ABSTRACT
Tumour mutational burden (TMB), defined as the total number of somatic non-synonymous mutations present within the cancer genome, varies across and within cancer types. A first wave of retrospective and prospective research identified TMB as a predictive biomarker of response to immune-checkpoint inhibitors and culminated in the disease-agnostic approval of pembrolizumab for patients with TMB-high tumours based on data from the Keynote-158 trial. Although the applicability of outcomes from this trial to all cancer types and the optimal thresholds for TMB are yet to be ascertained, research into TMB is advancing along three principal avenues: enhancement of TMB assessments through rigorous quality control measures within the laboratory process, including the mitigation of confounding factors such as limited panel scope and low tumour purity; refinement of the traditional TMB framework through the incorporation of innovative concepts such as clonal, persistent or HLA-corrected TMB, tumour neoantigen load and mutational signatures; and integration of TMB with established and emerging biomarkers such as PD-L1 expression, microsatellite instability, immune gene expression profiles and the tumour immune contexture. Given its pivotal functions in both the pathogenesis of cancer and the ability of the immune system to recognize tumours, a profound comprehension of the foundational principles and the continued evolution of TMB are of paramount relevance for the field of oncology.
ABSTRACT
BACKGROUND: Predictive biomarkers in use for immunotherapy in advanced non-small cell lung cancer are of limited sensitivity and specificity. We analysed the potential of activating KRAS and pathogenic TP53 mutations to provide additional predictive information. METHODS: The study cohort included 713 consecutive immunotherapy patients with advanced lung adenocarcinomas, negative for actionable genetic alterations. Additionally, two previously published immunotherapy and two surgical patient cohorts were analyzed. Therapy benefit was stratified by KRAS and TP53 mutations. Molecular characteristics underlying KRASmut/TP53mut tumours were revealed by the analysis of TCGA data. RESULTS: An interaction between KRAS and TP53 mutations was observed in univariate and multivariate analyses of overall survival (Hazard ratio [HR] = 0.56, p = 0.0044 and HR = 0.53, p = 0.0021) resulting in a stronger benefit for KRASmut/TP53mut tumours (HR = 0.71, CI 0.55-0.92). This observation was confirmed in immunotherapy cohorts but not observed in surgical cohorts. Tumour mutational burden, proliferation, and PD-L1 mRNA were significantly higher in TP53-mutated tumours, regardless of KRAS status. Genome-wide expression analysis revealed 64 genes, including CX3CL1 (fractalkine), as specific transcriptomic characteristic of KRASmut/TP53mut tumours. CONCLUSIONS: KRAS/TP53 co-mutation predicts ICI benefit in univariate and multivariate survival analyses and is associated with unique molecular tumour features. Mutation testing of the two genes can be easily implemented using small NGS panels.
Subject(s)
Adenocarcinoma of Lung , Immune Checkpoint Inhibitors , Lung Neoplasms , Mutation , Proto-Oncogene Proteins p21(ras) , Tumor Suppressor Protein p53 , Humans , Tumor Suppressor Protein p53/genetics , Proto-Oncogene Proteins p21(ras)/genetics , Female , Adenocarcinoma of Lung/genetics , Adenocarcinoma of Lung/drug therapy , Adenocarcinoma of Lung/immunology , Adenocarcinoma of Lung/pathology , Male , Immune Checkpoint Inhibitors/therapeutic use , Lung Neoplasms/genetics , Lung Neoplasms/drug therapy , Lung Neoplasms/pathology , Lung Neoplasms/immunology , Aged , Middle Aged , Biomarkers, Tumor/genetics , Immunotherapy/methods , Prognosis , Aged, 80 and over , Adult , Cohort StudiesABSTRACT
PURPOSE: The PI3K signaling pathway is frequently dysregulated in breast cancer (BC), and mutations in PIK3CA, are relevant for therapy resistance in HER2pos BC. Mutations in exons 9 or exon 20 may have different impact on response to neoadjuvant chemotherapy-based treatment regimens. EXPERIMENTAL DESIGN: We investigated PIK3CA mutations in 1691 early BC patients, randomized in four neoadjuvant multicenter trials: GeparQuattro (NCT00288002), GeparQuinto (NCT00567554), GeparSixto (NCT01426880) and GeparSepto (NCT01583426). The role of different PIK3CA exons and hotspots for pathological complete response (pCR) after neoadjuvant chemotherapy (NACT) and patient survival was evaluated for distinct molecular subgroups and anti-HER2 treatment procedures. RESULTS: A total of 302 patients (17.9%) of the full cohort of 1691 patients had a tumor with a PIK3CA mutation, with a different prevalence in molecular subgroups: luminal/HER2neg 95 of 404 patients (23.5%), HER2pos 170 of 819 patients (20.8%) and TNBC 37 of 431 patients (7.9%). We identified mutations in PIK3CA exon 20 to be linked with worse response to anti-HER2 treatment (OR=0.507, 95%CI 0.320-0.802, p=0.004), especially in HR positive HER2 positive BC (OR=0.445, 95%CI 0.237-0.837, p=0.012). In contrast, exon 9 hotspot mutations p.E452K and p.E545K revealed no noteworthy differences in response therapy response. Luminal/HER2neg patients show a trend to have worse treatment response when PIK3CA was mutated. Interestingly, patients with residual disease after neoadjuvant treatment, have better survival when PIK3CA was mutated. CONCLUSIONS: PIK3CA hotspot mutation p.H1047R are associated with worse pCR rates after NACT in HER2pos BC, while hotspot mutations in exon 9 seems to have less impact.
ABSTRACT
Targeted tumor only sequencing has become a standard practice in cancer diagnostics. This study aims to develop an approach for robust copy number variant calling in tumor samples using only off-target region (OTR) reads. We also established a clinical use case for homologous recombination deficiency (HRD) score estimation (HRDest) using the sum of telomeric-allelic imbalance and large-scale state transition scores without the need for loss of heterozygosity information. A strong correlation was found between HRD score and the sum of telomeric-allelic imbalance + large-scale state transition in The Cancer Genome Atlas cohort (ρ = 0.99, P < 2.2 × 10-16) and in a clinical in-house cohort of 34 tumors (ρ = 0.9, P = 5.1 × 10-13) comparing whole-exome sequencing and targeted sequencing data. HRDest scores from 1086 clinical cases were compared with The Cancer Genome Atlas data set. There were no significant differences in HRD score distribution within the analyzed tumor types. As a control, commercially available HRD standards were also sequenced, and the HRDest scores obtained from the OTR reads were well within the HRD reference range provided by the manufacturer. In conclusion, OTR reads of tumor-only panel sequencing can be used to determine genome-wide copy number variant profiles and to approximate HRD scores.
Subject(s)
DNA Copy Number Variations , Exome Sequencing , High-Throughput Nucleotide Sequencing , Neoplasms , Humans , Neoplasms/genetics , Exome Sequencing/methods , High-Throughput Nucleotide Sequencing/methods , Recombinational DNA Repair/genetics , Allelic ImbalanceABSTRACT
PURPOSE: Poly(ADP-ribose) polymerase inhibitors (PARPi) have shown promising clinical results in the treatment of ovarian cancer. Analysis of biomarker subgroups consistently revealed higher benefits for patients with homologous recombination deficiency (HRD). The test that is most often used for the detection of HRD in clinical studies is the Myriad myChoice assay. However, other assays can also be used to assess biomarkers, which are indicative of HRD, genomic instability (GI), and BRCA1/2 mutation status. Many of these assays have high potential to be broadly applied in clinical routine diagnostics in a time-effective decentralized manner. Here, we compare the performance of a multitude of alternative assays in comparison with Myriad myChoice in high-grade serous ovarian cancer (HGSOC). METHODS: DNA from HGSOC samples was extracted from formalin-fixed paraffin-embedded tissue blocks of cases previously run with the Myriad myChoice assay, and GI was measured by multiple molecular assays (CytoSNP, AmoyDx, Illumina TSO500 HRD, OncoScan, NOGGO GISv1, QIAseq HRD Panel and whole genome sequencing), applying different bioinformatics algorithms. RESULTS: Application of different assays to assess GI, including Myriad myChoice, revealed high concordance of the generated scores ranging from very substantial to nearly perfect fit, depending on the assay and bioinformatics pipelines applied. Interlaboratory comparison of assays also showed high concordance of GI scores. CONCLUSION: Assays for GI assessment not only show a high concordance with each other but also in correlation with Myriad myChoice. Thus, almost all of the assays included here can be used effectively to assess HRD-associated GI in the clinical setting. This is important as PARPi treatment on the basis of these tests is compliant with European Medicines Agency approvals, which are methodologically not test-bound.
Subject(s)
BRCA1 Protein , Ovarian Neoplasms , Humans , Female , BRCA1 Protein/genetics , Mutation , BRCA2 Protein/genetics , Ovarian Neoplasms/diagnosis , Ovarian Neoplasms/genetics , Ovarian Neoplasms/drug therapy , Genomic Instability/genetics , Homologous Recombination/geneticsABSTRACT
This study analyzed whether extended molecular profiling can predict the development of epidermal growth factor receptor (EGFR) gene T790M mutation, which is the most frequent resistance alteration in non-small cell lung cancer (NSCLC) after treatment with the first-/second-generation (1G/2G) EGFR inhibitors (tyrosine kinase inhibitors [TKIs]), but only weakly associated with clinical characteristics. Whole exome sequencing (WES) was performed on pretreatment tumor tissue with matched normal samples from NSCLC patients with (n = 25, detected in tissue or blood rebiopsies) or without (n = 14, negative tissue rebiopsies only) subsequent EGFR p.T790M mutation after treatment with 1G/2G EGFR TKI. Several complex genetic biomarkers were assessed using bioinformatic methods. After treatment with first-line afatinib (44%) or erlotinib/gefitinib (56%), median progression-free survival and overall survival were 12.1 and 33.7 months, respectively. Clinical and tumor genetic characteristics, including age (median, 66 years), sex (74% female), smoking (69% never/light smokers), EGFR mutation type (72% exon 19 deletions), and TP53 mutations (41%) were not significantly associated with T790M mutation (p > 0.05). By contrast, complex biomarkers including tumor mutational burden, the clock-like mutation signature SBS1 + 5, tumor ploidy, and markers of subclonality including mutant-allele tumor heterogeneity, subclonal copy number changes, and median tumor-adjusted variant allele frequency were significantly higher at baseline in tumors with subsequent T790M mutation (all p < 0.05). Each marker alone could predict subsequent development of T790M with an area under the curve (AUC) of 0.72-0.77, but the small number of cases did not allow confirmation of better performance for biomarker combinations in leave-one-out cross-validated logistic regression (AUC 0.69, 95% confidence interval: 0.50-0.87). Extended molecular profiling with WES at initial diagnosis reveals several complex biomarkers associated with subsequent development of T790M resistance mutation in NSCLC patients receiving first-/second-generation TKIs as the first-line therapy. Larger prospective studies will be necessary to define a forecasting model.
Subject(s)
Carcinoma, Non-Small-Cell Lung , Lung Neoplasms , Humans , Female , Aged , Male , ErbB Receptors/genetics , Lung Neoplasms/drug therapy , Lung Neoplasms/genetics , Carcinoma, Non-Small-Cell Lung/drug therapy , Carcinoma, Non-Small-Cell Lung/genetics , Prospective Studies , Mutation , Protein Kinase Inhibitors/therapeutic use , Genomics , BiomarkersABSTRACT
BACKGROUND: Regulation (EU) 2017/746 on in vitro diagnostic medical devices (IVDR) imposes several conditions on pathology departments that develop and use in-house in vitro diagnostic medical devices (IH-IVDs). However, not all of these conditions need to be implemented immediately after the IVDR entered into force on 26 May 2022. Based on an amending regulation of the European Parliament and the Council of the European Union, the requirements for IH-IVDs will be phased in. Conformity with the essential safety and performance requirements of annex I must be ensured from May 2022. OBJECTIVES: With this article, we would like to present the practical implementation of the currently valid conditions for IH-IVDs at the Institute of Pathology at the University Hospital of Heidelberg, in order to provide possible assistance to other institutions. CONCLUSIONS: In addition to the intensive work on the requirements for IH-IVDs, several guidance documents and handouts provide orientation for the implementation and harmonisation of the requirements for healthcare institutions mentioned in Article 5 (5). Exchange in academic network structures is also of great importance for the interpretation and practical implementation of the IVDR. For university and nonuniversity institutions, ensuring conformity with the IVDR represents a further challenge in terms of personnel and time, in addition to the essential tasks of patient care, teaching and research and the further development of methods for optimal and targeted diagnostics, as well as the maintenance of the constantly evolving quality management system.
Subject(s)
Reagent Kits, Diagnostic , Humans , European UnionABSTRACT
BACKGROUND: Regulation (EU) 2017/746 on in vitro diagnostic medical devices (IVDR) imposes several conditions on pathology institutes that develop and use in-house in vitro diagnostic medical devices (IH-IVDs). However, not all of these conditions need to be implemented immediately after the IVDR entered into force on 26 May 2022. Based on an amending regulation of the European Parliament and the Council of the European Union, the requirements for IH-IVDs will be phased in. Conformity with the essential safety and performance requirements of annex I must be ensured from May 2022. OBJECTIVES: With this article, we would like to present the practical implementation of the currently valid conditions for IH-IVDs at the Institute of Pathology at the University Hospital of Heidelberg, in order to provide possible assistance to other institutions. CONCLUSIONS: In addition to the intensive work on the requirements for IH-IVDs, several guidance documents and handouts provide orientation for the implementation and harmonisation of the requirements for healthcare institutions mentioned in Article 5 (5). Exchange in academic network structures is also of great importance for the interpretation and practical implementation of the IVDR. For university and nonuniversity institutions, ensuring conformity with the IVDR represents a further challenge in terms of personnel and time, in addition to the essential tasks of patient care, teaching and research and the further development of methods for optimal and targeted diagnostics, as well as the maintenance of the constantly evolving quality management system.
Subject(s)
Reagent Kits, Diagnostic , Humans , European UnionABSTRACT
A growing number of druggable targets and national initiatives for precision oncology necessitate broad genomic profiling for many cancer patients. Whole exome sequencing (WES) offers unbiased analysis of the entire coding sequence, segmentation-based detection of copy number alterations (CNAs), and accurate determination of complex biomarkers including tumor mutational burden (TMB), homologous recombination repair deficiency (HRD), and microsatellite instability (MSI). To assess the inter-institution variability of clinical WES, we performed a comparative pilot study between German Centers of Personalized Medicine (ZPMs) from five participating institutions. Tumor and matched normal DNA from 30 patients were analyzed using custom sequencing protocols and bioinformatic pipelines. Calling of somatic variants was highly concordant with a positive percentage agreement (PPA) between 91 and 95% and a positive predictive value (PPV) between 82 and 95% compared with a three-institution consensus and full agreement for 16 of 17 druggable targets. Explanations for deviations included low VAF or coverage, differing annotations, and different filter protocols. CNAs showed overall agreement in 76% for the genomic sequence with high wet-lab variability. Complex biomarkers correlated strongly between institutions (HRD: 0.79-1, TMB: 0.97-0.99) and all institutions agreed on microsatellite instability. This study will contribute to the development of quality control frameworks for comprehensive genomic profiling and sheds light onto parameters that require stringent standardization.
ABSTRACT
Molecular pathological diagnostics plays a central role in personalized oncology and requires multidisciplinary teamwork. It is just as relevant for the individual patient who is being treated with an approved therapy method or an individual treatment attempt as it is for prospective clinical studies that require the identification of specific therapeutic target structures or complex biomarkers for study inclusion. It is also of crucial importance for the generation of real-world data, which is becoming increasingly important for drug development. Future developments will be significantly shaped by improvements in scalable molecular diagnostics, in which increasingly complex and multi-layered data sets must be quickly converted into clinically useful information. One focus will be on the development of adaptive diagnostic strategies in order to be able to depict the enormous plasticity of a cancer disease over time.
Subject(s)
Neoplasms , Precision Medicine , Humans , Medical Oncology/methods , Neoplasms/diagnosis , Neoplasms/genetics , Neoplasms/therapy , Pathology, Molecular/methods , Precision Medicine/methods , Prospective StudiesABSTRACT
OBJECTIVES: Molecular diagnosis for targeted therapies has been improved significantly in non-small-cell lung cancer (NSCLC) patients in recent years. Here we report on the prevalence of rare fusions in NSCLC and dissect their genomic architecture and potential clinical implications. MATERIALS AND METHODS: Overall, n = 5554 NSCLC patients underwent next-generation sequencing (NGS) for combined detection of oncogenic mutations and fusions either at primary diagnosis (n = 5246) or after therapy resistance (n = 308). Panels of different sizes were employed with closed amplicon-based, or open assays, i.e. anchored multiplex PCR (AMP) and hybrid capture-based, for detection of translocations, including "rare" fusions, defined as those beyond ALK, ROS1, RET and <0.5 % frequency in NSCLC. RESULTS: Rare fusions involving EGFR, MET, HER2, BRAF and other potentially actionable oncogenes were detected in 0.5% (n = 26) of therapy-naive and 2% (n = 6) TKI-treated tumors. Detection was increased using open assays and/or larger panels, especially those covering >25 genes, by approximately 1-2% (p = 0.001 for both). Patient characteristics (age, gender, smoking, TP53 co-mutations (56%), or mean tumor mutational burden (TMB) (4.8 mut/Mb)) showed no association with presence of rare fusions. Non-functional alterations, i.e. out-of-frame or lacking kinase domains, comprised one-third of detected rare fusions and were significantly associated with simultaneous presence of classical oncogenic drivers, e.g. EGFR or KRAS mutations (p < 0.001), or use of larger panels (frequency of non-functional among the detected rare fusions 57% for 25+ gene- vs. 12% for smaller panels, p < 0.001). As many rare fusions were identified before availability of targeted therapy, mean survival for therapy-naïve patients was 23.8 months, comparable with wild-type tumors. CONCLUSION: Approximately 1-2% of advanced NSCLC harbor rare fusions, which are potentially actionable and may support diagnosis. Routine adoption of broad NGS assays capable to identify exact fusion points and potentially retained protein domains can increase the yield of therapeutically relevant molecular information in advanced NSCLC.
Subject(s)
Carcinoma, Non-Small-Cell Lung , Lung Neoplasms , Humans , Carcinoma, Non-Small-Cell Lung/drug therapy , Lung Neoplasms/pathology , Protein-Tyrosine Kinases/genetics , Proto-Oncogene Proteins/genetics , Genomics , Mutation , ErbB Receptors/geneticsABSTRACT
Homologous recombination deficiency (HRD) is a predictive marker for response to poly (ADP-ribose) polymerase inhibitors (PARPi) in ovarian carcinoma. HRD scores have entered routine diagnostics, but the influence of algorithms, parameters and confounders has not been analyzed comprehensively. A series of 100 poorly differentiated ovarian carcinoma samples was analyzed using whole exome sequencing (WES) and genotyping. Tumor purity was determined using conventional pathology, digital pathology, and two bioinformatic methods. HRD scores were calculated from copy number profiles determined by Sequenza and by Sclust either with or without fixed tumor purity. Tumor purity determination by digital pathology combined with a tumory purity informed variant of Sequenza served as reference method for HRD scoring. Seven tumors had deleterious mutations in BRCA1/2, 12 tumors had deleterious mutations in other homologous recombination repair (HRR) genes, 18 tumors had variants of unknown significance (VUS) in BRCA1/2 or other HRR genes, while the remaining 63 tumors had no relevant alterations. Using the reference method for HRD scoring, 68 tumors were HRD-positive. HRDsum determined by WES correlated strongly with HRDsum determined by single nucleotide polymorphism (SNP) arrays (R = 0.85). Conventional pathology systematically overestimated tumor purity by 8% compared to digital pathology. All investigated methods agreed on classifying the deleterious BRCA1/2-mutated tumors as HRD-positive, but discrepancies were observed for some of the remaining tumors. Discordant HRD classification of 11% of the tumors was observed comparing the tumor purity uninformed default of Sequenza and the reference method. In conclusion, tumor purity is a critical factor for the determination of HRD scores. Assistance by digital pathology helps to improve accuracy and imprecision of its estimation.
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OBJECTIVE: Intratumoral heterogeneity was found to be a significant factor causing resistance to lung cancer therapies, including immune checkpoint blockade. Lesser is known about spatial heterogeneity of the tumor microenvironment (TME) and its association with genetic properties of the tumor, which is of particular interest in the therapy-naïve setting. MATERIALS AND METHODS: We performed multi-region sampling (2-4 samples per tumor; total of 55 samples) from a cohort of 19 untreated stage IA-IIIB lung adenocarcinomas (n = 11 KRAS mutant, n = 1 ERBB2 mutant, n = 7 KRAS wildtype). For each sample the expression of 770 immunooncology-related genes was analyzed using the nCounter platform, while the mutational status was determined by hybrid capture-based next-generation sequencing (NGS) using a large panel covering more than 500 genes. RESULTS: Global unsupervised analyses revealed clustering of the samples into two groups corresponding to a 'hot' or 'cold' immunologic tumor contexture based on the abundance of immune cell infiltrates. All analyzed specific immune cell signatures (ICsig) showed a significantly higher intertumoral than intratumoral heterogeneity (p < 0.02), as most of the analyzed cases (14/19) showed a very homogenous spatial immune cell profile. PD-L1 exhibited a significantly higher intertumoral than intratumoral heterogeneity (p = 1.03e-13). We found a specific association with 'cold' TME for STK11 (11/14, p < 0.07), but not KRAS, TP53, LRP1B, MTOR, U2AF1 co-mutations, and validated this finding using The Cancer Genome Atlas (TCGA) data. CONCLUSION: Early-stage lung adenocarcinomas show considerable intertumoral, but limited intratumoral heterogeneity, which is clinically highly relevant as assessment before neoadjuvant treatment is based on small biopsies. STK11 mutations are specifically associated with a 'cold' TME, which could affect the efficacy of perioperative immunotherapy.
Subject(s)
AMP-Activated Protein Kinase Kinases , Adenocarcinoma of Lung , Immune Evasion , Lung Neoplasms , Adenocarcinoma of Lung/genetics , Adenocarcinoma of Lung/immunology , Adenocarcinoma of Lung/therapy , Lung Neoplasms/genetics , Lung Neoplasms/immunology , Lung Neoplasms/therapy , Immune Evasion/genetics , AMP-Activated Protein Kinase Kinases/genetics , Tumor Microenvironment/genetics , Tumor Microenvironment/immunology , Humans , Mutation , Neoplasm StagingABSTRACT
PURPOSE: Tumor microenvironment (TME) immune markers have been correlated with both response to neoadjuvant therapy and prognosis in patients with breast cancer. Here, immune-cell activity of breast cancer tumors was inferred by expression-based analysis to determine if it is prognostic and/or predictive of response to neoadjuvant paclitaxel-based therapy in the GeparSepto (G7) trial (NCT01583426). EXPERIMENTAL DESIGN: Pre-study biopsies from 279 patients with HER2-negative breast cancer in the G7 trial underwent RNA-seq-based profiling of 104 immune-cell-specific genes to assess inferred Immune Cell Activity (iICA) of 23 immune-cell types. Hierarchical clustering was used to classify tumors as iICA "hot," "warm," or "cold" by comparison of iICA in the G7 cohort relative to that of 1,467 samples from a tumor database established by Nantomics LLC. Correlations between iICA cluster, pathology-assessed tumor-infiltrating lymphocytes (TIL), and hormone receptor (HR) status for pathologic complete response (pCR), disease-free survival (DFS), and overall survival (OS) were determined. RESULTS: iICA cluster correlated with TIL levels. The highest pCR rates were observed in hot cluster tumors, and those with relatively higher TILs. Greater inferred activity of several T-cell types was significantly associated with pCR and survival. DFS and OS were prolonged in patients with hot or warm cluster tumors, the latter particularly for HR negative tumors, even if TILs were relatively low. CONCLUSIONS: Overall, TIL level better predicted pCR, but iICA cluster better predicted survival. Differences in associations between TILs, cluster, pCR, and survival were observed for HR-positive tumors versus HR-negative tumors, suggesting expanded study of the implication of these findings is warranted.
Subject(s)
Breast Neoplasms , Humans , Female , Breast Neoplasms/drug therapy , Breast Neoplasms/genetics , Breast Neoplasms/metabolism , Paclitaxel/therapeutic use , Prognosis , Lymphocytes, Tumor-Infiltrating , Disease-Free Survival , Neoadjuvant Therapy , Receptor, ErbB-2/metabolism , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Tumor Microenvironment/geneticsABSTRACT
TP53 is the most frequently mutated gene in human cancer. While no TP53-targeting drugs have been approved in the USA or Europe so far, preclinical and clinical studies are underway to investigate targeting of specific or all TP53 mutations, for example, by restoration of the functionality of mutated TP53 (TP53mut) or protecting wildtype TP53 (TP53wt) from negative regulation. We performed a comprehensive mRNA expression analysis in 24 cancer types of TCGA to extract (i) a consensus expression signature shared across TP53 mutation types and cancer types, (ii) differential gene expression patterns between tumors harboring different TP53 mutation types such as loss of function, gain of function or dominant-negative mutations, and (iii) cancer-type-specific patterns of gene expression and immune infiltration. Analysis of mutational hotspots revealed both similarities across cancer types and cancer type-specific hotspots. Underlying ubiquitous and cancer type-specific mutational processes with the associated mutational signatures contributed to explaining this observation. Virtually no genes were differentially expressed between tumors harboring different TP53 mutation types, while hundreds of genes were over- and underexpressed in TP53mut compared to TP53wt tumors. A consensus list included 178 genes that were overexpressed and 32 genes that were underexpressed in the TP53mut tumors of at least 16 of the investigated 24 cancer types. In an association analysis of immune infiltration with TP53 mutations in 32 cancer subtypes, decreased immune infiltration was observed in six subtypes, increased infiltration in two subtypes, a mixed pattern of decreased and increased immune cell populations in four subtypes, while immune infiltration was not associated with TP53 status in 20 subtypes. The analysis of a large cohort of human tumors complements results from experimental studies and supports the view that TP53 mutations should be further evaluated as predictive markers for immunotherapy and targeted therapies.
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BACKGROUND: The prognostic significance of tumour budding (TB) and minimal cell nest size (MCNS) was shown in human papillomavirus (HPV)-negative head and neck squamous cell carcinomas (HNSCC). However, the optimisation of cutpoints, the prognostic impact in HPV-positive HNSCC, and the comparison with other histopathological grading systems are insufficiently investigated. METHODS: TB and MCNS were analysed digitally in 1 and 10 high-power fields (HPF) of 331 HPV-positive and HPV-negative cases from TCGA. Optimising the cutpoints a new cellular dissociation grading (CDG) system was defined and compared to the WHO grading and the Brandwein-Gensler (BG) risk model. RESULTS: The two-tiered CDG system based solely on TB yielded optimal prognostic stratification with shortened overall survival for CDG-high cases. Optimal cut-offs were two buds (1 HPF) and six buds (10 HPF), respectively. Analysing MCNS did not add prognostic significance to quantifying TB. CDG was a significant prognostic marker in HPV-negative and HPV-positive tumours and prognostically superior to the WHO and BG systems. High CDG was associated with clinically occult lymph-node metastases. CONCLUSIONS: The most comprehensive study of TB in HNSCC so far confirmed its prognostic impact in HPV-negative tumours and for the first time in HPV-positive tumours. Further studies are warranted to evaluate its applicability for therapy guidance in HNSCC.
Subject(s)
Carcinoma, Squamous Cell , Head and Neck Neoplasms , Papillomavirus Infections , Humans , Squamous Cell Carcinoma of Head and Neck , Carcinoma, Squamous Cell/pathology , Prognosis , Papillomavirus Infections/complications , Papillomaviridae , BiomarkersABSTRACT
BACKGROUND: Regulation (EU) 2017/746 on in vitro diagnostic medical devices (IVDR) was passed by the European Parliament and the Council of the European Union on 5 April 2017 and came into force on 26 May 2017. A new amending regulation, which introduces a phased implementation of the IVDR with new transitional provisions for certain in vitro diagnostic medical devices (IVDs) and a later date of application of some requirements for in-house devices for healthcare facilities, was adopted on 15 December 2021. The combined use of CE-certified IVDs (CE-IVDs), in-house IVDs (IH-IVDs), and research use only (RUO) devices are a cornerstone of diagnostics in pathology departments and crucial for optimal patient care. The IVDR not only regulates the manufacture and placement on the market of industrially manufactured IVDs, but also imposes conditions on the manufacture and use of IH-IVDs for internal use by healthcare facilities. OBJECTIVES: Our work provides an overview of the background and structure of the IVDR and identifies core areas that need to be interpreted and fleshed out in the context of the legal framework as well as expert knowledge. CONCLUSIONS: The gaps and ambiguities in the IVDR crucially require the expertise of professional societies, alliances, and individual stakeholders to successfully facilitate the implementation and use of the IVDR in pathology departments and to avoid aberrant developments.
Subject(s)
Commerce , Reagent Kits, Diagnostic , Humans , European Union , Health FacilitiesABSTRACT
Leveraging real-world data (RWD) for drug access is necessary to overcome a key challenge of modern precision oncology: tackling numerous low-prevalence oncogenic mutations across cancers. Withholding a potentially active medication in patients with rare mutations for the sake of control chemotherapy or "best" supportive care is neither practicable nor ethically justifiable anymore, particularly as RWD could meanwhile be used instead, according to scientific principles outlined by the US Food and Drug Administration, European Medicines Agency and other stakeholders. However, practical implementation varies, with occasionally opposite recommendations based on the same evidence in different countries. In the face of growing need for precision drugs, more transparency of evaluation, a priori availability of guidance for the academia and industry, as well as a harmonized framework for health technology assessment across the European Union (EU) are imperative. These could in turn trigger infrastructural changes in national and pan-European registries, cancer management guidelines (e.g., frequency of routine radiologic restaging, inclusion of patient-reported outcomes), and the health data space, to ensure conformity with declared standards and facilitate extraction of RWD sets (including patient-level data) suitable for approval and pricing with minimal effort. For an EU-wide unification of precision cancer medicine, collective negotiation of drug supply contracts and funding solidarity would additionally be required to handle the financial burden. According to experience from pivotal European programs, off-label use could potentially also be harmonized across EU-states to accelerate availability of novel drugs, streamline collection of valuable RWD, and mitigate related costs through wider partnerships with pharmaceutical companies.