International journal of surgical pathology
Mannan, R;Wang, X;Bawa, PS;Zhang, Y;Skala, SL;Chinnaiyan, AK;Dagar, A;Wang, L;Zelenka-Wang, SB;McMurry, LM;Daniel, N;Cao, X;Sangoi, AR;Gupta, S;Vaishampayan, UN;Hafez, KS;Morgan, TM;Spratt, DE;Tretiakova, MS;Argani, P;Chinnaiyan, AM;Dhanasekaran, SM;Mehra, R;
PMID: 36250542 | DOI: 10.1177/10668969221125793
Introduction: Chromophobe renal cell carcinoma (chromophobe RCC) is the third major subcategory of renal tumors after clear cell RCC and papillary RCC, accounting for approximately 5% of all RCC subtypes. Other oncocytic neoplasms seen commonly in surgical pathology practice include the eosinophilic variant of chromophobe RCC, renal oncocytoma, and low-grade oncocytic unclassified RCC. Methods: In our recent next-generation sequencing based study, we nominated a lineage-specific novel biomarker LINC01187 (long intergenic non-protein coding RNA 1187) which was found to be enriched in chromophobe RCC. Like KIT (cluster of differentiation 117; CD117), a clinically utilized chromophobe RCC related biomarker, LINC01187 is expressed in intercalated cells of the nephron. In this follow-up study, we performed KIT immunohistochemistry and LINC01187 RNA in situ hybridization (RNA-ISH) on a cohort of chromophobe RCC and other renal neoplasms, characterized the expression patterns, and quantified the expression signals of the two biomarkers in both primary and metastatic settings. Results: LINC01187, in comparison to KIT, exhibits stronger and more uniform expression within tumors while maintaining temporal and spatial consistency. LINC01187 also is devoid of intra-tumoral heterogeneous expression pattern, a phenomenon commonly noted with KIT. Conclusions: LINC01187 expression can augment the currently utilized KIT assay and help facilitate easy microscopic analyses in routine surgical pathology practice.
Virchows Arch. 2015 Aug 5.
Gastrointestinal stromal tumors (GISTs) develop from interstitial cells of Cajal (ICCs) mainly by activating mutations in the KIT or PDGFRA genes. Immunohistochemical analysis for KIT, DOG1, and PKC-θ is used for the diagnosis of GIST. Recently, ETV1 has been shown to be a lineage survival factor for ICCs and required for tumorigenesis of GIST. We investigated the diagnostic value of ETV1expression in GIST. On fresh-frozen tissue samples, RT-PCR analysis showed that ETV1 as well as KIT, DOG1, and PKC-θ are highly expressed in GISTs. On tissue microarrays containing 407 GISTs and 120 non-GIST mesenchymal tumors of GI tract, we performed RNA in situ hybridization (ISH) for ETV1 together with immunohistochemical analysis for KIT, DOG1, PKC-θ, CD133, and CD44. Overall, 387 (95 %) of GISTs were positive for ETV1, while KIT and DOG1 were positive in 381 (94 %) and 392 (96 %) cases, respectively, showing nearly identical overall sensitivity of ETV1, KIT, and DOG1 for GISTs. In addition, ETV1 expression was positively correlated with that of KIT. Notably, ETV1 was positive in 15 of 26 (58 %) KIT-negative GISTs and even positive in 2 cases of GIST negative for KIT and DOG1, whereas only 6 (5 %) non-GIST mesenchymal GI tumors expressed ETV1. We conclude that ETV1 is specifically expressed in the majority of GISTs, even in some KIT-negative cases, suggesting that ETV1 may be useful as ancillary marker in diagnostically difficult select cases of GIST.
Ma D, Wang Z, Yang L, Mu X, Wang Y, Zhao X, Li J, Lin D.
PMID: 27418132 | DOI: 10.18632/oncotarget.10560.
Although the Ventana immunohistochemistry (IHC) platform for detecting anaplastic lymphoma kinase gene (ALK) (D5F3) expression was recently approved by the US Food and Drugs Administration (FDA), fluorescence in situ hybridization (FISH) is still the "gold-standard" method recommended by the US National Comprehensive Cancer Network (NCCN) guideline for NSCLC. We evaluated 6 ALK-positive lung adenocarcinoma patients who tested Ventana IHC-positive and FISH-negative and assessed their clinical responses to the ALK tyrosine kinase inhibitor (TKI) crizotinib. Histologic and cytologic specimens from the 6 patients were stained with Ventana anti-ALK(D5F3) rabbit monoclonal primary antibody using the OptiView™ DAB IHC detection kit and OptiView™ amplification kit on a Ventana BenchMark XT processor. In addition, they were also tested by FISH, qRT-PCR, next-generation sequencing (NGS), and RNAscope ISH analysis. All patients received crizotinib treatment and their follow-up clinical data were recorded. The objective response rate achieved with crizotinib therapy was 66.7% (4/6 partial responses and 2/6 stable disease). One patient in whom a new fusion type (EML4->EXOC6B->ALK fusion) was identified obtained a partial response. These findings indicate that patients with ALK-positive lung adenocarcinoma who test Ventana IHC-positive and FISH-negative may still respond to crizotinib therapy.
Fomitcheva-Khartchenko, A;Kashyap, A;Geiger, T;Kaigala, GV;
PMID: 35995681 | DOI: 10.1016/j.trecan.2022.07.008
Tumor cells present complex behaviors in their interactions with other cells. This intricate behavior is driving the need to develop new tools to understand these ecosystems. The surge of spatial technologies allows evaluation of the complexity of relationships between cells present in a tumor, giving insights about tumor heterogeneity and the tumor microenvironment while providing clinically relevant metrics for tumor classification. In this review, we describe key results obtained using spatial techniques, present recent advances in methods to uncover spatially relevant biological significance, and summarize their main characteristics. We expect spatial technologies to significantly broaden our understanding of tumor biology and to generate clinically relevant tools that will ultimately impact personalized medicine.
Journal of Hematopathology
Shi, Y;Mi, L;Lai, Y;Zhao, M;Jia, L;Du, T;Song, Y;Li, X;
| DOI: 10.1007/s12308-023-00530-1
Overexpression of PD-L1 can be a predictive marker for anti-PD-1 therapeutic efficacy in classic Hodgkin lymphoma (CHL); however, harmonization of different IHC assays remains to be accomplished, and interpretations of PD-L1 immunostaining results remain controversial in CHL. In this study, we sought to optimize the PD-L1 immunohistochemistry (IHC) assay in CHL. All tests were performed on a tumour tissue microarray established from 54 CHL cases. Three IHC antibodies (405.9A11, SP142, 22C3) for detecting PD-L1 expression were compared semi quantitatively with the RNAscope assay (No. 310035, ACD), and the difference in the expression in background immune cells (ICs) between assays and the associations of expression levels with densities of TILs/TAMs were also analysed. 405.9A11 demonstrated best specificity in HRS cells and best sensitivity in ICs. Positive expression of PD-L1 was more frequent in ICs (85.2%) than in HRS cells (48.1%). Different subgroups of background ICs, including tumour-associated macrophages (TAMs), were assessed and scored for CD4, CD8, FOXP3, and CD163 expression. PD-L1 expression on ICs was the factor most associated with the density of TAMs. 405.9A11 provided the most convincing PD-L1 expression results. Pathologists should report PD-L1 expression in a combined manner, including both the status of HRS cells and the percentage of PD-L1-positive ICs.
bioRxiv : the preprint server for biology
Chen, J;Zheng, Q;Hicks, JL;Trabzonlu, L;Ozbek, B;Jones, T;Vaghasia, A;Larman, TC;Wang, R;Markowski, MC;Denmeade, SR;Pienta, KJ;Hruban, RH;Antonaraskis, ES;Gupta, A;Dang, CV;Yegnasubramanian, S;De Marzo, AM;
PMID: 36865273 | DOI: 10.1101/2023.02.20.529259
Increased mitochondrial function may render some cancers vulnerable to mitochondrial inhibitors. Since mitochondrial function is regulated partly by mitochondrial DNA copy number (mtDNAcn), accurate measurements of mtDNAcn could help reveal which cancers are driven by increased mitochondrial function and may be candidates for mitochondrial inhibition. However, prior studies have employed bulk macrodissections that fail to account for cell type-specific or tumor cell heterogeneity in mtDNAcn. These studies have often produced unclear results, particularly in prostate cancer. Herein, we developed a multiplex in situ method to spatially quantify cell type specific mtDNAcn. We show that mtDNAcn is increased in luminal cells of high-grade prostatic intraepithelial neoplasia (HGPIN), is increased in prostatic adenocarcinomas (PCa), and is further elevated in metastatic castration-resistant prostate cancer. Increased PCa mtDNAcn was validated by two orthogonal methods and is accompanied by increases in mtRNAs and enzymatic activity. Mechanistically, MYC inhibition in prostate cancer cells decreases mtDNA replication and expression of several mtDNA replication genes, and MYC activation in the mouse prostate leads to increased mtDNA levels in the neoplastic prostate cells. Our in situ approach also revealed elevated mtDNAcn in precancerous lesions of the pancreas and colon/rectum, demonstrating generalization across cancer types using clinical tissue samples.
Li, Y;Lih, TM;Dhanasekaran, SM;Mannan, R;Chen, L;Cieslik, M;Wu, Y;Lu, RJ;Clark, DJ;Kołodziejczak, I;Hong, R;Chen, S;Zhao, Y;Chugh, S;Caravan, W;Naser Al Deen, N;Hosseini, N;Newton, CJ;Krug, K;Xu, Y;Cho, KC;Hu, Y;Zhang, Y;Kumar-Sinha, C;Ma, W;Calinawan, A;Wyczalkowski, MA;Wendl, MC;Wang, Y;Guo, S;Zhang, C;Le, A;Dagar, A;Hopkins, A;Cho, H;Leprevost, FDV;Jing, X;Teo, GC;Liu, W;Reimers, MA;Pachynski, R;Lazar, AJ;Chinnaiyan, AM;Van Tine, BA;Zhang, B;Rodland, KD;Getz, G;Mani, DR;Wang, P;Chen, F;Hostetter, G;Thiagarajan, M;Linehan, WM;Fenyö, D;Jewell, SD;Omenn, GS;Mehra, R;Wiznerowicz, M;Robles, AI;Mesri, M;Hiltke, T;An, E;Rodriguez, H;Chan, DW;Ricketts, CJ;Nesvizhskii, AI;Zhang, H;Ding, L;Clinical Proteomic Tumor Analysis Consortium, ;
PMID: 36563681 | DOI: 10.1016/j.ccell.2022.12.001
Clear cell renal cell carcinomas (ccRCCs) represent ∼75% of RCC cases and account for most RCC-associated deaths. Inter- and intratumoral heterogeneity (ITH) results in varying prognosis and treatment outcomes. To obtain the most comprehensive profile of ccRCC, we perform integrative histopathologic, proteogenomic, and metabolomic analyses on 305 ccRCC tumor segments and 166 paired adjacent normal tissues from 213 cases. Combining histologic and molecular profiles reveals ITH in 90% of ccRCCs, with 50% demonstrating immune signature heterogeneity. High tumor grade, along with BAP1 mutation, genome instability, increased hypermethylation, and a specific protein glycosylation signature define a high-risk disease subset, where UCHL1 expression displays prognostic value. Single-nuclei RNA sequencing of the adverse sarcomatoid and rhabdoid phenotypes uncover gene signatures and potential insights into tumor evolution. In vitro cell line studies confirm the potential of inhibiting identified phosphoproteome targets. This study molecularly stratifies aggressive histopathologic subtypes that may inform more effective treatment strategies.
The Journal of clinical investigation
Duhen, R;Fesneau, O;Samson, KA;Frye, AK;Beymer, M;Rajamanickam, V;Ross, D;Tran, E;Bernard, B;Weinberg, AD;Duhen, T;
PMID: 35439168 | DOI: 10.1172/JCI156821
CD4 T helper (Th) cells play a key role in orchestrating immune responses, but the identity of the CD4 Th cells involved in the anti-tumor immune response remains to be defined. We analyzed the immune cell infiltrates of head and neck squamous cell carcinoma and colorectal cancers and identified a subset of CD4 Th cells distinct from FOXP3+ regulatory T cells that co-express PD-1 and ICOS. These tumor-infiltrating CD4 Th cells (CD4 Th TIL) have a tissue-resident memory phenotype, are present in MHC class II-rich areas and proliferate in the tumor suggesting local antigen recognition. The T-cell receptor repertoire of the PD-1+ICOS+ CD4 Th TIL is oligoclonal, with T-cell clones expanded in the tumor, but present at low frequencies in the periphery. Finally, these PD-1+ICOS+ CD4 Th TIL were shown to recognize both tumor-associated antigens and tumor-specific neoantigens. Our findings provide an approach for isolating tumor-reactive CD4 Th TIL directly ex vivo that will help define their role in the anti-tumor immune response and potentially improve future adoptive T-cell therapy approaches.
Cancer immunology, immunotherapy : CII
Kawaguchi, S;Kawahara, K;Fujiwara, Y;Ohnishi, K;Pan, C;Yano, H;Hirosue, A;Nagata, M;Hirayama, M;Sakata, J;Nakashima, H;Arita, H;Yamana, K;Gohara, S;Nagao, Y;Maeshiro, M;Iwamoto, A;Hirayama, M;Yoshida, R;Komohara, Y;Nakayama, H;
PMID: 35044489 | DOI: 10.1007/s00262-022-03149-w
The CD169+ macrophages in lymph nodes are implicated in cytotoxic T lymphocyte (CTL) activation and are associated with improved prognosis in several malignancies. Here, we investigated the significance of CD169+ macrophages in oral squamous cell carcinoma (OSCC). Further, we tested the anti-tumor effects of naringenin, which has been previously shown to activate CD169+ macrophages, in a murine OSCC model. Immunohistochemical analysis for CD169 and CD8 was performed on lymph node and primary tumor specimens from 89 patients with OSCC. We also evaluated the effects of naringenin on two murine OSCC models. Increased CD169+ macrophage counts in the regional lymph nodes correlated with favorable prognosis and CD8+ cell counts within tumor sites. Additionally, naringenin suppressed tumor growth in two murine OSCC models. The mRNA levels of CD169, interleukin (IL)-12, and C-X-C motif chemokine ligand 10 (CXCL10) in lymph nodes and CTL infiltration in tumors significantly increased following naringenin administration in tumor-bearing mice. These results suggest that CD169+ macrophages in lymph nodes are involved in T cell-mediated anti-tumor immunity and could be a prognostic marker for patients with OSCC. Moreover, naringenin is a new potential agent for CD169+ macrophage activation in OSCC treatment.
Epigenomic landscape of human colorectal cancer unveils an aberrant core of pan-cancer enhancers orchestrated by YAP/TAZ
Della Chiara, G;Gervasoni, F;Fakiola, M;Godano, C;D'Oria, C;Azzolin, L;Bonnal, RJP;Moreni, G;Drufuca, L;Rossetti, G;Ranzani, V;Bason, R;De Simone, M;Panariello, F;Ferrari, I;Fabbris, T;Zanconato, F;Forcato, M;Romano, O;Caroli, J;Gruarin, P;Sarnicola, ML;Cordenonsi, M;Bardelli, A;Zucchini, N;Ceretti, AP;Mariani, NM;Cassingena, A;Sartore-Bianchi, A;Testa, G;Gianotti, L;Opocher, E;Pisati, F;Tripodo, C;Macino, G;Siena, S;Bicciato, S;Piccolo, S;Pagani, M;
PMID: 33879786 | DOI: 10.1038/s41467-021-22544-y
Cancer is characterized by pervasive epigenetic alterations with enhancer dysfunction orchestrating the aberrant cancer transcriptional programs and transcriptional dependencies. Here, we epigenetically characterize human colorectal cancer (CRC) using de novo chromatin state discovery on a library of different patient-derived organoids. By exploring this resource, we unveil a tumor-specific deregulated enhancerome that is cancer cell-intrinsic and independent of interpatient heterogeneity. We show that the transcriptional coactivators YAP/TAZ act as key regulators of the conserved CRC gained enhancers. The same YAP/TAZ-bound enhancers display active chromatin profiles across diverse human tumors, highlighting a pan-cancer epigenetic rewiring which at single-cell level distinguishes malignant from normal cell populations. YAP/TAZ inhibition in established tumor organoids causes extensive cell death unveiling their essential role in tumor maintenance. This work indicates a common layer of YAP/TAZ-fueled enhancer reprogramming that is key for the cancer cell state and can be exploited for the development of improved therapeutic avenues.
Archives of pathology & laboratory medicine
Haqshenas, G;Molano, M;Phillips, S;Balgovind, P;Garland, SM;Hawkes, D;Brotherton, JM;Machalek, DA;Murray, G;
PMID: 37226838 | DOI: 10.5858/arpa.2022-0317-OA
Detection of human papillomavirus (HPV) in formalin-fixed, paraffin-embedded (FFPE) tissues may identify the cause of lesions and has value for the development of new diagnostic assays and epidemiologic studies. Seegene Anyplex II assays are widely used for HPV screening, but their performance using FFPE samples has not been fully explored.To validate Anyplex II HPV HR Detection (Anyplex II, Seegene) using FFPE samples.We used 248 stored DNA extracts from cervical cancer FFPE samples collected during 2005-2015 and that tested HPV positive using the RHA kit HPV SPF10-LiPA25, v1 (SPF10, Labo Biomedical Products) HPV genotyping assay, manufacturer-validated for FFPE samples.Of the selected 248 samples, 243 were used in our analysis. Consistent with SPF10 genotyping results, Anyplex II detected all 12 oncogenic types and had an overall HPV detection rate of 86.4% (210 of 243 samples). Anyplex II and SPF10 showed very high agreement for the detection of the 2 most important oncogenic genotypes: HPV 16 (219 of 226; 96.9%; 95% CI, 93.7-98.75) and HPV 18 (221 of 226; 97.8%; 95% CI, 94.9-99.3).Overall results showed that both platforms produced comparable HPV genotyping results, indicating the suitability of Anyplex II for FFPE samples. The Anyplex II assay has the added convenience of being an efficient, single-well semiquantitative polymerase chain reaction assay. Further optimization of Anyplex II may enhance its performance using FFPE samples by improving the detection limit.
Tahara, S;de Faria, FCC;Sarchet, P;Calore, F;Sharick, J;Leight, JL;Casadei, L;Pollock, RE;
PMID: 36763259 | DOI: 10.1007/s13577-023-00865-y
Sarcomas are rare malignancies, the number of reports is limited, and this rarity makes further research difficult even though liposarcoma is one of major sarcomas. 2D cell culture remains an important role in establishing basic tumor biology research, but its various shortcomings and limitations are still of concern, and it is now well-accepted that the behavior of 3D-cultured cells is more reflective of in vivo cellular responses compared to 2D models. This study aimed to establish 3D cell culture of liposarcomas using two different methods: scaffold-based (Matrigel extracellular matrix [ECM] scaffold method) and scaffold-free (Ultra-low attachment [ULA] plate). Lipo246, Lipo224 and Lipo863 cell lines were cultured, and distinctive differences in structures were observed in Matrigel 3D model: Lipo224 and Lipo863 formed spheroids, whereas Lipo246 grew radially without forming spheres. In ULA plate approaches, all cell lines formed spheroids, but Lipo224 and Lipo863 spheroids showed bigger size and looser aggregation than Lipo246. Formalin fixed, paraffin embedded (FFPE) blocks were obtained from all 3D models, confirming the spheroid structures. The expression of MDM2, Ki-67 positivity and MDM2 amplification were confirmed by IHC and DNAscope , respectively. Protein and DNA were extracted from all samples and MDM2 upregulation was confirmed by western blot and qPCR analysis. After treatment with MDM2 inhibitor SAR405838, DDLPS spheroids demonstrated different sensitivity patterns from 2D models. Taken together, we believed that 3D models would have a possibility to provide us a new predictability of efficacy and toxicity, and considered as one important process in in vitro pre-clinical phase prior to moving forward to clinical trials.
