Predictors of Recurrence and Progression in Poorly Differentiated Cutaneous Squamous Cell Carcinomas: Insights from a Real-Life Experience.

INTRODUCTION: Surgery represents the primary treatment option for cutaneous squamous cell carcinoma (cSCC) aiming for complete tumor resection (R0). Recurrence and metastasis significantly affect survival and outcomes, and poorly differentiated (G3) cSCC is associated with a higher risk of recurrence. However, the specific clinical and histopathological features that predict recurrence and progression in G3-cSCC remain unclear. METHODS: A retrospective analysis was conducted on a series of patients with primary G3-cSCC diagnosed at the Turin University Hospital between January 2016 and January 2021. After independent histological revision, logistic regression models were used to identify clinico-pathological predictors of cutaneous recurrence, lymphnode/metastatic progression, and both types of progression. RESULTS: Among the 161 G3-cSCC patients, 80.1% (129/161) showed no signs of local recurrence or metastatic progression, while 19.9% (32 patients) had progressed. In the univariate logistic regression, tumor clinical diameter, depth of infiltration (DOI), and lymphovascular invasion (LVI) were identified as significant predictors across the various types of progression (p < 0.05). In the context of multivariate logistic regression, distinct models proved to be significant. For skin recurrence, a 3-variable model incorporating DOI (OR 1.16, 95% CI, 1.01-1.35, p = 0.050), LVI (OR 3.61, 95% CI, 1.11-11.8, p = 0.034), and desmoplasia (OR 3.45, 95% CI, 1.25-9.5, p = 0.017) was selected. Regarding lymphnode/metastatic progression, a 3-variable model combining pT2 (OR 6.10, 95% CI, 1.15-32.35, p = 0.034), pT3 (OR 14.33, 95% CI, 2.79-73.63, p = 0.001), and LVI (OR 3.86, 95% CI, 1.10-13.62, p = 0.036) was identified. Lastly, a 2-variable model for both types of progression consisted of vertical tumor thickness (OR 5.45, 95% CI, 1.11-27.32, p = 0.039) and LVI (OR 1.15, 95% CI, 1.04-1.26, p = 0.006). CONCLUSION: Tumor size, DOI, and LVI were significant predictors of recurrence and metastatic progression. Notably, the size of histologically defined tumor-free margins did not affect the risk of recurrence, whilst LVI emerged as a key predictor of all forms of progression. These findings provide insights into risk stratification and suggest that close monitoring and potential adjuvant therapies, such as radiation therapy, may be necessary especially for patients with lymphovascular involvement.

Pulmonary Low Malignant Potential Adenocarcinoma: A Validation of the Proposed Criteria for This Novel Subtype.

Adenocarcinoma (ADC) is the most common histologic type of lung cancer, including in situ (lepidic), minimally invasive, and invasive forms. While the former 2 types are associated with a favorable outcome, the latter includes tumors with variable behavior, often tumor stage-related. A recent study proposed strict morphologic criteria defining a new subgroup of resected stage I invasive ADC (16% of cases) with favorable outcomes (100% disease-specific survival), named "ADC of low malignant potential (LMP-ADC)." The following criteria were met: ≤3 cm size, nonmucinous histotype, ≥15% lepidic growth, and the absence of the following: high-grade patterns, >1 mitosis/2 mm 2 , necrosis, and vascular/pleural invasion. The aim of the present study was to validate the performance of such criteria to identify LMP-ADC in a series of 274 stage IA resected lung ADCs from a single institution. Thirty-four tumors (12.4%) met the proposed criteria for LMP-ADC, as confirmed by additional stains for mitotic figures, Ki67 index, and elastic fibers (helpful to assess alveolar wall invasion). Minor differences between the lepidic and invasive components were observed regarding cell atypia and proliferation. p53 was normally expressed by invasive tumor cells. Mutations occurred in known lung cancer genes (mostly KRAS and EGFR). Five patients (14.7%) developed disease progression and 2 of them (5.9%) died of the disease. In our series, the disease-specific survival was 94.1%. In conclusion, in resected invasive lung ADC, a subgroup presenting low-grade morphologic features and associated with favorable prognosis does exist. Morphologic criteria for LMP-ADC supported by ancillary techniques represent a valid tool to better define this novel subgroup and to refine the stratification of invasive lung ADC, possibly suggesting modified follow-up protocols, based on the observed indolent behavior in most cases.

SS18-SSX Antibody: A Useful Tool to Save Time and Reduce Costs in Synovial Sarcoma Diagnosis. Proposal of a Novel Diagnostic Algorithm.

Synovial sarcoma is a rare malignant mesenchymal neoplasm mostly affecting young adults, characterized by a specific translocation which results in the fusion of the SS18 gene on chromosome 18 with one of the three highly homologous SSX genes on chromosome X. Its morphological diagnosis, especially in monophasic or poorly differentiated variants, can be challenging because histological features often overlap with other malignant mesenchymal tumors. Until recently, the differential diagnosis mostly relied on the use of cytogenetic or molecular analyses to detect the specific t(X;18)(p11;q11) translocation, thus virtually restricting its correct identification to referral centers with a high histological and molecular pathology workflow. The recently commercialized highly sensitive and fusion-specific SS18-SSX antibody has significantly improved the approach to these tumors, representing a relatively cheap and easy to access tool for synovial sarcoma diagnosis. Through a retrospective analysis of 79 synovial sarcomas and histological mimickers, this study confirms the usefulness of the SS18-SSX antibody in the diagnosis of synovial sarcoma, particularly focusing on its application in the pathological response evaluation after neoadjuvant treatment as well as its time- and cost-saving advantages. Finally, we here propose a new diagnostic algorithm to apply into the routine practice.

COVID-19 and Breast Cancer: Analysis of Surgical Management of a Large Referral Center during the 2020-2021 Pandemic Period.

BACKGROUND: Coronavirus disease-19 (COVID-19) has spread worldwide since December 2019 and was officially declared a pandemic in March 2020. Due to the rapid transmission and the high fatality rate, drastic emergency restrictions were issued, with a negative impact on routine clinical activities. In particular, in Italy, many authors have reported a reduction in the number of breast cancer diagnoses and critical problems in the management of patients who accessed the breast units during the dramatic first months of the pandemic. Our study aims to analyze the global impact of COVID-19 in the two years of the pandemic (2020-2021) on the surgical management of breast cancer by comparing them with the previous two years. METHODS: In our retrospective study, we analyzed all cases of breast cancer diagnosed and surgically treated at the breast unit of "Città della Salute e della Scienza" in Turin, Italy, making a comparison between the 2018-2019 pre-pandemic period and the 2020-2021 pandemic period. RESULTS: We included in our analysis 1331 breast cancer cases surgically treated from January 2018 to December 2021. A total of 726 patients were treated in the pre-pandemic years and 605 in the pandemic period (-121 cases, 9%). No significant differences were observed regarding diagnosis (screening vs. no screening) and timing between radiological diagnosis and surgery for both in situ and invasive tumors. There were no variations in the breast surgical approach (mastectomy vs. conservative surgery), while a reduction in axillary dissection compared to the sentinel lymph node in the pandemic period was observed (p-value < 0.001). Regarding the biological characteristics of breast cancers, we observed a greater number of grades 2-3 (p-value = 0.007), pT stage 3-4 breast cancer surgically treated without previous neoadjuvant chemotherapy (p-value = 0.03), and a reduction in luminal B tumors (p-value = 0.007). CONCLUSIONS: Overall, we report a limited reduction in surgical activity for breast cancer treatment considering the entire pandemic period (2020-2021). These results suggest a prompt resumption of surgical activity similar to the pre-pandemic period.

Clinical and Pathological Features and Gene Expression Profiles of Clinically Aggressive Papillary Thyroid Carcinomas.

Papillary thyroid carcinoma (PTC) is considered an indolent neoplasm but it may demonstrate aggressive behavior. We aimed to identify clinical and pathological characteristics and molecular signatures associated with aggressive forms of PTCs. We selected 43 aggressive PTC cases based on the presence of metastases at the time of diagnosis, the development of distant metastasis during follow-up, and/or biochemical recurrence, and 43 PTC patients that were disease-free upon follow-up, matching them according to age, sex, pT, and pN parameters. Twenty-four pairs (a total of 48 cases) and 6 normal thyroid tissues were studied using targeted mRNA screening of cancer-associated genes employing NanoString nCounter® technology. In general, aggressive PTCs showed distinctive clinical and morphological features. Among adverse prognostic parameters, the presence of necrosis and an increased mitotic index were associated with shorter disease-free and overall survivals. Other parameters associated with shorter disease-free or overall survivals include a lack of tumor capsule, the presence of vascular invasion, tumor-infiltrating lymphocytes, fibrosclerotic changes, age > 55 years, and a high pTN stage. Various pathways were differentially regulated in non-aggressive as compared to aggressive PTC, including the DNA damage repair, the MAPK, and the RAS pathways. In particular, the hedgehog pathway was differentially de-regulated in aggressive PTC as compared to non-aggressive PTC cases, being WNT10A and GLI3 genes significantly up- and down-regulated in aggressive PTC and GSK3B up-regulated in non-aggressive PTC cases. In conclusion, our study revealed specific molecular signatures and morphological features in aggressive PTC that may be useful to predict more aggressive behavior in a subset of PTC patients. These findings may be useful when developing novel, tailored treatment options for these patients.

Predicting outcome in dogs with diffuse large B-cell lymphoma with a novel immune landscape signature.

Canine diffuse large B-cell lymphoma (cDLBCL) is characterized by high mortality and clinical heterogeneity. Although chemo-immunotherapy improves outcome, treatment response remains mainly unpredictable. To identify a set of immune-related genes aberrantly regulated and impacting the prognosis, we explored the immune landscape of cDLBCL by NanoString. The immune gene expression profile of 48 fully clinically characterized cDLBCLs treated with chemo-immunotherapy was analyzed with the NanoString nCounter Canine IO Panel using RNA extracted from tumor tissue paraffin blocks. A Cox proportional-hazards model was used to design a prognostic gene signature. The Cox model identified a 6-gene signature (IL2RB, BCL6, TXK, C2, CDKN2B, ITK) strongly associated with lymphoma-specific survival, from which a risk score was calculated. Dogs were assigned to high-risk or low-risk groups according to the median score. Thirty-nine genes were differentially expressed between the 2 groups. Gene set analysis highlighted an upregulation of genes involved in complement activation, cytotoxicity, and antigen processing in low-risk dogs compared with high-risk dogs, whereas genes associated with cell cycle were downregulated in dogs with a lower risk. In line with these results, cell type profiling suggested the abundance of natural killer and CD8+ cells in low-risk dogs compared with high-risk dogs. Furthermore, the prognostic power of the risk score was validated in an independent cohort of cDLBCL. In conclusion, the 6-gene-derived risk score represents a robust biomarker in predicting the prognosis in cDLBCL. Moreover, our results suggest that enhanced tumor antigen recognition and cytotoxic activity are crucial in achieving a more effective response to chemo-immunotherapy.

First pilot case-control interventional study using autologous extracellular vesicles to treat chronic venous ulcers unresponsive to conventional treatments.

Current therapeutic approaches for chronic venous ulcers (CVUs) still require evidence of effectiveness. Diverse sources of extracellular vesicles (EVs) have been proposed for tissue regeneration, however the lack of potency tests, to predict in-vivo effectiveness, and a reliable scalability have delayed their clinical application. This study aimed to investigate whether autologous serum-derived EVs (s-EVs), recovered from patients with CVUs, may be a proper therapeutic approach to improve the healing process. A pilot case-control interventional study (CS2/1095/0090491) has been designed and s-EVs recovered from patients. Patient eligibility included two or more distinct chronic lesions in the same limb with 11 months as median persistence of active ulcer before enrollment. Patients were treated three times a week, for 2 weeks. Qualitative CVU analysis demonstrated that s-EVs-treated lesions displayed a higher percentage of granulation tissue compared to the control group (Sham) (s-EVs 3 out of 5: 75-100 % vs Sham: none), further confirmed at day 30. s-EVs-treated lesions also displayed higher sloughy tissue reduction at the end of treatment even increased at day 30. Additionally, s-EV treatment led to a median surface reduction of 151 mm2 compared to 84 mm2 in the Sham group, difference even more evident at day 30 (s-EVs 385 mm2vs Sham 106 mm2p = 0.004). Consistent with the enrichment of transforming growth factor-ß1 in s-EVs, histological analyses showed a regenerative tissue with an increase in microvascular proliferation areas. This study first demonstrates the clinical effectiveness of autologous s-EVs in promoting the healing process of CVUs unresponsive to conventional treatments.

Antagonist of Growth Hormone-Releasing Hormone Potentiates the Antitumor Effect of Pemetrexed and Cisplatin in Pleural Mesothelioma.

Pleural mesothelioma (PM) is an aggressive cancer with poor prognosis and no effective therapies, mainly caused by exposure to asbestos. Antagonists of growth hormone-releasing hormone (GHRH) display strong antitumor effects in many experimental cancers, including lung cancer and mesothelioma. Here, we aimed to determine whether GHRH antagonist MIA-690 potentiates the antitumor effect of cisplatin and pemetrexed in PM. In vitro, MIA-690, in combination with cisplatin and pemetrexed, synergistically reduced cell viability, restrained cell proliferation and enhanced apoptosis, compared with drugs alone. In vivo, the same combination resulted in a strong growth inhibition of MSTO-211H xenografts, decreased tumor cell proliferation and increased apoptosis. Mechanistically, MIA-690, particularly with chemotherapeutic drugs, inhibited proliferative and oncogenic pathways, such as MAPK ERK1/2 and cMyc, and downregulated cyclin D1 and B1 mRNAs. Inflammatory pathways such as NF-kB and STAT3 were also reduced, as well as oxidative, angiogenic and tumorigenic markers (iNOS, COX-2, MMP2, MMP9 and HMGB1) and growth factors (VEGF and IGF-1). Overall, these findings strongly suggest that GHRH antagonists of MIA class, such as MIA-690, could increase the efficacy of standard therapy in PM.

Overexpression of INSM1, NOTCH1, NEUROD1, and YAP1 genes is associated with adverse clinical outcome in pediatric neuroblastoma.

Pediatric neuroblastoma is responsible for approximately 8-10% of pediatric tumors, and it is one of the leading causes of tumor-related deaths in children. Although significant progress has been made in the characterization of neuroblastoma in recent years, the mechanisms influencing the prognosis of neuroblastoma patients remain largely unknown. Our aim was to investigate if the major neuroendocrine-associated transcriptional drivers, including ASCL1, NEUROD1, DLL3, NOTCH1, INSM1, MYCL1, POU2F3 and YAP1 are correlated with specific clinical and pathological characteristics. We selected a retrospective series of 46 primary pediatric neuroblastoma, composed of 30 treatment-naïve and 16 post-chemotherapy cases. Gene expression levels were explored by means of quantitative real-time PCR. An increased expression of NOTCH1 (p = 0.005), NEUROD1 (p = 0.0059), and YAP1 (p = 0.0008) was found in stage IV tumors, while the highest levels of MYCL1 and ASCL1 were seen in stages IVS and III, respectively (p = 0.0182 and p = 0.0134). A higher level of NOTCH1 (p = 0.0079) and YAP1 (p = 0.0026) was found in cases with differentiating morphology, while high mitosis-karyorrhexis index cases demonstrated significantly lower levels of POU2F3 (p = 0.0277). High expression of NOTCH1 (p = 0.008), NEUROD1 (p = 0.026), INSM1 (p = 0.010), and YAP1 (p = 0.005) together with stage IV (p = 0.043) was associated with shorter disease-free survival. In summary, our data indicate that the assessment of gene expression levels of neuroendocrine-lineage transcription factors might help to identify neuroblastoma patients with the risk of relapse.

Prognostic Analysis of the IDH1 G105G (rs11554137) SNP in IDH-Wildtype Glioblastoma.

The G105G SNP (rs11554137) in the IDH1 gene is observed in about 10-15% of patients with a diffuse glioma. Data regarding its impact on gliomas are poor and partially conflicting, possibly due to the evolving classification of CNS tumors. The aim of this study was to investigate the G105G SNP prognostic significance in a homogenous cohort of IDH-wildtype glioblastomas, in agreement with the 2021 WHO classification. The study analyzed 211 patients by collecting several clinico-pathological and molecular characteristics, including the age, lesion localization, number of involved lobes, type of surgical treatment, disease outcome and MGMT promoter methylation status. PFS and DSS curves were plotted according to the Kaplan-Meier method and statistical analyses were performed using parametric and non-parametric tests. A total of 32 patients out of 211 (15.2%) were found to be G105G SNP carriers. No significant impact of the IDH1 G105G SNP on patients' outcomes was observed in terms of PFS and DSS, while MGMT promoter methylation and gross total resection resulted as key prognostic factors in our cohort as expected. No prognostic impact of the IDH1 G105G SNP was detected in this strict cohort of IDH-wildtype glioblastomas. Analysis of larger cohorts is warranted to address the sample size limitations.

Transcriptome-wide association study of multiple myeloma identifies candidate susceptibility genes.

BACKGROUND: While genome-wide association studies (GWAS) of multiple myeloma (MM) have identified variants at 23 regions influencing risk, the genes underlying these associations are largely unknown. To identify candidate causal genes at these regions and search for novel risk regions, we performed a multi-tissue transcriptome-wide association study (TWAS). RESULTS: GWAS data on 7319 MM cases and 234,385 controls was integrated with Genotype-Tissue Expression Project (GTEx) data assayed in 48 tissues (sample sizes, N = 80-491), including lymphocyte cell lines and whole blood, to predict gene expression. We identified 108 genes at 13 independent regions associated with MM risk, all of which were in 1 Mb of known MM GWAS risk variants. Of these, 94 genes, located in eight regions, had not previously been considered as a candidate gene for that locus. CONCLUSIONS: Our findings highlight the value of leveraging expression data from multiple tissues to identify candidate genes responsible for GWAS associations which provide insight into MM tumorigenesis. Among the genes identified, a number have plausible roles in MM biology, notably APOBEC3C, APOBEC3H, APOBEC3D, APOBEC3F, APOBEC3G, or have been previously implicated in other malignancies. The genes identified in this TWAS can be explored for follow-up and validation to further understand their role in MM biology.

Insight into genetic predisposition to chronic lymphocytic leukemia from integrative epigenomics.

Genome-wide association studies have provided evidence for inherited genetic predisposition to chronic lymphocytic leukemia (CLL). To gain insight into the mechanisms underlying CLL risk we analyze chromatin accessibility, active regulatory elements marked by H3K27ac, and DNA methylation at 42 risk loci in up to 486 primary CLLs. We identify that risk loci are significantly enriched for active chromatin in CLL with evidence of being CLL-specific or differentially regulated in normal B-cell development. We then use in situ promoter capture Hi-C, in conjunction with gene expression data to reveal likely target genes of the risk loci. Candidate target genes are enriched for pathways related to B-cell development such as MYC and BCL2 signalling. At 14 loci the analysis highlights 63 variants as the probable functional basis of CLL risk. By integrating genetic and epigenetic information our analysis reveals novel insights into the relationship between inherited predisposition and the regulatory chromatin landscape of CLL.

Association analyses identify 31 new risk loci for colorectal cancer susceptibility.

Colorectal cancer (CRC) is a leading cause of cancer-related death worldwide, and has a strong heritable basis. We report a genome-wide association analysis of 34,627 CRC cases and 71,379 controls of European ancestry that identifies SNPs at 31 new CRC risk loci. We also identify eight independent risk SNPs at the new and previously reported European CRC loci, and a further nine CRC SNPs at loci previously only identified in Asian populations. We use in situ promoter capture Hi-C (CHi-C), gene expression, and in silico annotation methods to identify likely target genes of CRC SNPs. Whilst these new SNP associations implicate target genes that are enriched for known CRC pathways such as Wnt and BMP, they also highlight novel pathways with no prior links to colorectal tumourigenesis. These findings provide further insight into CRC susceptibility and enhance the prospects of applying genetic risk scores to personalised screening and prevention.

Author Correction: Identification of multiple risk loci and regulatory mechanisms influencing susceptibility to multiple myeloma.

The original version of this Article contained an error in the spelling of a member of the PRACTICAL Consortium, Manuela Gago-Dominguez, which was incorrectly given as Manuela Gago Dominguez. This has now been corrected in both the PDF and HTML versions of the Article. Furthermore, in the original HTML version of this Article, the order of authors within the author list was incorrect. The PRACTICAL consortium was incorrectly listed after Richard S. Houlston and should have been listed after Nora Pashayan. This error has been corrected in the HTML version of the Article; the PDF version was correct at the time of publication.

Identification of recurrent noncoding mutations in B-cell lymphoma using capture Hi-C.

The identification of driver mutations is fundamental to understanding oncogenesis. Although genes frequently mutated in B-cell lymphoma have been identified, the search for driver mutations has largely focused on the coding genome. Here we report an analysis of the noncoding genome using whole-genome sequencing data from 117 patients with B-cell lymphoma. Using promoter capture Hi-C data in naive B cells, we define cis-regulatory elements, which represent an enriched subset of the noncoding genome in which to search for driver mutations. Regulatory regions were identified whose mutation significantly alters gene expression, including copy number variation at cis-regulatory elements targeting CD69, IGLL5, and MMP14, and single nucleotide variants in a cis-regulatory element for TPRG1 We also show the commonality of pathways targeted by coding and noncoding mutations, exemplified by MMP14, which regulates Notch signaling, a pathway important in lymphomagenesis and whose expression is associated with patient survival. This study provides an enhanced understanding of lymphomagenesis and describes the advantages of using chromosome conformation capture to decipher noncoding mutations relevant to cancer biology.

Author Correction: Genome-wide association study of classical Hodgkin lymphoma identifies key regulators of disease susceptibility.

The original version of this Article contained an error in the spelling of a member of the PRACTICAL Consortium, Manuela Gago-Dominguez, which was incorrectly given as Manuela Gago Dominguez. This has now been corrected in both the PDF and HTML versions of the Article.

Promoter capture Hi-C-based identification of recurrent noncoding mutations in colorectal cancer.

Efforts are being directed to systematically analyze the non-coding regions of the genome for cancer-driving mutations1-6. cis-regulatory elements (CREs) represent a highly enriched subset of the non-coding regions of the genome in which to search for such mutations. Here we use high-throughput chromosome conformation capture techniques (Hi-C) for 19,023 promoter fragments to catalog the regulatory landscape of colorectal cancer in cell lines, mapping CREs and integrating these with whole-genome sequence and expression data from The Cancer Genome Atlas7,8. We identify a recurrently mutated CRE interacting with the ETV1 promoter affecting gene expression. ETV1 expression influences cell viability and is associated with patient survival. We further refine our understanding of the regulatory effects of copy-number variations, showing that RASL11A is targeted by a previously identified enhancer amplification1. This study reveals new insights into the complex genetic alterations driving tumor development, providing a paradigm for employing chromosome conformation capture to decipher non-coding CREs relevant to cancer biology.

Identification of multiple risk loci and regulatory mechanisms influencing susceptibility to multiple myeloma.

Genome-wide association studies (GWAS) have transformed our understanding of susceptibility to multiple myeloma (MM), but much of the heritability remains unexplained. We report a new GWAS, a meta-analysis with previous GWAS and a replication series, totalling 9974 MM cases and 247,556 controls of European ancestry. Collectively, these data provide evidence for six new MM risk loci, bringing the total number to 23. Integration of information from gene expression, epigenetic profiling and in situ Hi-C data for the 23 risk loci implicate disruption of developmental transcriptional regulators as a basis of MM susceptibility, compatible with altered B-cell differentiation as a key mechanism. Dysregulation of autophagy/apoptosis and cell cycle signalling feature as recurrently perturbed pathways. Our findings provide further insight into the biological basis of MM.

Genome-wide association study implicates immune dysfunction in the development of Hodgkin lymphoma.

To further our understanding of inherited susceptibility to Hodgkin lymphoma (HL), we performed a meta-analysis of 7 genome-wide association studies totaling 5325 HL cases and 22 423 control patients. We identify 5 new HL risk loci at 6p21.31 (rs649775; P = 2.11 × 10-10), 6q23.3 (rs1002658; P = 2.97 × 10-8), 11q23.1 (rs7111520; P = 1.44 × 10-11), 16p11.2 (rs6565176; P = 4.00 × 10-8), and 20q13.12 (rs2425752; P = 2.01 × 10-8). Integration of gene expression, histone modification, and in situ promoter capture Hi-C data at the 5 new and 13 known risk loci implicates dysfunction of the germinal center reaction, disrupted T-cell differentiation and function, and constitutive NF-κB activation as mechanisms of predisposition. These data provide further insights into the genetic susceptibility and biology of HL.