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Immunotherapy has become the standard-of-care in many solid tumors. Despite the significant recent achievements in the diagnosis and treatment of cancer, several issues related to patients’ selection for immunotherapy remain unsolved. Multiple lines of evidence suggest that, in this setting, the vision of a single biomarker is somewhat naïve and imprecise, given that immunotherapy does not follow the rules that we have experienced in the past for targeted therapies. On the other hand, additional immune-related biomarkers that are reliable in real-life clinical practice remain to be identified. Recently, the immune-checkpoint blockade has been approved in the US irrespective of the tumor site of origin. Further histology-agnostic approvals, coupled with with tumor-specific companion diagnostics and guidelines, are expected in this field. In addition, immune-related biomarkers can also have a significant prognostic value. In this review, we provide an overview of the role of these biomarkers and their characterization in the management of lung cancer, melanoma, colorectal cancer, gastric cancer, head and neck cancer, renal cell carcinoma, urothelial cancers, and breast cancer.

Immune Checkpoint Inhibitors (ICIs) have remarkably modified the way solid tumors are managed, including breast cancer. Unfortunately, only a relatively small number of breast cancer patients significantly respond to these treatments. To maximize the immunotherapy benefit in breast cancer, several efforts are currently being put forward for the identification of i) the best therapeutic strategy (i.e. ICI monotherapy or in association with chemotherapy, radiotherapy, or other drugs); ii) optimal timing for administration (e.g. early/advanced stage of disease; adjuvant/ neoadjuvant setting); iii) most effective and reliable predictive biomarkers of response (e.g. tumor-infiltrating lymphocytes, programmed death-ligand 1, microsatellite instability associated with mismatch repair deficiency, and tumor mutational burden). In this article, we review the impacts and gaps in the characterization of immune-related biomarkers raised by clinical and translational research studies with immunotherapy treatments. Particular emphasis has been put on the documented evidence of significant clinical benefits of ICI in different randomized clinical trials, along with preanalytical and analytical issues in predictive biomarkers pathological assessment.

IntroductionBRCA tumour testing is a crucial tool for personalised therapy of patients with ovarian cancer. Since different next-generation sequencing (NGS) platforms and BRCA panels are available, the NGS Italian Network proposed to assess the robustness of different technologies.MethodsSix centres, using four different technologies, provided raw data of 284 cases, including 75 cases with pathogenic/likely pathogenic variants, for a revision blindly performed by an external bioinformatic platform.ResultsThe third-party revision assessed that all the 284 raw data reached good quality parameters. The variant calling analysis confirmed all the 75 pathogenic/likely pathogenic variants, including challenging variants, achieving a concordance rate of 100% regardless of the panel, instrument and bioinformatic pipeline adopted. No additional variants were identified in the reanalysis of a subset of 41 cases.ConclusionsBRCA tumour testing performed with different technologies in different centres, may achieve the realibility and reproducibility required for clinical diagnostic procedures.

The advances in understanding the inherited biological mechanisms of non-small cell lung cancer harbouring epidermal growth factor receptor (EGFR) mutations led to a significant improvement in the outcomes of patients treated with EGFR tyrosine kinase inhibitors. Despite these clinically impressive results, clinical results are not always uniform, suggesting the need for deepening the molecular heterogeneity of this molecularly defined subgroup of patients beyond the clinical and biological surface. The availability of tissue and blood-based tumour genotyping allows us to improve the understanding of molecular and genetic intratumor heterogeneity, driving the measurement of clonal evaluation in patients with lung cancer carrying EGFR mutations. Genetic diversification, clonal expansion and selection are highly variable patterns of genetic diversity, resulting in different biological entities, also a prerequisite for Darwinian selection and therapeutic failure. Such emerging pieces of evidence on the genetic diversity, including adaptive and immunomodulated aspects, provide further evidence for the role of the tumour microenvironment (TME) in drug-resistance and immune-mediated mechanisms. Matching in daily clinical practice, the detailed genomic profile of lung cancer disease and tracking the clonal evolution could be the way to individualise the further target treatments in EGFR-positive disease. Characterising the tumour and immune microenvironment during the time of the cancer evaluation could be the way forward for the qualitative leap needed from bench to bedside. Such a daring approach, aiming at personalising treatment selection in order to exploit the TME properties and weaken tumour adaptivity, should be integrated into clinical trial design to optimise patient outcome.

BackgroundWith the approval of the poly (ADP-ribose) polymerase (PARP) inhibitor olaparib for newly diagnosed, breast cancer gene (BRCA)1/2 mutated, ovarian cancer women, the assessment of BRCA1/2 tumour status will be shortly required at the time of diagnosis.AimTo investigate the feasibility of next-generation sequencing (NGS)-based BRCA tumour test on cytological specimens from ovarian cancer ascites.MethodsWe evaluated the BRCA1/2 status on neoplastic ascites and corresponding tumour tissue of 11 patients with ovarian cancer, using the NGS ‘Oncomine BRCA Research Assay’.ResultsThe NGS-based BRCA test on cytological samples had a success rate of 100%, with 11 of 11 concordant BRCA1/2 results between ascites and tumour tissues analyses, including two wild type samples and nine cases harbouring somatic or germline variants.ConclusionBRCA test may be performed on ovarian cancer ascites, reproducing BRCA1/2 tumour status and representing a useful tool for clinical decision-making.

Background: Temozolomide (TEM) is an active treatment in metastatic neuroendocrine tumors (NETs). Patients affected by glioblastoma multiforme or advanced melanoma treated with TEM who have deficiency of O6-methylguanine DNA methyltransferase (MGMT) have a better responses and survival. However, the predictive role of MGMT in patients with NETs treated with TEM is still debated. Methods: We conducted a systematic review of the literature and meta-analysis, based on PRISMA methodology, searching in the main databases (PubMed, Embase, Scopus, Web of Science, Cochrane Library and clinical trial.gov) and the proceedings of the main international congresses, until April 26, 2021. Results: Twelve out of 616 articles were selected for our analysis, regarding a total of 858 NET patients treated with TEM-based chemotherapy. The status of MGMT had been tested in 513 (60%) patients, using various methods. The pooled overall response rate (ORR) was higher in MGMT-deficient compared with MGMT-proficient NETs, with a risk difference of 0.31 (95% confidence interval, CI: 0.13–0.50; p < 0.001; I2: 73%) and risk ratio of 2.29 (95% CI: 1.34–3.91; p < 0.001; I2: 55%). The pooled progression free survival (PFS) (hazard ratio, HR = 0.56; 95% CI: 0.43–0.74; p < 0.001) and overall survival (OS) (HR = 0.41; 95% CI: 0.20–0.62; p = 0.011) were longer in MGMT-deficient versus MGMT-proficient NETs. Conclusions: Our meta-analysis suggested that MGMT status may be predictive of TEM efficacy. However, due to the high heterogeneity of the evaluated studies the risk of biases should be considered. On this hypothesis future homogeneous prospective studies are warranted.

The PARP inhibitor olaparib has been approved in the maintenance setting of platinum-sensitive epithelial ovarian cancer patients with germline or somatic BRCA1/2 mutation. Therefore, the availability of a tumor BRCA test has become a clinical need. We report the results of the clinical implementation of a tumor BRCA test within the frame of an institutional workflow for the management of patients with nonmucinous and nonborderline epithelial ovarian cancer. In total, 223 patients with epithelial ovarian cancer were prospectively analyzed. BRCA1/2 status was evaluated on formalin-fixed, paraffin-embedded tumor specimens using next-generation sequencing technology. The tumor BRCA test had a success rate of 99.1% (221 of 223 successfully analyzed cases) and a median turnaround time of 17 calendar days. Among the 221 cases, BRCA1 or BRCA2 pathogenic/likely pathogenic mutations were found in 62 (28.1%) cases and variants of uncertain significance in 25 (11.3%) cases. The concordance rate between tumor BRCA test results and germline BRCA1/2 status was 87%, with five cases harboring pathogenic/likely pathogenic somatic-only mutations. The next-generation, sequencing-based tumor BRCA test showed a high success rate and a turnaround time compatible with clinical purposes. The tumor BRCA test could be implemented in a molecular diagnostic setting and it may guide the clinical management of patients with epithelial ovarian cancer.

Introduction Among the several agents targeting the programmed cell death 1 (PD-1) pathway, pembrolizumab is currently the only one approved for the treatment of patients with NSCLC in association with a companion diagnostic assay, the anti–PD-L1 immunohistochemical (IHC) 22C3 PharmDx (Agilent Technologies, Santa Clara, CA) using the Dako Autostainer (Dako, Carpinteria, CA). However, the Dako platform is not present in each pathology department, and this technical limitation is a major problem for the diffusion of the PD-L1 IHC predictive test for pembrolizumab. Methods The Italian Society of Anatomic Pathology and Cytopathology and the Italian Association of Medical Oncology in an independent, multicenter study compared the in vitro diagnostics PD-L1 IHC 22C3 pharmDx test (Agilent) on the Dako Autostainer and the in vitro diagnostics Ventana PD-L1 (SP263) test on the Ventana BenchMark platform (Ventana Medical Systems, Tucson, AZ). Using serial sections from tissue microarrays, 100 lung adenocarcinomas were locally stained and scored in four centers with the same antibody batches. Results A high analytical correlation (more than 90% at the lower 95% confidence interval [CI] value) between PD-L1 expression levels obtained with the 22C3 and SP263 assays was observed. At the proposed clinically relevant cutoffs (≥50% and ≥1%), the overall concordances between 22C3 and SP263 data were 0.99 (95% CI: 0.96–1) and 0.80 (95% CI: 0.68–0.91), respectively. The lower agreement between data obtained with the 22C3 and SP263 clones at the cutoff of 1% or higher was mainly related to the lower (about 80%) interrater agreement at this cutoff with each clone. Conclusions These results indicate a high correlation between PD-L1 IHC expression data obtained with the Agilent PD-L1 IHC 22C3 pharmDx and the Ventana PD-L1 (SP263) tests in NSCLC and suggest that the two assays could be utilized interchangeably as an aid to select patients for first-line and second-line treatment with pembrolizumab and potentially with other anti–PD-1/PD-L1 checkpoint inhibitors.

Aims: Pulmonary adenofibromas are rare benign fibroepithelial tumours of the lung with unknown histogenesis and an indolent clinical behaviour. Their stroma resembles that of solitary fibrous tumours, whereas the glands are composed of respiratory epithelium organized in a phyllodes-like architecture. Differentiation of pulmonary adenofibromas from other more aggressive intrathoracic tumours is clinically relevant. However, their biology is unknown. Here, we sought to characterize pulmonary adenofibromas at a clinicopathological level and to define whether they could be underpinned by a highly recurrent somatic genetic alteration akin to tumours with similar morphology. Methods and results: Seven pulmonary adenofibromas were subjected to immunohistochemical analysis for thyroid transcription factor 1 (TTF1), napsin A, cytokeratin 7, E-cadherin, CD99, CD34, CD31, STAT6, oestrogen receptor (ER), progesterone receptor, androgen receptor, bcl-2, and vimentin, as well as electron microscopy and capillary sequencing on microdissected samples to evaluate the presence of NAB2–STAT6 fusion genes and MED12 exon 2 mutations in their discrete components. A control group comprising pulmonary solitary fibrous tumours, pulmonary hamartomas and breast fibroadenomas was also analysed. We confirmed that the stromal elements of pulmonary adenofibromas pertain to the fibroblastic lineage, and show ER overexpression in 71% of cases, whereas the epithelium consists of TTF1-positive, E-cadherin positive bronchiolar elements. A highly recurrent NAB2–STAT6 fusion variant (exon 4–exon 2) was detected in the stroma but not in the epithelium. No MED12 mutations were identified. Conclusions: Here, we demonstrate that pulmonary adenofibromas are neoplastic lesions harbouring the molecular hallmark of solitary fibrous tumours