Diagnostics (Basel, Switzerland)
Bumrungthai, S;Ekalaksananan, T;Kleebkaow, P;Pongsawatkul, K;Phatnithikul, P;Jaikan, J;Raumsuk, P;Duangjit, S;Chuenchai, D;Pientong, C;
PMID: 36980391 | DOI: 10.3390/diagnostics13061084
The current practice of determining histologic grade with a single molecular biomarker can facilitate differential diagnosis but cannot predict the risk of lesion progression. Cancer is caused by complex mechanisms, and no single biomarker can both make accurate diagnoses and predict progression risk. Modelling using multiple biomarkers can be used to derive scores for risk prediction. Mathematical models (MMs) may be capable of making predictions from biomarker data. Therefore, this study aimed to develop MM-based scores for predicting the risk of precancerous cervical lesion progression and identifying precancerous lesions in patients in northern Thailand by evaluating the expression of multiple biomarkers. The MMs (Models 1-5) were developed in the test sample set based on patient age range (five categories) and biomarker levels (cortactin, p16INK4A, and Ki-67 by immunohistochemistry [IHC], and HPV E6/E7 ribonucleic acid (RNA) by in situ hybridization [ISH]). The risk scores for the prediction of cervical lesion progression ("risk biomolecules") ranged from 2.56-2.60 in the normal and low-grade squamous intraepithelial lesion (LSIL) cases and from 3.54-3.62 in cases where precancerous lesions were predicted to progress. In Model 4, 23/86 (26.7%) normal and LSIL cases had biomolecule levels that suggested a risk of progression, while 5/86 (5.8%) cases were identified as precancerous lesions. Additionally, histologic grading with a single molecular biomarker did not identify 23 cases with risk, preventing close patient monitoring. These results suggest that biomarker level-based risk scores are useful for predicting the risk of cervical lesion progression and identifying precancerous lesion development. This multiple biomarker-based strategy may ultimately have utility for predicting cancer progression in other contexts.
Lewis, JS;Smith, MH;Wang, X;Tong, F;Mehrad, M;Lang-Kuhs, KA;
PMID: 35802245 | DOI: 10.1007/s12105-022-01467-0
HPV-associated oral cavity squamous cell carcinoma (SCC) is not well-characterized in the literature, and also has a clinical significance that is poorly understood.We gathered a cohort of oral cavity (OC) SCC with nonkeratinizing morphology, either in the invasive or in situ carcinoma (or both), tested for p16 by immunohistochemistry and high risk HPV E6/E7 mRNA by RTPCR (reference standard for transcriptionally-active high risk HPV) and gathered detailed morphologic and clinicopathologic data.Thirteen patients from two institutions were proven to be HPV-associated by combined p16 and high risk HPV mRNA positivity. All 13 patients (100%) were males, all were heavy smokers (average 57 pack/year), and most were active drinkers (9/11 or 81.8%). All 13 (100%) involved the tongue and/or floor of mouth. All had nonkeratinizing features, but maturing squamous differentiation varied widely (0-90%; mean 37.3%). Nonkeratinizing areas had high N:C ratios and larger nests, frequently with pushing borders, and minimal (or no) stromal desmoplasia. The carcinoma in situ, when present, was Bowenoid/nonkeratinizing with cells with high N:C ratios, full thickness loss of maturation, and abundant apoptosis and mitosis. HPV was type 16 in 11 patients (84.6%) and type 33 in two (15.4%). Nine patients had treatment data available. These underwent primary surgical resection with tumors ranging from 1.6 to 5.2 cm. Most had bone invasion (6/9-66.7% were T4a tumors), and most (6/9-66.7%) had extensive SCC in situ with all 6 of these patients having final margins positive for in situ carcinoma.HPV-associated OCSCC is an uncommon entity that shows certain distinct clinical and pathologic features. Recognition of these features may help pathologic diagnosis and could potentially help guide clinical management.
J Neurooncol. 2014 May 28.
Abiria SA, Williams TV, Munden AL, Grover VK, Wallace A, Lundberg CJ, Valadez JG, Cooper MK.
PMID: 24867209
Hedgehog (Hh) signaling regulates the growth of malignant gliomas by a ligand-dependent mechanism. The cellular source of Sonic Hh ligand and mode of signaling have not been clearly defined due to the lack of methods to definitively identify neoplastic cells in glioma specimens. Using an antibody specific for mutant isocitrate dehydrogenase protein expression to identify glioma cells, we demonstrate that Sonic Hh ligand and the pathway components Patched1 (PTCH1) and GLI1 are expressed in neoplastic cells. Further, Sonic Hh ligand and its transcriptional targets, PTCH1 and GLI1, are expressed in mutually distinct populations of neoplastic cells. These findings support a paracrine mode of intratumoral Hh signaling in malignant gliomas.
Diagnostic Histopathology
Assessment of human papillomavirus (HPV) status is a requirement for the diagnosis of HPV-associated oropharyngeal squamous cell carcinoma (OPSCC) and metastatic squamous cell carcinoma in cervical lymph nodes where the location of the primary neoplasm is unknown. Within the diagnostic histopathology laboratory, there should be a validated and reproducible HPV testing strategy that can provide HPV status within a reasonable timeframe to inform patient care. Although these requirements are recognized by the head and neck oncology community, there is no internationally accepted standard for HPV testing. A two-tiered approach incorporating p16 immunohistochemistry with specific HPV testing by DNA in situ hybridization is a pragmatic way of providing HPV testing in clinical practice. A novel RNA in situ hybridization methodology targeting E6 and E7 mRNA has been validated and is likely to be available as an in vitro diagnostic device soon. This review will outline the current concepts around the diagnosis of HPV-associated head and neck SCC and suggest a diagnostic algorithm that can be instituted in most diagnostic cellular pathology laboratories.
Oral oncology, 50(1):1–9.
Mirghani H1, Amen F2, Moreau F3, Guigay J4, Ferchiou M5, Melkane AE6, Hartl DM7, Lacau St Guily J (2014).
PMID: 24169585 | DOI: 10.1016/j.oraloncology.2013.10.008.
High risk Human Papilloma virus (HR-HPV) associated oropharyngeal cancers are on the increase. Although, the scientific community is aware of the importance of Human Papilloma Virus (HPV) testing, there is no consensus on the assays that are required to reliably identify HR-HPV related tumors. A wide range of methods have been developed. The most widely used techniques include viral DNA detection, with polymerase chain reaction (PCR) or In Situ Hybridization, and p16 detected by immunohistochemistry. However, these tests provide different information and have their own specific limitations. In this review, we summarize these different techniques, in light of the recent literature. p16 Overexpression, which is an indirect marker of HPV infection, is considered by many head and neck oncologists to be the most important marker for patient stratification. We describe the frequent lack of concordance of this marker with other assays and the possible reasons for this. The latest developments in HPV testing are also reported, such as the RNAscope™ HPV test, and how they fit into the existing framework of techniques. HPV testing must not be considered in isolation, as there are important interactions with other parameters, such as tobacco exposure. This is an important and rapidly evolving field and is likely to become pivotal to staging and choice of treatment of oropharyngeal carcinoma in the future.
Head and neck pathology, 1–7.
Chernock RD, Nussenbaum B, Thorstad WL, Luo Y, Ma XJ, El-Mofty SK, Lewis JS Jr (2013).
PMID: 24151062.
Over the past several decades, it has become clear that human papillomavirus (HPV) is important for the development and progression of many head and neck squamous cell carcinomas, particularly those arising in the oropharyngeal tonsillar crypts. Yet, our understanding of HPV's role in premalignant squamous lesions remains relatively poor. This is in part because premalignant lesions of the oropharyngeal tonsillar crypt tissue, where most HPV-related carcinomas arise, are difficult if not impossible to identify. Recent evidence does suggest a role for HPV in a subset of premalignant lesions of the surface epithelium, especially the oral cavity, despite the rarity of HPV-related invasive squamous cell carcinomas at this site. Furthermore, these HPV-related oral cavity dysplasias appear to have unique, bowenoid histologic features described as 'basaloid' with full-thickness loss of squamous maturation, mitotic figures and apoptosis throughout. Here, we present a unique case of an HPV-related premalignant lesion (squamous cell carcinoma in situ) extensively involving the surface epithelium of the oral cavity, oropharynx and larynx that had 'nonkeratinizing' histologic features typical of HPV-related invasive squamous cell carcinoma. This case was strongly p16 positive by immunohistochemistry and harbored transcriptionally active HPV as demonstrated by E6/E7 RNA in situ hybridization. Furthermore, the patient had an excellent response to radiation treatment.
The Journal of Molecular Diagnostics, 14(1), 22–29.
Wang, F, Flanagan, J, Su N, Wang LC, Bui S, Nielson A, Wu X, Vo HT, Ma XJ, Luo Y. (2012).
PMID: 22166544 | DOI: 10.1016/j.jmoldx.2011.08.002.
In situ analysis of biomarkers is highly desirable in molecular pathology because it allows the examination of biomarker status within the histopathological context of clinical specimens. Immunohistochemistry and DNA in situ hybridization (ISH) are widely used in clinical settings to assess protein and DNA biomarkers, respectively, but clinical use of in situ RNA analysis is rare. This disparity is especially notable when considering the abundance of RNA biomarkers discovered through whole-genome expression profiling. This is largely due to the high degree of technical complexity and insufficient sensitivity and specificity of current RNA ISH techniques. Here, we describe RNAscope, a novel RNA ISH technology with a unique probe design strategy that allows simultaneous signal amplification and background suppression to achieve single-molecule visualization while preserving tissue morphology. RNAscope is compatible with routine formalin-fixed, paraffin-embedded tissue specimens and can use either conventional chromogenic dyes for bright-field microscopy or fluorescent dyes for multiplex analysis. Unlike grind-and-bind RNA analysis methods such as real-time RT-PCR, RNAscope brings the benefits of in situ analysis to RNA biomarkers and may enable rapid development of RNA ISH-based molecular diagnostic assays.
Case Reports in Otolaryngology
Brobst T, García J, Rowe Price KA, Gao G, Smith DI, Price D.
PMID: - | DOI: -
Abstract
Background:
Although alcohol and tobacco use are known risk factors for development of squamous cell carcinoma in the head and neck, human papillomavirus (HPV) has been increasingly associated with this group of cancers. We describe the case of a married couple who presented with HPV-positive oropharynx squamous cell carcinoma within two months of each other.
Methods:
Tumor biopsies were positive for p16 and high-risk HPV in both patients. Sanger sequencing showed a nearly identical HPV16 strain in both patients. Both patients received chemoradiation, and one patient also underwent transoral robotic tongue base resection with bilateral neck dissection.
Results:
Both patients showed no evidence of recurrent disease on follow-up PET imaging.
Conclusions:
New head and neck symptoms should be promptly evaluated in the partner of a patient with known HPV-positive oropharynx cancer. This case expands the limited current literature on concurrent presentation of HPV-positive oropharynx squamous cell carcinoma in couples.
Gerling M, Büller NV, Kirn LM, Joost S, Frings O, Englert B, Bergström Å, Kuiper RV, Blaas L, Wielenga MC, Almer S, Kühl AA, Fredlund E, van den Brink GR, Toftgård R.
PMID: 27492255 | DOI: 10.1038/ncomms12321
A role for Hedgehog (Hh) signalling in the development of colorectal cancer (CRC) has been proposed. In CRC and other solid tumours, Hh ligands are upregulated; however, a specific Hh antagonist provided no benefit in a clinical trial. Here we use Hh reporter mice to show that downstream Hh activity is unexpectedly diminished in a mouse model of colitis-associated colon cancer, and that downstream Hh signalling is restricted to the stroma. Functionally, stroma-specific Hh activation in mice markedly reduces the tumour load and blocks progression of advanced neoplasms, partly via the modulation of BMP signalling and restriction of the colonic stem cell signature. By contrast, attenuated Hh signalling accelerates colonic tumourigenesis. In human CRC, downstream Hh activity is similarly reduced and canonical Hh signalling remains predominantly paracrine. Our results suggest that diminished downstream Hh signalling enhances CRC development, and that stromal Hh activation can act as a colonic tumour suppressor.
El Shahawy M, Reibring CG, Neben CL, Hallberg K, Marangoni P, Harfe BD, Klein OD, Linde A, Gritli-Linde A.
PMID: 28715412 | DOI: 10.1371/journal.pgen.1006914
The interaction between signaling pathways is a central question in the study of organogenesis. Using the developing murine tongue as a model, we uncovered unknown relationships between Sonic hedgehog (SHH) and retinoic acid (RA) signaling. Genetic loss of SHH signaling leads to enhanced RA activity subsequent to loss of SHH-dependent expression of Cyp26a1 and Cyp26c1. This causes a cell identity switch, prompting the epithelium of the tongue to form heterotopic minor salivary glands and to overproduce oversized taste buds. At developmental stages during which Wnt10b expression normally ceases and Shh becomes confined to taste bud cells, loss of SHH inputs causes the lingual epithelium to undergo an ectopic and anachronic expression of Shh and Wnt10b in the basal layer, specifying de novo taste placode induction. Surprisingly, in the absence of SHH signaling, lingual epithelial cells adopted a Merkel cell fate, but this was not caused by enhanced RA signaling. We show that RA promotes, whereas SHH, acting strictly within the lingual epithelium, inhibits taste placode and lingual gland formation by thwarting RA activity. These findings reveal key functions for SHH and RA in cell fate specification in the lingual epithelium and aid in deciphering the molecular mechanisms that assign cell identity.
Nandadasa S, Kraft CM, Wang LW, O'Donnell A, Patel R, Gee HY, Grobe K, Cox TC, Hildebrandt F, Apte SS.
PMID: 30814516 | DOI: 10.1038/s41467-019-08520-7
Although hundreds of cytosolic or transmembrane molecules form the primary cilium, few secreted molecules are known to contribute to ciliogenesis. Here, homologous secreted metalloproteases ADAMTS9 and ADAMTS20 are identified as ciliogenesis regulators that act intracellularly. Secreted and furin-processed ADAMTS9 bound heparan sulfate and was internalized by LRP1, LRP2 and clathrin-mediated endocytosis to be gathered in Rab11 vesicles with a unique periciliary localization defined by super-resolution microscopy. CRISPR-Cas9 inactivation of ADAMTS9 impaired ciliogenesis in RPE-1 cells, which was restored by catalytically active ADAMTS9 or ADAMTS20 acting in trans, but not by their proteolytically inactive mutants. Their mutagenesis in mice impaired neural and yolk sac ciliogenesis, leading to morphogenetic anomalies resulting from impaired hedgehog signaling, which is transduced by primary cilia. In addition to their cognate extracellular proteolytic activity, ADAMTS9 and ADAMTS20 thus have an additional proteolytic role intracellularly, revealing an unexpected regulatory dimension in ciliogenesis.
J Invest Dermatol. 2015 Mar;135(3):701-9.
Chen J, Laclef C, Moncayo A, Snedecor ER, Yang N, Li L, Takemaru K, Paus R, Schneider-Maunoury S, Clark RA.
PMID: 25398052 | DOI: 10.1038/jid.2014.483.
The primary cilium is essential for skin morphogenesis through regulating the Notch, Wnt, and hedgehog signaling pathways. Prior studies on the functions of primary cilia in the skin were based on the investigations of genes that are essential for cilium formation. However, none of these ciliogenic genes has been linked to ciliopathy, a group of disorders caused by abnormal formation or function of cilia. To determine whether there is a genetic and molecular link between ciliopathies and skin morphogenesis, we investigated the role of RPGRIP1L, a gene mutated in Joubert (JBTS) and Meckel (MKS) syndromes, two severe forms of ciliopathy, in the context of skin development. We found that RPGRIP1L is essential for hair follicle morphogenesis. Specifically, disrupting the Rpgrip1l gene in mice resulted in reduced proliferation and differentiation of follicular keratinocytes, leading to hair follicle developmental defects. These defects were associated with significantly decreased primary cilium formation and attenuated hedgehog signaling. In contrast, we found that hair follicle induction and polarization and the development of interfollicular epidermis were unaffected. This study indicates that RPGRIP1L, a ciliopathy gene, is essential for hair follicle morphogenesis likely through regulating primary cilia formation and the hedgehog signaling pathway.
