted DNA sequencing platform (OncoPanel, 447 genes). RESULTS: The tumours included one adenocarcinoma with intestinal phenotype presenting after external beam radiation (for prostatic adenocarcinoma), one carcinoma with Müllerian-type clear cell phenotype, two mucinous tumours resembling low-grade mucinous neoplasms of the appendix (LAMN) and one mucinous cystadenoma. The post-radiation mucinous adenocarcinoma had genomic findings consistent with bi-allelic inactivation of TP53, as well as multiple copy-number changes with regional and chromosomal arm-level copy-number losses. The Müllerian-type clear cell carcinoma exhibited a complex copy-number profile with numerous regional and arm-level copy-number changes, as well as focal amplification events, including copy-number gain of 8q and amplification of a region within 20q13. Both low-grade mucinous tumours resembling LAMN harboured hot-spot gain-of-function KRAS variants (p.G12V and p.G13D) as the only genomic alteration. No genomic alterations were detected inthe lesion diagnosed as mucinous cystadenoma. CONCLUSION: Our results suggest that primary low-grade mucinous neoplasms of the seminal vesicle may represent a distinct entity equivalent to appendiceal counterparts, driven by gain-of-function variants of RAS GTPases. The remaining tumours showed genomic features that closely resembled those of neoplasms with comparable phenotypes and/or biological characteristics arising in other sites, suggesting that they could be managed similarly, with special considerations related to their anatomical location.

STK11 (LKB1) immunohistochemistry is a sensitive and specific marker for STK11 adnexal tumours.

AIMS: STK11 adnexal tumour is a rare, recently described malignant neoplasm that is associated with Peutz-Jeghers syndrome. [Correction added on 3 October 2024, after first online publication: 'ST11' in preceding sentence has been corrected to 'STK11' in this version.] It predominantly originates from the para-adnexal soft tissues and often shows secondary involvement of the fallopian tube and ovary. STK11 adnexal tumours have a broad differential diagnosis due to their variable morphology and non-specific immunoprofile, and diagnostic confirmation currently requires sequencing to identify an STK11 mutation. We investigate the diagnostic utility of STK11 (LKB1) immunohistochemistry (IHC) in a cohort of STK11 adnexal tumours and morphological mimics. METHODS AND RESULTS: IHC for STK11 was performed on 122 tumours, including 17 STK11 adnexal tumours and 105 morphological mimics (10 female adnexal tumours of Wolffian origin, 22 adult granulosa cell tumours, 10 juvenile granulosa cell tumours, four Sertoli-Leydig cell tumours, two Leydig cell tumours, one Sertoli cell tumour, one steroid cell tumour, four extra-ovarian sex cord-stromal tumours, 16 ovarian endometrioid carcinomas, eight tubo-ovarian high-grade serous carcinomas, five ovarian mesonephric-like adenocarcinomas, 14 ovarian carcinosarcomas, five peritoneal malignant mesotheliomas, two pelvic plexiform leiomyomata and one ovarian solid pseudopapillary tumour). All STK11 adnexal tumours showed complete loss of cytoplasmic staining for STK11. All other tumour types showed retained cytoplasmic staining, except for one endometrioid carcinoma with mucinous differentiation which showed complete loss of STK11 expression and a high-grade serous carcinoma with subclonal loss. CONCLUSIONS: STK11 is a highly sensitive and specific immunohistochemical marker for distinguishing STK11 adnexal tumour from its histological mimics, and can obviate the need for confirmatory molecular studies in the appropriate morphological context.

Reliability and Management Outcomes Following a Percutaneous Biopsy Diagnosis of Oncocytoma: A 15-year Retrospective Analysis.

Background There is uncertainty in the management of renal masses diagnosed as oncocytomas with image-guided percutaneous biopsy. Purpose To assess the reliability of a diagnosis of oncocytoma based on image-guided percutaneous renal mass biopsy and evaluate patient outcomes following different management strategies. Materials and Methods In this retrospective study, image-guided percutaneous biopsy pathology reports from April 2004 to April 2019 were searched for keywords "oncocytoma" and "oncocytic neoplasm" and compared with surgical pathology or repeat biopsy results. Patients with at least 12 months of clinical follow-up and known cause of death were grouped according to management strategies, and disease-specific survival and metastatic renal cell carcinoma (RCC)-free survival were compared. Mass growth rates were calculated with use of a normal linear mixed model. Results The database yielded 160 biopsy reports of 149 renal masses in 139 patients; 149 masses were categorized as oncocytoma (n = 107), likely oncocytoma (n = 12), oncocytic neoplasm (n = 28), and indeterminate with oncocytoma in differential (n = 2). Biopsied masses categorized as oncocytoma or likely oncocytoma were oncocytomas in 16 of 17 masses (94%) based on surgical pathology or repeat biopsy; four of eight masses (50%) categorized as oncocytic neoplasms were low-grade RCCs. Outcome analysis included 121 patients (mean age ± SD, 68 years ± 9.1; 82 men); 80 patients initially underwent active surveillance (11 were later treated), 33 underwent ablation, and eight underwent surgery. Disease-specific survival and metastatic-free survival were 100% after each management strategy (median follow-up, 86.6 months; range, 14.2-207.9 months). Mass growth rate (mean, 1.7 mm per year) showed no evidence of a significant difference among biopsy result categories (P = .37) or initial (P = .84) or final management strategies (P = .11). Conclusion Image-guided percutaneous biopsy diagnosis of renal oncocytoma was reliable. Although some masses diagnosed as oncocytic neoplasms were low-grade renal cell carcinomas (RCCs) at final diagnosis, no patients died of RCC, including those managed with active surveillance. © RSNA, 2023 See also the editorial by Lockhart in this issue.

Integrating Molecular Sequencing Into the Pathological Diagnosis of Clinically Suspected Non-Small Cell Lung Carcinomas.

Cancer panel sequencing has been rapidly adopted into clinical practice for the detection of actionable genetic variants in advanced non-small cell lung carcinomas. Unexpected molecular results may lead to reexamination of the clinical diagnosis; however, this scenario has not been systematically evaluated. We reviewed cancer panel next-generation sequencing results from 1007 consecutive patients performed for the clinical indication of non-small cell lung carcinoma along with the corresponding clinical history and anatomical pathology findings. The final integrative diagnosis was a cancer of extrapulmonary origin in 12 of 1007 patients (1.2%). Molecular evidence supporting the final diagnoses included the detection of an UV radiation-associated mutational signature (n = 6), gene fusions (n = 2), and mutations (n = 4). The integrative diagnoses included undifferentiated melanoma, cutaneous squamous cell, and basal cell carcinomas, thyroid carcinoma, urothelial carcinoma, hepatocellular carcinoma, pancreatic adenocarcinoma, intrahepatic cholangiocarcinoma, and synovial sarcoma. A small but nonnegligible proportion of clinically suspected non-small cell lung carcinomas had a final diagnosis of cancer of extrapulmonary origin after clinical next-generation sequencing. The integration of clinical, microscopic, and molecular evidence can aid diagnosis and guide personalized oncology care.

Large cell calcifying Sertoli cell tumour: molecular and immunohistochemical assessment of a series comprising non-metastasising and metastasising neoplasms.

Large cell calcifying Sertoli cell tumour (LCCSCT) is a type of testicular sex cord-stromal tumour that may occur sporadically or in the context of Carney complex and other genetic syndromes. A subset is clinically malignant, and the molecular mechanisms that drive such aggressive behaviour remain unknown. METHODS AND RESULTS: We analysed 21 samples from 20 patients with LCCSCT (12 non-metastasising and eight metastasising) using PRKAR1A immunohistochemistry (IHC) and next-generation sequencing. All tumours except two (cases 17 and 20, both metastasising) demonstrated loss of PRKAR1A expression. Among 11 cases with interpretable sequencing results, all harboured pathogenic single nucleotide variants of PRKAR1A. Evidence of loss of heterozygosity (LOH) of PRKAR1A was present in all tumours with interpretable zygosity data, but the mechanisms of LOH were different for non-metastasising and metastasising tumours. Non-metastasising tumours demonstrated only copy-neutral LOH, while metastasising tumours demonstrated a spectrum of mechanisms of LOH, including copy-loss LOH, two concurrent mutations or copy-neutral LOH. Relevant molecular findings in non-metastasising LCCSCT were limited to PRKAR1A variants. In contrast, all metastasising LCCSCTs with interpretable data harboured additional pathogenic variants, including (but not restricted to) BRCA2 mutations with evidence of LOH and bi-allelic CDKN2A/B deletions. Three patients harboured PRKAR1A variants of inferred germline origin, including one with Carney complex and two without known syndromic features. CONCLUSIONS: This study further confirms that PRKAR1A IHC is a useful diagnostic tool for both non-metastasising and metastasising tumours and suggests that molecular analyses can be helpful to identify non-metastasising tumours with malignant potential in selected patients. Importantly, these results highlight that germline assessment could be beneficial for all patients presenting with LCCSCT.

Myoid gonadal stromal tumours are characterised by recurrent chromosome-level copy number gains: molecular assessment of a multi-institutional series.

Myoid gonadal stromal tumours (MGST) represent a rare type of testicular sex cord-stromal tumour that has recently been recognised as a distinct entity by the World Health Organization (WHO) classification of genitourinary tumours. MGSTs affect adult men and have been reported to behave in an indolent fashion. Histologically, MGSTs are pure spindle cell neoplasms that coexpress SMA and S100 protein. Given that the molecular features of these neoplasms remain largely undescribed, we evaluated a multi-institutional series of MGSTs using DNA and RNA sequencing. This study included 12 tumours from 12 patients aged 28 to 57 years. Tumour sizes ranged from 0.6 to 4.3 cm. Aggressive histologic features, such as vascular invasion, necrosis, invasive growth, and atypical mitoses were invariably absent. Mitotic activity was low, with a median of less than 1 mitosis per 10 high power fields (HPF; maximum: 3 mitoses per 10 HPF). Molecular analyses did not identify recurrent mutations or gene fusions. All cases with interpretable copy number variant data (9/10 cases sequenced successfully) demonstrated a consistent pattern of chromosome arm-level and whole-chromosome-level copy number gains indicative of ploidy shifts, with recurrent gains involving chromosomes 3, 6, 7, 8, 9, 11, 12, 14q, 15q, 17, 18q, 20, and 21q. Similar findings have also been recognised in pure spindle cell and spindle-cell predominant sex cord-stromal tumours without S100 protein expression. MGSTs are characterised by ploidy shifts and may be part of a larger spectrum of spindle cell-predominant sex cord-stromal tumours, including cases without S100 protein expression.

Deriving tumor purity from cancer next generation sequencing data: applications for quantitative ERBB2 (HER2) copy number analysis and germline inference of BRCA1 and BRCA2 mutations.

Tumor purity, or the relative contribution of tumor cells out of all cells in a pathological specimen, influences mutation identification and clinical interpretation of cancer panel next generation sequencing results. Here, we describe a method of calculating tumor purity using pathologist-guided copy number analysis from sequencing data. Molecular calculation of tumor purity showed strong linear correlation with purity derived from driver KRAS or BRAF variant allele fractions in colorectal cancers (R2 = 0.79) compared to histological estimation in the same set of colorectal cancers (R2 = 0.01) and in a broader dataset of cancers with various diagnoses (R2 = 0.35). We used calculated tumor purity to quantitate ERBB2 copy number in breast carcinomas with equivocal immunohistochemical staining and demonstrated strong correlation with fluorescence in situ hybridization (R2 = 0.88). Finally, we used calculated tumor purity to infer the germline status of variants in breast and ovarian carcinomas with concurrent germline testing. Tumor-only next generation sequencing correctly predicted the somatic versus germline nature of 26 of 26 (100%) pathogenic TP53, BRCA1 and BRCA2 variants. In this article, we describe a framework for calculating tumor purity from cancer next generation sequencing data. Accurate tumor purity assessment can be assimilated into interpretation pipelines to derive clinically useful information from cancer genomic panels.

Molecular assessment of testicular adult granulosa cell tumor demonstrates significant differences when compared to ovarian counterparts.

Testicular adult granulosa cell tumor (AGCT) is a rare type of sex-cord stromal tumor that affects patients of a wide age range and has the potential for late metastasis. In the testis, the diagnosis of AGCTs often requires the exclusion of other more common types of sex-cord stromal tumors. Immunohistochemistry is of limited utility, being used mostly to confirm sex-cord lineage and to exclude other entities when morphology is not typical. Unlike ovarian AGCTs, which are molecularly homogeneous and harbor a specific activating FOXL2 mutation (c.7558C > T p.C134W) in >90% of cases, the molecular characteristics of testicular AGCTs remain largely unknown. In the current study, we analyzed 13 testicular AGCTs diagnosed at multiple institutions using massively parallel DNA sequencing to evaluate single nucleotide variants, copy number alterations, and structural variants. In all, 10/13 cases were sequenced successfully. Notably, the FOXL2 c.7558C > T (p.C134W) mutation was identified in only a single case (1/10, 10%). The remaining cases were molecularly heterogeneous, with largely nonrecurrent genetic variants. Putative driver events in individual cases included a well-characterized gain-of-function NRAS mutation, as well as inactivation of ATM and TP53, among others. The only highly recurrent finding was single copy loss of 22q (7/10 cases, 70%). Comparatively, the frequencies of FOXL2 c.7558C > T (p.C134W) and 22q loss in 12 metastatic ovarian AGCTs identified in our database were 92% (11/12) and 42% (5/12), respectively. The results of the present study suggest that testicular AGCTs are different from their ovarian counterparts in that they appear to be molecularly heterogeneous and only rarely harbor FOXL2 mutations.

Clinicopathologic and molecular spectrum of testicular sex cord-stromal tumors not amenable to specific histopathologic subclassification.

A subset of testicular sex cord-stromal tumors (SCST), which includes neoplasms with mixed histology, cannot be classified into a specific histologic subtype. This study evaluated the clinicopathologic, immunophenotypic and molecular features of 26 SCST not amenable to specific classification by expert uropathologists. Median age at diagnosis was 43 years and median tumor size was 2.4 cm. Follow-up information was available for 18 (69%) patients, with evidence of an aggressive clinical course in 6 patients (4 alive with disease, 2 dead of disease 3 months and 6 months after orchiectomy). Microscopically, SCST not amenable to specific classification demonstrated monophasic epithelioid (9/26, 35%), monophasic spindle cell (5/26, 19%), and biphasic or mixed histology (12/26, 46%). One or more aggressive histopathologic features were seen in 11 cases. DNA sequencing was successful in 22 tumors. Pathogenic CTNNB1 and APC alterations were seen in 7 (33%) and 2 (10%) cases, respectively, with additional variants (e.g., CDKN2A, RB1, TP53, BRCA2) being identified in individual cases. Combined evaluation of morphology, sequencing data and beta-catenin immunohistochemistry resulted in reclassification of 6 (23%) tumors as Sertoli cell tumor, not otherwise specified. This was supported by comparing the methylation profiles of a subset of these tumors and those of typical Sertoli cell tumors. Additionally, a subset of 5 neoplasms (19%) with spindle cell or biphasic histology and SMA expression was characterized by hyperdiploid genomes with recurrent chromosomal gains and absence of driver mutations, possibly representing a distinct tumor type. The SCST that remained not amenable to specific histologic classification (15/26, 58%) were enriched for aggressive histologic features and malignant clinical behavior. In conclusion, this study demonstrated that a subset of testicular SCST that were originally not amenable to specific classification could be reclassified by combined evaluation of morphology, immunohistochemistry and molecular data.

Low-dose rapamycin extends lifespan in a mouse model of mtDNA depletion syndrome.

Mitochondrial disorders affecting oxidative phosphorylation (OxPhos) are caused by mutations in both the nuclear and mitochondrial genomes. One promising candidate for treatment is the drug rapamycin, which has been shown to extend lifespan in multiple animal models, and which was previously shown to ameliorate mitochondrial disease in a knock-out mouse model lacking a nuclear-encoded gene specifying an OxPhos structural subunit (Ndufs4). In that model, relatively high-dose intraperitoneal rapamycin extended lifespan and improved markers of neurological disease, via an unknown mechanism. Here, we administered low-dose oral rapamycin to a knock-in (KI) mouse model of authentic mtDNA disease, specifically, progressive mtDNA depletion syndrome, resulting from a mutation in the mitochondrial nucleotide salvage enzyme thymidine kinase 2 (TK2). Importantly, low-dose oral rapamycin was sufficient to extend Tk2KI/KI mouse lifespan significantly, and did so in the absence of detectable improvements in mitochondrial dysfunction. We found no evidence that rapamycin increased survival by acting through canonical pathways, including mitochondrial autophagy. However, transcriptomics and metabolomics analyses uncovered systemic metabolic changes pointing to a potential 'rapamycin metabolic signature.' These changes also implied that rapamycin may have enabled the Tk2KI/KI mice to utilize alternative energy reserves, and possibly triggered indirect signaling events that modified mortality through developmental reprogramming. From a therapeutic standpoint, our results support the possibility that low-dose rapamycin, while not targeting the underlying mtDNA defect, could represent a crucial therapy for the treatment of mtDNA-driven, and some nuclear DNA-driven, mitochondrial diseases.

Low-grade oncocytic tumor: a review of radiologic and clinical features.

PURPOSE: The 2022 World Health Organization classification of renal neoplasia expanded the spectrum of oncocytic neoplasms to encompass newly established and emerging entities; one of the latter is the low-grade oncocytic tumor (LOT). This study reports the radiologic appearance and clinical behavior of LOT. METHODS: In this IRB-approved, HIPPA-compliant retrospective study, our institution's pathology database was searched for low-grade oncocytic tumors or neoplasms. Patient age, gender, and comorbidities were obtained from a review of electronic medical records, and imaging characteristics of the tumors were assessed through an imaging platform. RESULTS: The pathology database search yielded 14 tumors in 14 patients. Four patients were excluded, as radiologic images were not available in three, and one did not fulfill diagnostic criteria after pathology re-review. The resulting cohort consisted of 10 tumors (median diameter 2.3 cm, range 0.7-5.1) in 10 patients (median age 68 years, range 53-91, six women). All tumors presented as a solitary, well-circumscribed, mass with solid components. All enhanced as much or almost as much as adjacent renal parenchyma; all but one enhanced heterogeneously. None had lymphadenopathy, venous invasion, or metastatic disease at presentation or at clinical follow-up (median, 22.2 months, range 3.4-71.6). Among five tumors undergoing active surveillance, mean increase in size was 0.4 cm/year at imaging follow-up (median 16.7 months, range 8.9-25.4). CONCLUSION: LOT, a recently described pathologic entity in the kidney, can be considered in the differential diagnosis of an avidly and typically heterogeneously enhancing solid renal mass in an adult patient.

Epidermodysplasia verruciformis-associated eccrine neoplasm: a rare entity with distinctive clinical and histopathologic features.

Most tumors are caused by inherited or acquired genetic changes. However, a subset of tumors is driven by viral infection including Kaposi sarcoma, nasopharyngeal carcinoma, and others. Human papillomavirus (HPV) is an especially common cause of epithelial cancers and hyperplasias. Epidermodysplasia verruciformis (EDV) is a rare type of HPV infection with characteristic histopathologic features and a unique spectrum of HPV subtypes. We report here a distinctive form of EDV-associated eccrine neoplasia. Seven tumors from two patients were analyzed and show highly uniform features including multiple clustered clinical lesions, multifocal epidermal origin, eccrine differentiation with close association with the acrosyringium, an anastomosing growth pattern, and a bland monotonous poroid-to-basaloid cytomorphology. Clinical follow-up for one patient has been benign to date. These tumors show strong similarity to two previously reported cases, suggesting that this type of EDV-associated eccrine neoplasia may represent a rare but reproducible form of skin adnexal tumor with distinctive clinicopathologic features.

Concordance of MTOR Pathway Mutations and the Diagnosis of Renal Low-Grade Oncocytic Tumor (LOT).

The differential diagnosis for oncocytic renal tumors spans the spectrum from benign entities to more aggressive renal cell carcinomas (RCC). Recent work has characterized a provisional renal oncocytic neoplasm, namely the low-grade oncocytic tumor (LOT), which demonstrates overlapping morphologic features with oncocytoma and chromophobe RCC, but also has a unique immunoprofile (ie, diffusely positive for KRT7, negative for KIT) and a high rate (80% to 100%) of mTOR pathway gene alterations. Given the diagnostic overlap among oncocytic tumors, we looked for concordance between mTOR pathway mutations and LOT. Thirty low-grade renal oncocytic neoplasms underwent histologic review and immunohistochemistry for KRT7 and KIT. Tumors were classified as "determinate" (eg, LOT) for tumors with solid, nested or vaguely tubular growth and diffuse KRT7 staining and negative KIT, or "indeterminate" if the morphology and/or immunostains did not fully support a definitive LOT diagnosis. Next-generation sequencing was performed without any knowledge of the diagnoses, and identified mTOR pathway mutations in 80% (12/15) of the determinate tumors, compared with 7% (1/15) in the indeterminate group. One determinate tumor was reclassified as papillary RCC (MTOR mutation negative) and 6 indeterminate tumors were confirmed to be oncocytoma (N = 4), clear cell RCC or papillary RCC with reverse polarity, respectively. Overall, integration of morphology, immunohistochemistry, and molecular data enabled a final definitive diagnosis for 70% of tumors (21 of the total 30), with a high concordance (93%) for LOT specifically in the determinate group; the remaining 9 tumors (30%) were classified as renal oncocytic neoplasm, not otherwise specified.

FGFR Alterations in Thyroid Carcinoma: A Novel Class of Primary Drivers with Significant Therapeutic Implications and Secondary Molecular Events Potentially Mediating Resistance in Thyroid Malignancy.

Background: Diagnostic classification of thyroid malignancy is primarily accomplished through examination of histomorphological features and may be substantiated and clarified by molecular data. Individual molecular drivers show relatively robust and specific associations with histological subtypes of thyroid malignancy, including BRAF sequence variants and kinase gene fusions in papillary thyroid carcinoma, predominantly RAS variants in follicular-patterned neoplasia, and additional "late" mutations affecting TERT promoter, TP53, and the PI3K/AKT/PTEN pathway in high-grade malignancies. Given the oncogenic role of FGFR, particularly FGFR1-3, the goal of this study was to explore the role of FGFR in thyroid carcinoma biology. Methods: We completed a multicenter retrospective observational study for thyroid carcinomas with pathogenic alterations in the FGFR gene family. We performed this study by querying the molecular data accumulated for thyroid carcinomas from each center. Results: Overall, 5030 sequenced thyroid malignancies were reviewed, yielding 17 tumors with FGFR alterations, including 11 where FGFR was the primary molecular driver and 6 where FGFR was a secondary pathogenic alteration, with a subset for which there was available clinical follow-up data. Of the 11 carcinomas with an FGFR driver, 9 were gene fusions involving FGFR2:VCL (4 tumors), TG::FGFR1 (3 tumors), FGFR2::CIT, and FGFR2::SHTN1, and the remaining 2 were driven by FGFR1 amplification. In the 6 tumors where a canonical driver of thyroid neoplasia was present (5 cases) or no clear primary driver was detected (1 case), sequencing detected secondary FGFR2 p.W290C, p.Y375C, and p.N549K, as well as FGFR1 p.N546K in the respective tyrosine kinase domains, some at subclonal variant allele frequencies. Conclusions: This study presents the first description of a collection of thyroid carcinomas grouped by primary driver alterations in FGFR, as well as a cohort of thyroid tumors with secondary alterations that potentially lead to tumor progression or resistance to targeted therapy. Given the availability of small molecular inhibitors targeting oncogenic FGFR, this study emphasizes the significant implications for patients from identification of FGFR alterations as they are currently under-recognized in the literature and, most importantly, have potential novel treatment options.

A pleiotropic drug resistance transporter in Nicotiana tabacum is involved in defense against the herbivore Manduca sexta.

Pleiotropic drug resistance (PDR) transporters are a group of membrane proteins belonging to the ABCG sub-family of ATP binding cassette (ABC) transporters. There is clear evidence for the involvement of plant ABC transporters in resistance to fungal and bacterial pathogens, but not in the biotic stress response to insect or herbivore attack. Here, we describe a PDR transporter, ABCG5/PDR5, from Nicotiana tabacum. GFP fusion and subcellular fractionation studies revealed that ABCG5/PDR5 is localized to the plasma membrane. Staining of transgenic plants expressing the GUS reporter gene under the control of the ABCG5/PDR5 transcription promoter and immunoblotting of wild-type plants showed that, under standard growth conditions, ABCG5/PDR5 is highly expressed in roots, stems and flowers, but is only expressed at marginal levels in leaves. Interestingly, ABCG5/PDR5 expression is induced in leaves by methyl jasmonate, wounding, pathogen infiltration, or herbivory by Manduca sexta. To address the physiological role of ABCG5/PDR5, N. tabacum plants silenced for the expression of ABCG5/PDR5 were obtained. No phenotypic modification was observed under standard conditions. However, a small increase in susceptibility to the fungus Fusarium oxysporum was observed. A stronger effect was observed in relation to herbivory: silenced plants allowed better growth and faster development of M. sexta larvae than wild-type plants, indicating an involvement of this PDR transporter in resistance to M. sexta herbivory.

An update on diagnostic tissue-based biomarkers in testicular tumors.

Testicular cancer is rare overall but comprises the most common solid malignancy diagnosed in young men aged ∼20-40 years. Most testicular neoplasms generally fall into 2 broad categories: germ cell tumors (GCTs; ∼95%) and sex cord-stromal tumors (SCSTs ∼5%). Given the relative rarity of these tumors, diagnostic biomarkers are highly relevant for their diagnosis. Over the past several decades, diagnostic biomarkers have improved dramatically through targeted immunohistochemical and molecular characterization. Despite these recent advances, most markers are not perfectly sensitive or entirely specific. Therefore, they need to be used in combination and interpreted in context. In this review, we summarize tissue-based biomarkers relevant to the pathologist, with a focus on practical diagnostic issues that relate to testicular GCT and SCST.

Germline APC Alterations May Predispose to Testicular Sex Cord-Stromal Tumors.

Sertoli cell tumor is a type of testicular sex cord-stromal tumor (TSCST) typically driven by gain-of-function CTNNB1 variants. Recently, molecular studies have identified TSCSTs (including Sertoli cell tumors) with loss-of-function APC variants, raising the possibility that germline APC alterations may predispose to TSCSTs. In this study, we evaluated 4 TSCSTs from 4 individual patients, including 3 APC -mutant neoplasms identified in prior studies (1 in a patient with familial adenomatous polyposis [FAP] and 2 in patients with unknown syndromic status) and 1 tumor of unknown mutational status diagnosed in a patient with known FAP. Three neoplasms were typical Sertoli cell tumors, and 1 was a malignant unclassified TSCT. All neoplasms exhibited diffuse nuclear beta-catenin expression. Non-neoplastic tissue could be obtained for DNA sequencing in the 3 Sertoli cell tumors. Comparative assessment of non-neoplastic and lesional tissue in these cases suggested that germline APC variants with subsequent inactivation of the gene (loss of heterozygosity) were the likely oncogenic driver of these Sertoli cell tumors. In the malignant unclassified TSCSTs, APC inactivation was also interpreted as the most likely driver event, and the germline origin of the variant was inferred using a recently published method. The results of this study suggest that pathogenic germline APC alterations (eg, FAP and variants thereof) may predispose to TSCSTs.

Clinical Problem Solving: A 56-Year-Old Woman With Right Facial Weakness, Numbness, and Diplopia.

We discuss a case of a middle-aged woman with a history of urothelial carcinoma, who presented with subacute progression of double vision, right facial numbness, and back pain. Her MRI brain and spine demonstrated multifocal enhancing lesions at the skull base and throughout the spine. Since her initial serum and cerebrospinal fluid (CSF) studies were unrevealing, she underwent a biopsy of her skull base lesion, which was initially concerning for infection. However, her symptoms worsened despite appropriate antibiotic therapy, necessitating a spinal root biopsy that was consistent with leptomeningeal urothelial carcinoma. Her case illustrates the challenge of accurately diagnosing isolated leptomeningeal metastatic disease, which can be difficult to diagnose from CSF analysis and often requires multiple lumbar punctures to improve sensitivity. While genitourinary cancers rarely metastasize to the CNS, clinicians should retain high suspicion for neoplastic etiologies of leptomeningeal disease in patients with a history of cancer and new neurologic deficits.

Measuring Digital Pathology Throughput and Tissue Dropouts.

BACKGROUND: Digital pathology operations that precede viewing by a pathologist have a substantial impact on costs and fidelity of the digital image. Scan time and file size determine throughput and storage costs, whereas tissue omission during digital capture ("dropouts") compromises downstream interpretation. We compared how these variables differ across scanners. METHODS: A 212 slide set randomly selected from a gynecologic-gestational pathology practice was used to benchmark scan time, file size, and image completeness. Workflows included the Hamamatsu S210 scanner (operated under default and optimized profiles) and the Leica GT450. Digital tissue dropouts were detected by the aligned overlay of macroscopic glass slide camera images (reference) with images created by the slide scanners whole slide images. RESULTS: File size and scan time were highly correlated within each platform. Differences in GT450, default S210, and optimized S210 performance were seen in average file size (1.4 vs. 2.5 vs. 3.4 GB) and scan time (93 vs. 376 vs. 721 s). Dropouts were seen in 29.5% (186/631) of successful scans overall: from a low of 13.7% (29/212) for the optimized S210 profile, followed by 34.6% (73/211) for the GT450 and 40.4% (84/208) for the default profile S210 profile. Small dislodged fragments, "shards," were dropped in 22.2% (140/631) of slides, followed by tissue marginalized at the glass slide edges, 6.2% (39/631). "Unique dropouts," those for which no equivalent appeared elsewhere in the scan, occurred in only three slides. Of these, 67% (2/3) were "floaters" or contaminants from other cases. CONCLUSIONS: Scanning speed and resultant file size vary greatly by scanner type, scanner operation settings, and clinical specimen mix (tissue type, tissue area). Digital image fidelity as measured by tissue dropout frequency and dropout type also varies according to the tissue type and scanner. Dropped tissues very rarely (1/631) represent actual specimen tissues that are not represented elsewhere in the scan, so in most cases cannot alter the diagnosis. Digital pathology platforms vary in their output efficiency and image fidelity to the glass original and should be matched to the intended application.