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Abstract Background The KEYNOTE-048 trial showed that pembrolizumab-based first-line treatment for R/M HNSCC led to improved OS in the PD-L1 CPS ≥ 1 population when compared to the EXTREME regimen. However, the R/M HNSCC real-world population is generally frailer, often presenting with multiple comorbidities, worse performance status and older age than the population included in phase III clinical trials. Methods This is a retrospective, single-centre analysis of patients with R/M HNSCC treated with pembrolizumab-based first-line treatment. Results From February 2021 to March 2023, 92 patients were treated with pembrolizumab-based first-line treatment. Patients treated with pembrolizumab-based chemoimmunotherapy had better ECOG PS and younger age than those treated with pembrolizumab monotherapy. Median PFS and OS were 4 months and 8 months, respectively. PFS was similar among patients treated with pembrolizumab-based chemoimmunotherapy and pembrolizumab monotherapy, while patients treated with pembrolizumab monotherapy had worse OS (log-rank p =.001, HR 2.7). PFS and OS were improved in patients with PD-L1 CPS > = 20 (PFS: log-rank p =.005, HR 0.50; OS: log-rank p =.04, HR 0.57). Patients with higher ECOG PS scores had worse PFS and OS (PFS, log-rank p =.004; OS, log-rank p = 6e-04). In multivariable analysis, ECOG PS2 was associated with worse PFS and OS. Conclusions PFS in our real-world cohort was similar to the KEYNOTE-048 reference while OS was numerically inferior. A deeper understanding of clinical variables that might affect survival outcomes of patients with R/M HNSCC beyond ECOG PS and PD-L1 CPS is urgently needed.

Effective risk assessment in early breast cancer is essential for informed clinical decision-making, yet consensus on defining risk categories remains challenging. This paper explores evolving approaches in risk stratification, encompassing histopathological, immunohistochemical, and molecular biomarkers alongside cutting-edge artificial intelligence (AI) techniques. Leveraging machine learning, deep learning, and convolutional neural networks, AI is reshaping predictive algorithms for recurrence risk, thereby revolutionizing diagnostic accuracy and treatment planning. Beyond detection, AI applications extend to histological subtyping, grading, lymph node assessment, and molecular feature identification, fostering personalized therapy decisions. With rising cancer rates, it is crucial to implement AI to accelerate breakthroughs in clinical practice, benefiting both patients and healthcare providers. However, it is important to recognize that while AI offers powerful automation and analysis tools, it lacks the nuanced understanding, clinical context, and ethical considerations inherent to human pathologists in patient care. Hence, the successful integration of AI into clinical practice demands collaborative efforts between medical experts and computational pathologists to optimize patient outcomes.

Abstract Background Breast cancer (BC) includes different pathological and molecular subtypes. This study aimed to investigate whether multiparametric magnetic resonance imaging (mpMRI) could reliably predict the molecular status of BC, comparing mpMRI features with pathological and immunohistochemical results. Methods This retrospective study included 156 patients with an ultrasound-guided biopsy-proven BC, who underwent breast mpMRI (including diffusion-weighted imaging) on a 3-T scanner from 2017 to 2020. Histopathological analyses were performed on the surgical specimens. Kolmogorov–Smirnov Z, χ 2, and univariate and multivariate logistic regression analyses were performed. Results Fifteen patients were affected with ductal carcinoma in situ, 122 by invasive carcinoma of no special type, and 19 with invasive lobular carcinoma. Out of a total of 141 invasive cancers, 45 were luminal A-like, 54 luminal B-like, 5 human epidermal growth factor receptor 2 (HER2) positive, and 37 triple negative. The regression analyses showed that size

Renal involvement is a common occurrence in patients with immuno-rheumatological diseases (IRDs). Several instances of glomerulonephritis (GN) occur in the setting of IRD and complicate the clinical course of an underlying condition. The aim of this study was to observe the spectrum of nephropathies according to age, kidney function, history of IRD at the time of biopsy, and histopathological kidney diagnosis. We evaluated data relating to 699 consecutive kidney native biopsies (female 52.1%) with a median age of 48 years (IQR 34–62) performed in adult patients collected over 15 years. The study population was divided into three groups: patients with kidney histological findings correlated to underlying IRD (Group 1), patients with kidney histological findings not correlated to underlying IRD (Group 2), and patients with kidney histological findings compatible with “de novo” IRD (absent in personal medical history) (Group 3). Kidney involvement related to IRD was found in 25.2% of patients. Group 1 was mostly represented by lupus nephritis (76.6%), with a younger age than Group 3 (p < 0.001) and by a higher percentage of females than other groups (p < 0.001). Group 3 was the most represented by microscopic polyangiitis (50.8%) when compared with the other two groups (p < 0.001). Acute nephritic syndrome (p < 0.001), acute kidney injury (AKI), and abnormal urinalysis (p < 0.001) were more represented in Group 3 than the other groups. In conclusion, IRDs are characterized by different clinical presentations and heterogeneous histological findings. Kidney biopsy remains fundamental to achieving the correct diagnosis and starting targeted therapy.

Background: Microvascular disorders represent an uncommon site of tissue hypo-perfusion and damage. Various genetic and acquired causes can be involved. A 65-year-old man was admitted because of refractory angina, which he had had since the age of 30 years, micro-hematuria, and recurrent transitory ischemic attacks from the age of 64. Methods: Hematochemical studies, ECG, Holter monitoring, 2D-echo, cardiac magnetic resonance (CMR), CTA of cerebral vessels, endomyocardial coronary angiography, and kidney biopsy processes were undertaken. Gene mutation analysis was conducted using next-generation sequencing, which included more than 5000 genes associated with inherited diseases. Results: Hematochemical findings were unremarkable. The ECG, Holter, 2D-echo, and CTA of brain vessels were normal. Cerebral magnetic resonance showed the presence of multiple small foci of ischemia. Coronary and ventricular angiography showed normal arteries with remarkably slow flow and multiple biventricular micro-aneurysms. At the endomyocardial biopsy, five of seven arterioles presented severe lumen obstruction due to hypertrophy and disarray of the muscular coat. Similarly, obstructed pre-glomerular arteries with glomerular sclerosis were seen at the renal biopsy. Genetics identified mutations in the ABCC6, MMP2, and XYLT1 genes, which play pivotal roles in the extracellular matrix. Conclusion: This study described a new genetic microvascular obstructive disease causing progressive hypo-perfusion of the human brain, heart, and kidney.

IntroductionCompared with breast cancer (BC) in women, BC in men is a rare disease with genetic and molecular peculiarities. Therapeutic approaches for male BC (MBC) are currently extrapolated from the clinical management of female BC, although the disease does not exactly overlap in males and females. Data on specific molecular biomarkers in MBC are lacking, cutting out male patients from more appropriate therapeutic strategies. Growing evidence indicates that Next Generation Sequencing (NGS) multigene panel testing can be used for the detection of predictive molecular biomarkers, including Tumor Mutational Burden (TMB) and Microsatellite Instability (MSI).MethodsIn this study, NGS multigene gene panel sequencing, targeting 1.94 Mb of the genome at 523 cancer-relevant genes (TruSight Oncology 500, Illumina), was used to identify and characterize somatic variants, Copy Number Variations (CNVs), TMB and MSI, in 15 Formalin-Fixed Paraffin-Embedded (FFPE) male breast cancer samples.Results and discussionA total of 40 pathogenic variants were detected in 24 genes. All MBC cases harbored at least one pathogenic variant. PIK3CA was the most frequently mutated gene, with six (40.0%) MBCs harboring targetable PIK3CA alterations. CNVs analysis showed copy number gains in 22 genes. No copy number losses were found. Specifically, 13 (86.7%) MBCs showed gene copy number gains. MYC was the most frequently amplified gene with eight (53.3%) MBCs showing a median fold-changes value of 1.9 (range 1.8-3.8). A median TMB value of 4.3 (range 0.8-12.3) mut/Mb was observed, with two (13%) MBCs showing high-TMB. The median percentage of MSI was 2.4% (range 0-17.6%), with two (13%) MBCs showing high-MSI. Overall, these results indicate that NGS multigene panel sequencing can provide a comprehensive molecular tumor profiling in MBC. The identification of targetable molecular alterations in more than 70% of MBCs suggests that the NGS approach may allow for the selection of MBC patients eligible for precision/targeted therapy.

BackgroundDespite the efficacy of immunotherapy, only a small percentage of patients achieves a long-term benefit in terms of overall survival. The aim of this study was to define an immune profile predicting the response to immune checkpoint inhibitors (ICIs).MethodsPatients with advanced solid tumors, who underwent ICI treatment were enrolled in this prospective study. Blood samples were collected at the baseline. Thirteen soluble immune checkpoints, 3 soluble adhesion molecules, 5 chemokines and 11 cytokines were analyzed. The results were associated with oncological outcomes.ResultsRegardless of tumor type, patients with values of sTIM3, IFNα, IFNγ, IL1β, IL1α, IL12p70, MIP1β, IL13, sCD28, sGITR, sPDL1, IL10 and TNFα below the median had longer overall survival (p

Primary Sjögren’s Syndrome (pSS) is a systemic autoimmune disease that primarily attacks the lacrimal and salivary glands, resulting in impaired secretory function characterized by xerostomia and xerophthalmia. Patients with pSS have been shown to have impaired salivary gland innervation and altered circulating levels of neuropeptides thought to be a cause of decreased salivation, including substance P (SP). Using Western blot analysis and immunofluorescence studies, we examined the expression levels of SP and its preferred G protein-coupled TK Receptor 1 (NK1R) and apoptosis markers in biopsies of the minor salivary gland (MSG) from pSS patients compared with patients with idiopathic sicca syndrome. We confirmed a quantitative decrease in the amount of SP in the MSG of pSS patients and demonstrated a significant increase in NK1R levels compared with sicca subjects, indicating the involvement of SP fibers and NK1R in the impaired salivary secretion observed in pSS patients. Moreover, the increase in apoptosis (PARP-1 cleavage) in pSS patients was shown to be related to JNK phosphorylation. Since there is no satisfactory therapy for the treatment of secretory hypofunction in pSS patients, the SP pathway may be a new potential diagnostic tool or therapeutic target.

Abstract Objective First-line therapy for metastatic squamous cell carcinoma of the head and neck (R/M HNSCC) has been revolutionized by the introduction of anti-checkpoint monoclonal antibodies, which have shown a significant improvement in overall survival (OS) gaining approval in a first line setting. Efficacy and safety of first-line weekly chemotherapy, compared to 3-weeks treatment, was retrospectively evaluated in a frail patient population with R/M HNSCC with the aim to evaluate its role as part of a personalized first-line approach. Methods A total of 124 patients with locally incurable R/M HNSCC receiving weekly (21) or three-weekly (103) chemotherapy plus cetuximab in a first line setting from December 2010 to September 2020 were retrospectively reviewed. Treatment outcomes in terms of objective response rate (ORR), progression-free survival (PFS), overall survival (OS) and toxicities were analysed. Results Patients in the three-week subgroup were ECOG PS 0 (39) and 1 (64) while patients in weekly group (21) were all PS 2. No significant differences were reported in terms of age, sex, smoking and previous alcohol abuse considering the two distinct subgroups. Moreover, no statistically significant difference was found in PFS and OS between the two treatment subgroups. The response rate was 35% (36 patients) and 34% (7 patients) in three-week and weekly treatment group, respectively. Seventy patients (68%) in the three-week group experienced chemotherapy-related toxicities, predominantly G3. In the weekly group a predominantly low-grade toxicity was found in a lower number of patients (52%). Conclusion The weekly schedule appears to be an active and safe strategy in frail patients with R/M HNSCC. Based on these data, a weekly schedule could be considered as a first line treatment in all frail patients excluded from pembrolizumab treatment and a study on the combination of weekly chemotherapy and immunotherapy should be performed.

Abstract Background The response to immunotherapy can be impaired by several factors including external intervention such as drug interactions with immune system. We aimed to examine the immunomodulatory action of opioids, since immune cells express opioid receptors able to negatively influence their activities. Methods This observational, multicenter, retrospective study, recruited patients with different metastatic solid tumors, who have received immunotherapy between September 2014 and September 2019. Immunotherapy was administered according to the standard schedule approved for each primary tumor and line of treatment. The concomitant intake of antibiotics, antifungals, corticosteroids and opioids were evaluated in all included patients. The relationship between tumor response to immunotherapy and the oncological outcomes were evaluated. A multivariate Cox-proportional hazard model was used to identify independent prognostic factors for survival. Results One hundred ninety-three patients were recruited. Overall, progression-free survival (PFS) and overall survival (OS) were significantly shorter in those patients taking opioids than in those who didn’t (median PFS, 3 months vs. 19 months, HR 1.70, 95% CI 1.37–2.09, p

Pembrolizumab, an anti-PD-1 antibody, has been approved as first-line treatment for recurrent or metastatic head and neck squamous cell carcinoma ((R/M) HNSCC). However, only a minority of patients benefit from immunotherapy, which highlights the need to identify novel biomarkers to optimize treatment strategies. CD137+ T cells have been identified as tumour-specific T cells correlated with immunotherapy responses in several solid tumours. In this study, we investigated the role of circulating CD137+ T cells in (R/M) HNSCC patients undergoing pembrolizumab treatment. PBMCs obtained from 40 (R/M) HNSCC patients with a PD-L1 combined positive score (CPS) ≥1 were analysed at baseline via cytofluorimetry for the expression of CD137, and it was found that the percentage of CD3+CD137+ cells is correlated with the clinical benefit rate (CBR), PFS, and OS. The results show that levels of circulating CD137+ T cells are significantly higher in responder patients than in non-responders (p = 0.03). Moreover, patients with CD3+CD137+ percentage ≥1.65% had prolonged OS (p = 0.02) and PFS (p = 0.02). Multivariate analysis, on a combination of biological and clinical parameters, showed that high levels of CD3+CD137+ cells (≥1.65%) and performance status (PS) = 0 are independent prognostic factors of PFS (CD137+ T cells, p = 0.007; PS, p = 0.002) and OS (CD137+ T cells, p = 0.006; PS, p = 0.001). Our results suggest that levels of circulating CD137+ T cells could serve as biomarkers for predicting the response of (R/M) HNSCC patients to pembrolizumab treatment, thus contributing to the success of anti-cancer treatment.

Background: Programmed death-ligand 1 (PD-L1) checkpoint inhibitors represent a mainstay of therapy in head and neck squamous cell cancer (HNSCC). However, little is known about the influence of combined therapy on PD-L1 expression. The study aims to gather evidence on this topic. Methods: A systematic search was carried out in electronic databases Pubmed-MEDLINE and Embase to retrieve studies on the comparison of PD-L1 expression before and after conventional therapy. Data were extracted and a quantitative analysis with pooled odds ratios (ORs) was performed when applicable. Results: Of 5688 items, 15 were finally included. Only a minority of studies assessed PD-L1 with the recommended combined positive score (CPS). The results are highly heterogeneous, with some studies reporting an increase in PD-L1 expression and others reporting a decrease. Three studies allowed for quantitative analysis and showed a pooled OR of 0.49 (CI 0.27–0.90). Conclusions: From the present evidence, a clear conclusion towards an increase or decrease in PD-L1 expression after combined therapy cannot be drawn, but even with few studies available, a trend towards an increase in expression in tumor cells at a cutoff of 1% can be noted in patients undergoing platinum-based therapy. Future studies will provide more robust data on the effect of combined therapy on PD-L1 expression.

Abstract Backgrounds The organization of minor salivary glands (MSG) infiltrates, in patients with Sjögren’s syndrome (SS), associates with disease severity and progression. Aberrant regulation of lymphocyte autophagy is involved in autoimmunity, and in previous work, we provided the first evidence of upregulated autophagy in CD4+ T cells infiltrating SS MSG. The aim of this study was to further explore autophagy in SS infiltrating and circulating lymphocytes and to investigate its role in disease histopathological progression. Methods After collection of 20 SS MSG, the presence of lymphocyte aggregates (foci) and the formation of germinal center (GC)-like structures were observed by H&E and confirmed by immunohistochemistry. The expression of autophagy-related genes, Atg5 and MAP1LC3A, was detected by RT-PCR on microdissected salivary gland tissue and control tonsils. In MSG and tonsils, autophagic lymphocytes were identified by the detection of the autophagosome protein LC3B visualized as LC3 puncta staining by immunofluorescence. Peripheral blood autophagy was assessed by flow cytometry in SS and healthy controls (HC). Results Real-time PCR demonstrated higher expression in the autophagy genes Atg5 and MAP1LC3A in MSG GCs as compared to both small foci (p = 0.0075, p = 0.0002) and GCs from tonsils (p = 0.0001, p = 0.0037). In MSG, LC3 puncta staining was detectable on both CD3+ and CD20+ lymphocytes; in tonsils, LC3 puncta was almost undetectable on all lymphocytes. Compared to HC (n = 20), flow cytometry did not reveal any increase of autophagy in SS circulating lymphocytes (n = 30). Conclusions In SS MSG, lymphocytes’ autophagy is a feature of infiltrating T and B cells and is associated with histological severity. Interestingly, in MSG aberrant regulation of autophagy is detectable in GC-like structures possibly indicating its involvement in the development and persistence of the autoimmune process within the lesions.

Abstract Background Despite the efficacy of immune checkpoint inhibitors (ICIs) only the 20–30% of treated patients present long term benefits. The metabolic changes occurring in the gut microbiota metabolome are herein proposed as a factor potentially influencing the response to immunotherapy. Methods The metabolomic profiling of gut microbiota was characterized in 11 patients affected by non-small cell lung cancer (NSCLC) treated with nivolumab in second-line treatment with anti-PD-1 nivolumab. The metabolomics analyses were performed by GC–MS/SPME and 1H-NMR in order to detect volatile and non-volatile metabolites. Metabolomic data were processed by statistical profiling and chemometric analyses. Results Four out of 11 patients (36%) presented early progression, while the remaining 7 out of 11 (64%) presented disease progression after 12 months. 2-Pentanone (ketone) and tridecane (alkane) were significantly associated with early progression, and on the contrary short chain fatty acids (SCFAs) (i.e., propionate, butyrate), lysine and nicotinic acid were significantly associated with long-term beneficial effects. Conclusions Our preliminary data suggest a significant role of gut microbiota metabolic pathways in affecting response to immunotherapy. The metabolic approach could be a promising strategy to contribute to the personalized management of cancer patients by the identification of microbiota-linked “indicators” of early progressor and long responder patients.

ObjectiveThe monoclonal antibodies anti-programmed death protein-1 (anti–PD-1) nivolumab and pembrolizumab are the first immune checkpoint inhibitors (ICIs) approved for treatment of recurrent/metastatic head and neck carcinoma R/M HNSCC in first line and in platinum refractory disease. This network meta-analysis aims to investigate the efficacy of anti–PD-1- vs anti–PD-L1-based therapy in R/M HNSCC cancer patients through a systematic review of the literature to provide support for evidence-based treatment decisions. In particular, the effectiveness of ICIs for R/M HNSCC is analyzed according to the different mechanisms of action of the check-points inhibitory drugs in different subgroups of patients.MethodsWe did a systematic literature review and network meta-analysis (NMA) of randomized controlled trials (RCTs) in PubMed, ClinicalTrials.gov, Embase, Medline, the Cochrane Central Register of Controlled Trials, Web of Science. Our search identified a total of five randomized controlled trials: Keynote 040, Keynote 048, Eagle, Condor, Checkmate 141. These trials included 3001 patients. Treatment was sub-categorized into PD-L1–based, PD-1–based, and standard chemotherapy. Treatments were indirectly compared with anti–PD-L1-based therapy.ResultsThe network meta-analysis demonstrated no significant differences in OS between different subgroups except for the metastatic patients in which anti–PD-1-based therapy was associated with significantly less risk of death. Furthermore, anti–PD-1-based therapy appeared to be effective in smoker patients and in human papilloma–negative (HPV) patients. Conversely, anti–PD-L1-based therapy seems to be better efficient in female patients, in locally recurrent setting and in HPV positive patients.ConclusionThis is the first NMA study that aimed to indirectly compare anti–PD-1- and anti–PD-L1-based therapy in HNSCC patients. The results of our NMA could help define a profile of patient responder or resistant to specific classes of immune drugs and can be used to guide/design future studies in the novel scenario of precision immune-oncology.

Introduction: Only a minority of patients with platinum refractory head and neck squamous cell carcinoma (PR/HNSCC) gain some lasting benefit from immunotherapy. Methods: The combined role of the comprehensive genomic (through the FoundationOne Cdx test) and immune profiles of 10 PR/HNSCC patients treated with the anti-PD-1 nivolumab was evaluated. The immune profiles were studied both at baseline and at the second cycle of immunotherapy, weighing 20 circulating cytokines/chemokines, adhesion molecules, and 14 soluble immune checkpoints dosed through a multiplex assay. A connectivity map was obtained by calculating the Spearman correlation between the expression profiles of circulating molecules. Results: Early progression occurred in five patients, each of them showing TP53 alteration and three of them showing a mutation/loss/amplification of genes involved in the cyclin-dependent kinase pathway. In addition, ERB2 amplification (1 patient), BRCA1 mutation (1 patient), and NOTCH1 genes alteration (3 patients) occurred. Five patients achieved either stable disease or partial response. Four of them carried mutations in PI3K/AKT/PTEN pathways. In the only two patients, with a long response to immunotherapy, the tumor mutational burden (TMB) was high. Moreover, a distinct signature, in terms of network connectivity of the circulating soluble molecules, characterizing responder and non-responder patients, was evidenced. Moreover, a strong negative and statistically significant (p-value ≤ 0.05) correlation with alive status was evidenced for sE-selectin at T1. Conclusions: Our results highlighted the complexity and heterogeneity of HNSCCs, even though it was in a small cohort. Molecular and immune approaches, combined in a single profile, could represent a promising strategy, in the context of precision immunotherapy.

Background and Aim: Metachronous, isolated adrenal metastases from breast cancer are extremely rare. The aim of this study was to evaluate the results of adrenalectomy as a treatment of this uncommon condition.Methods: Twelve female patients (median age: 68 years) underwent 13 adrenalectomies for isolated, metachronous metastases of breast cancer. Ten resections were performed thorugh open surgery and two were preformed through a laparoscopic approach. As main study endpoints, postoperative mortality, postoperative morbidity and disease-free survival were considered. Median length of follow-up was 40 months.Results: Postoperative mortality was absent. Postoperative morbidity was 17%: one patient presented a postoperative pneumothorax requiring drainage and one patient required re-hospitalization 8 days after contralateral adrenalectomy for electrolyte imbalance. Two patients died of recurrent metastatic disease, 28 and 33 months respectively after adrenalectomy. One patient remained alive with hepatic metastases at 32 months from resection of adrenal recurrence. All in all, disease-free survival at 48 months was 75%.Conclusions: Adrenalectomy for metachronous, isolated metastases of breast cancer can be performed with no postoperative mortality and minimal postoperative morbidity, enabling good long-term disease-free survival.

Abstract The aim of the study was to verify whether neuromuscular magnetic stimulation (NMMS) improves muscle function in spinal-onset amyotrophic lateral sclerosis (ALS) patients. Twenty-two ALS patients were randomized in two groups to receive, daily for two weeks, NMMS in right or left arm (referred to as real-NMMS, rNMMS), and sham NMMS (sNMMS) in the opposite arm. All the patients underwent a median nerve conduction (compound muscle action potential, CMAP) study and a clinical examination that included a handgrip strength test and an evaluation of upper limb muscle strength by means of the Medical Research Council Muscle Scale (MRC). Muscle biopsy was then performed bilaterally on the flexor carpi radialis muscle to monitor morpho-functional parameters and molecular changes. Patients and physicians who performed examinations were blinded to the side of real intervention. The primary outcome was the change in the muscle strength in upper arms. The secondary outcomes were the change from baseline in the CMAP amplitudes, in the nicotinic ACh currents, in the expression levels of a selected panel of genes involved in muscle growth and atrophy, and in histomorphometric parameters of ALS muscle fibers. The Repeated Measures (RM) ANOVA with a Greenhouse-Geisser correction (sphericity not assumed) showed a significant effect [F(3, 63) = 5.907, p

Primary squamous cell carcinoma of salivary gland (SCG) is an extremely rare type of malignant salivary gland tumor, which in turn results in scarcity of data available regarding both its treatment and associated genetic alterations. A retrospective analysis of 12 patients with primary SCG was conducted, along with analysis of the association between treatment, clinical/pathological characteristics, and outcomes. Most patients (8) were staged IVa, with the majority of them (10) having G3 fast growing cancer. Local and systemic recurrence were reported in only three out of nine parotid cases (0 out of 2 submandibular SCGs). In two out of eight patients local relapse occurred after integrated treatment, while recurrence occurred in two out of three patients undergoing exclusive surgery. Five patients eventually died. Treatment of resectable disease must be aggressive and multimodal, with achievement of loco-regional control in order to reduce rate of recurrence and improve outcomes. Metastatic disease would require a therapeutic strategy tailored to the molecular profile in order to improve the currently disappointing results.

Introduction: Seroma formation represents one of the most frequent postoperative complications of axillary dissection in breast cancer (BC) patients. We aimed to retrospectively explore the effectiveness of the intraoperative use of a synthetic cyanoacrylate glue (specifically Glubran®2) vs. the intraoperative use of a fibrin sealant (specifically Tisseel) in reducing seroma formation compared to the use of nonsealant in BC patients who underwent breast surgery and axillary dissection.Materials and Methods: We conducted a retrospective, monocentric observational study on BC patients who underwent axillary dissection associated with breast surgery. The axillary dissection was completed with the application of a closed suction drain and was preceded by the application of either Glubran®2 glue or Tisseel sealant or nonsealant. We analyzed the quantity of serum drained in the first 3 postoperative days, length of hospitalization, days of permanence of axillary drain, seroma development, and presence of postoperative infection signs.Results: Forty-one BC patients were considered. Based on the device used during the surgical treatment, the patients were divided into three groups: group A (17 patients), to whom suction axillary drain was applied; group B (7 patients), to whom Tisseel and axillary suction drain were applied; and group C (17 patients), to whom Glubran®2 and axillary suction drain were applied. Among the three groups, we did not find significant differences in terms of amount of serum drained in the first 3 postoperative days, length of hospitalization, and incidence of seroma. Group C maintained the axillary drain in a significantly lower number of days compared to the other two groups (p = 0.02); it also had a lower incidence of postoperative infections (6%) compared to group A (23%) and group B (57%) (p = 0.02).Conclusions: We did not find any evidence that the use of surgical glues may reduce the formation of seroma following axillary dissection in BC patients. Nevertheless, the use of cyanoacrylate glue in association with closed suction axillary drain seems to contribute to the reduction in days of axillary drain permanence and of postoperative infections, which are known factors delaying the schedule of any adjuvant oncological therapies.

Background: Clinical trials showed that only a subset of patients benefits from immunotherapy, suggesting the need to identify new predictive biomarker of resistance. Indoleamine-2,3-dioxygenase (IDO) has been proposed as a mechanism of resistance to anti-PD-1 treatment, and serum kynurenine/tryptophan (kyn/trp) ratio represents a possible marker of IDO activity.Methods: Metastatic non-small cell lung cancer (NSCLC), renal cell carcinoma (RCC), and head and neck squamous cell carcinoma (HNSCC) treated with nivolumab as second-line treatment were included in this prospective study. Baseline serum kyn and trp levels were measured by high-performance liquid chromatography to define the kyn/trp ratio. The χ2-test and t-test were applied to compare frequencies and mean values of kyn/trp ratio between subgroups with distinct clinical/pathological features, respectively. Median baseline kyn/trp ratio was defined and used as cutoff in order to stratify the patients. The association between kyn/trp ratio, clinical/pathological characteristics, response, progression-free survival (PFS), and overall survival (OS) was analyzed.Results: Fifty-five patients were included. Mean baseline serum kyn/trp ratio was significantly lower in female than in male patients (0.048 vs. 0.059, respectively, p = 0.044) and in patients with lung metastasis than in others (0.053 vs. 0.080, respectively, p = 0.017). Mean baseline serum kyn/trp ratio was significantly higher in early progressor patients with both squamous and non-squamous NSCLC (p = 0.003) and with a squamous histology cancer (19 squamous NSCLC and 14 HNSCC, p = 0.029). The median value of kyn/trp ratio was 0.06 in the overall population. With the use of median value as cutoff, patients with kyn/trp ratio > 0.06 had a higher risk to develop an early progression (within 3 months) to nivolumab with a trend toward significance (p = 0.064 at multivariate analysis). Patients presenting a baseline kyn/trp ratio ≤0.06 showed a longer PFS [median 8 vs. 3 months; hazard ratio (HR): 0.49; 95% confidence interval (CI) 0.24–1.02; p = 0.058] and a significantly better OS than did those with a kyn/trp ratio > 0.06 (median 16 vs. 4 months; HR: 0.39; 95% CI 0.19–0.82; p = 0.013).Conclusion: Serum kyn/trp ratio could have both prognostic and predictive values in patients with solid tumor treated with immunotherapy, probably reflecting a primary immune-resistant mechanism regardless of the primary tumor histology. Its relative weight is significantly related to gender, site of metastasis, NSCLC, and squamous histology, although these suggestive data need to be confirmed in larger studies.

Abstract Background Immune checkpoint inhibitors have revolutionized the treatment paradigm of highly lethal malignancies like advanced non-small cell lung cancer (NSCLC), demonstrating long-term tumour control and extended patient survival. Unfortunately, only 25–30% of patients experience a durable benefit, while the vast majority demonstrate primary or acquired resistance. Recently, indoleamine 2,3-dioxygenase (IDO) activity has been proposed as a possible mechanism of resistance to anti-PD-1 treatment leading to an immunosuppressive microenvironment. Methods Pre-treatment serum concentrations of tryptophan (trp) and kynurenine (kyn) were measured by high-performance liquid chromatography tandem mass spectrometry in NSCLC patients treated with second-line nivolumab. The IDO activity was expressed with kyn/trp ratio. The associations between kyn/trp ratio and early progression, performance status (PS), age, sex, brain metastases, pleural effusion, progression free survival (PFS) and overall survival (OS) were analyzed using Spearman test and Mann–Whitney test. Results Twenty-six NSCLC patients were included in our study; 14 of them (54%) presented early progression (

Background: to evaluate whether Apparent Diffusion Coefficient (ADC) values of invasive breast cancer, provided by 3T Diffusion Weighted-Images (DWI), may represent a non-invasive predictor of pathophysiologic tumor aggressiveness. Methods: 100 Patients with histologically proven invasive breast cancers who underwent a 3T-MRI examination were included in the study. All MRI examinations included dynamic contrast-enhanced and DWI/ADC sequences. ADC value were calculated for each lesion. Tumor grade was determined according to the Nottingham Grading System, and immuno-histochemical analysis was performed to assess molecular receptors, cellularity rate, on both biopsy and surgical specimens, and proliferation rate (Ki-67 index). Spearman’s Rho test was used to correlate ADC values with histological (grading, Ki-67 index and cellularity) and MRI features. ADC values were compared among the different grading (G1, G2, G3), Ki-67 (20%) and cellularity groups (70%), using Mann–Whitney and Kruskal-Wallis tests. ROC curves were performed to demonstrate the accuracy of the ADC values in predicting the grading, Ki-67 index and cellularity groups. Results: ADC values correlated significantly with grading, ER receptor status, Ki-67 index and cellularity rates. ADC values were significantly higher for G1 compared with G2 and for G1 compared with G3 and for Ki-67 < 20% than Ki-67 > 20%. The Kruskal-Wallis test showed that ADC values were significantly different among the three grading groups, the three biopsy cellularity groups and the three surgical cellularity groups. The best ROC curves were obtained for the G3 group (AUC of 0.720), for G2 + G3 (AUC of 0.835), for Ki-67 > 20% (AUC of 0.679) and for surgical cellularity rate > 70% (AUC of 0.805). Conclusions: 3T-DWI ADC is a direct predictor of cellular aggressiveness and proliferation in invasive breast carcinoma, and can be used as a supporting non-invasive factor to characterize macroscopic lesion behavior especially before surgery.

Unresectable recurrent and/or metastatic head and neck squamous cell carcinoma (R/M HNSCC) has a very poor prognosis. Soluble immune checkpoints (sICs) are circulating proteins that result from the alternative splicing of membrane proteins and can modulate the immune response to cancer cells. The aim of our pilot study was to determine the possible role of a comprehensive evaluation of sICs in the classification of prognosis and response to treatment in patients with advanced disease. We evaluated several sICs (CD137, CTLA-4, PD-1, PD-L1, PD-L2, TIM3, LAG3, GITR, HVEM, BTLA, IDO, CD80, CD27, and CD28) from peripheral blood at baseline and investigated the association with clinical characteristics and outcomes. A high baseline soluble LAG3 (sLAG3 > 377 pg/mL) resulted in an association with poor PFS and OS (p = 0.047 and p = 0.003, respectively). Moreover, sLAG3 emerged as an independent prognostic factor using an MVA (p = 0.005). The evaluation of sICs, in particular sLAG3, may be relevant for identifying patients with worse prognoses, or resistance to treatments, and may lead to the development of novel targeted strategies.

The pathologic definition of triple negative breast cancer (TNBC) relies on the absence of expression of estrogen, progesterone and HER2 receptors. However, this BC subgroup is distinguished by a wide biological, molecular and clinical heterogeneity. Among the intrinsic TNBC subtypes, the mesenchymal type is defined by the expression of genes involved in the epithelial to mesenchymal transition, stromal interaction and cell motility. Moreover, it shows a high expression of genes involved in proliferation and an immune-suppressive microenvironment. Several molecular alterations along different pathways activated during carcinogenesis and tumor progression have been outlined and could be involved in immune evasion mechanisms. Furthermore, reverting epithelial to mesenchymal transition process could lead to the overcoming of immune-resistance. This paper reviews the current knowledge regarding the mesenchymal TNBC subtype and its response to conventional therapeutic strategies, as well as to some promising molecular target agents and immunotherapy. The final goal is a tailored combination of cytotoxic drugs, target agents and immunotherapy in order to restore immunocompetence in mesenchymal breast cancer patients.

Glycosylation, the posttranslational linking of sugar molecules to proteins, is notoriously altered during tumor transformation. More specifically in carcinomas, GalNAc-type O-glycosylation, is characterized by biosynthetically immature truncated glycans present on the cancer cell surface, which profoundly impact anti-tumor immune recognition. The tumor-associated glycan pattern may thus be regarded as a biomarker of immune modulation. In epithelial ovarian cancer (EOC) there is a particular lack of specific biomarkers and molecular targets to aid early diagnosis and develop novel therapeutic interventions. The aim of this study was to investigate the ovarian cancer O-glycoproteome and identify tumor-associated glycoproteins relevant in tumor–dendritic cell (DC) interactions, mediated by macrophage galactose-like C type lectin (MGL), which recognizes the tumor-associated Tn O-glycan. Lectin weak affinity chromatography (LWAC) was employed to probe the O-glycopeptidome by MGL and Vicia villosa agglutinin (VVA) lectin using glycoengineered ovarian cancer cell lines and ovarian cancer tissues as input material. Biochemical and bioinformatics analysis gave information on the glycan arrangement recognized by MGL in tumor cells. The potential MGL binders identified were located, as expected, at the cell membrane, but also within the intracellular compartment and the matrisome, suggesting that MGL in vivo may play a complex role in sensing microenvironmental cues. The tumor glycoproteins binders for MGL may become relevant to characterize the interaction between the immune system and tumor progression and contribute to the design of glycan targeting-based strategies for EOC immunotherapeutic interventions.

Pathological complete response (pCR) after neoadjuvant chemotherapy (NACT) can predict better survival outcomes in patients with early triple negative breast cancer (TNBC). Tumor infiltrating lymphocytes (TILs), Programmed Death-Ligand 1 (PD-L1), and Cluster of Differentiation 73 (CD73) are immune-related biomarkers that can be evaluated in the tumor microenvironment. We investigated if the contemporary expression of these biomarkers combined in a tissue immune profile (TIP) can predict pCR better than single biomarkers in TNBC. Tumor infiltrating lymphocytes (TILs), CD73 expression by cancer cells (CC), and PD-L1 expression by immune cells (IC) were evaluated on pre-NACT biopsies. We defined TIP positive (TIP+) as the simultaneous presence of TILS ≥ 50%, PD-L1 ≥ 1%, and CD73 ≤ 40%. To consider the effects of all significant variables on the pCR, multivariate analysis was performed. Akaike information criterion (AIC) and Bayesian information criterion (BIC) were used for model selection. We retrospectively analyzed 60 biopsies from patients with TNBC who received standard NACT. Pathological complete response was achieved in 23 patients (38.0%). Twelve (20.0%) cases resulted to be TIP+. The pCR rate was significantly different between TIP+ (91.7%) and TIP− (25.0%) (p < 0.0001). Using a multivariate analysis, TIP was confirmed as an independent predictive factor of pCR (OR 49.7 (6.30–392.4), p < 0.0001). Finally, we compared the efficacy of TIP versus each single biomarker in predicting pCR by AIC and BIC. The combined immune profile is more accurate in predicting pCR (AIC 68.3; BIC 74.5) as compared to single biomarkers. The association between TIP+ and pCR can be proposed as a novel link between immune background and response to chemotherapy in TNBC, highlighting the need to consider an immunological patients’ profile rather than single biomarkers.

Background: Previous locoregional treatment could affect the response to nivolumab in platinum-refractory recurrent/metastatic head and neck squamous cell carcinoma (R/M HNSCC). The aim of this study is to evaluate the impact of the clinicopathological characteristics and previous treatment in predicting early progression to nivolumab in a real-world population. Methods: This is an observational, multicenter retrospective/prospective study including patients (pts) with platinum refractory R/M HNSCC who received nivolumab 240 mg every 2 weeks from October 2018 to October 2019. We analyzed the association between previous treatment, clinicopathological characteristics, and early progression (within 3 months). Results: Data from 61 pts were reviewed. Median age was 67 years (30–82). Forty-two pts (69%) received previous locoregional treatment. Early progression to nivolumab occurred in 36 pts (59%), while clinical benefit (stable disease and partial response) was achieved in 25 pts (41%). Early progression to nivolumab was significantly associated to previous locoregional treatment both at univariate and multivariate analysis (p = 0.005 and p = 0.048, respectively). Conclusion: nivolumab in R/M HNSCC is burdened with a high early progression rate. Previous wide neck dissection and high dose radiotherapy may compromise the efficacy of nivolumab, distorting the anatomy of the local lymphatic system and hindering the priming of immune response.

Immune checkpoint inhibitors have revolutionized treatment and outcome of melanoma and many other solid malignancies including non-small cell lung cancer (NSCLC) and renal cell carcinoma (RCC). Unfortunately, only a minority of patients have a long-term benefit, while the remaining demonstrate primary or acquired resistance. Recently, it has been demonstrated that the prevalence of programmed death-ligand 1 (PD-L1) and tumor-infiltrating lymphocytes (TILs) varies based on the anatomical site of metastases. In particular, liver seems to have more immunosuppressive microenvironment while both the presence of lymph nodal disease and lung metastases seem to have the highest prevalence of PD-L1 and TILs. The aim of the present study is to investigate the possible role of site of metastases as a predictive factor for response or resistance to immunotherapy in several types of cancer. In this multicenter retrospective study, we enrolled patients with metastatic NSCLC, melanoma, RCC, urothelial, merkel carcinoma, and colon cancer who received immunotherapy from April 2015 to August 2019. Major clinicopathological parameters were retrieved and correlated with patients’ survival outcomes in order to assess their prognostic value and build a useful tool to assist in the decision-making process. A total of 291 patients were included in this study. One hundred eighty-seven (64%) patients were male and 104 (36%) female. The tumor histology was squamous NSCLC in 56 (19%) patients, non-squamous NSCLC in 99 (34%) patients, melanoma in 101 (35%) patients, RCC in 28 (10%) patients, and other tumors in the remaining 7 (2%) patients. The number of metastatic sites was 1 in 103 patients (35%), 2 in 104 patients (36%) and 3 in 84 patients (29%). Out of 183 valuable patients, the entity of response was complete response (CR), partial response (PR), stable disease (SD), and progression disease (PD) in 15, 53, 31, and 79 patients, respectively. Using an univariate analysis (UVA), tumor burden (p = 0.0004), the presence of liver (p = 0.0009), bone (p = 0.0016), brain metastases (p < 0.0001), the other metastatic sites (p = 0.0375), the number of metastatic sites (p = 0.0039), the histology (p = 0.0034), the upfront use of immunotherapy (p = 0.0032), and Eastern Cooperative Oncology Group (ECOG) Perfomance status (PS) ≥ 1 (p < 0.0001) were significantly associated with poor overall survival (OS). Using a multivariate analysis (MVA) the presence of liver (p = 0.0105) and brain (p = 0.0026) metastases, the NSCLC diagnosis (p < 0.0001) and the ECOG PS (p < 0.0001) resulted as significant prognostic factors of survival. Regarding the progression free survival (PFS), using a UVA of the tumor burden (p = 0.0004), bone (p = 0.0098) and brain (p = 0.0038) metastases, the presence of other metastatic sites (p = 0.0063), the number of metastatic sites (p = 0.0007), the histology (p = 0.0007), the use of immunotherapy as first line (p = 0.0031), and the ECOG PS ≥ 1 (p ≤ 0.0001) were associated with a lower PFS rate. Using an MVA, the presence of brain (p = 0.0088) and liver metastases (p = 0.024) and the ECOG PS (p < 0.0001) resulted as predictors of poor PFS. Our study suggests that the site of metastases could have a role as prognostic and predictive factor in patients treated with immunotherapy. Indeed, regardless of the histology, the presence of liver and brain metastases was associated with a shorter PFS and OS, but these results must be confirmed in further studies. In this context, a deep characterization of microenvironment could be crucial to prepare patients through novel strategies with combination or sequential immunotherapy in order to improve treatment response.

BACKGROUND:5-FU based chemotherapy is the most common first line regimen used for metastatic colorectal cancer (mCRC). Identification of predictive markers of response to chemotherapy is a challenging approach for drug selection. The present study analyzes the predictive role of 5-FU degradation rate (5-FUDR) and genetic polymorphisms (MTHFR, TSER, DPYD) on survival. MATERIALS AND METHODS:Genetic polymorphisms of MTHFR, TSER and DPYD, and the 5-FUDR of homogenous patients with mCRC were retrospectively studied. Genetic markers and the 5-FUDR were correlated with clinical outcome. RESULTS:133 patients affected by mCRC, treated with fluoropyrimidine-based chemotherapy from 2009 to 2014, were evaluated. Patients were classified into three metabolic classes, according to normal distribution of 5-FUDR in more than 1000 patients, as previously published: poor-metabolizer (PM) with 5-FU-DR ≤ 0,85 ng/ml/106 cells/min (8 pts); normal metabolizer with 0,85 < 5-FU-DR < 2,2 ng/ml/106 cells/min (119 pts); ultra-rapid metabolizer (UM) with 5-FU-DR ≥ 2,2 ng/ml/106 cells/min (6 pts). PM and UM groups showed a longer PFS respect to normal metabolizer group (14.5 and 11 months respectively vs 8 months; p = 0.029). A higher G3-4 toxicity rate was observed in PM and UM, respect to normal metabolizer (50% in both PM and UM vs 18%; p = 0.019). No significant associations between genes polymorphisms and outcomes or toxicities were observed. CONCLUSION:5-FUDR seems to be significantly involved in predicting survival of patients who underwent 5-FU based CHT for mCRC. Although our findings require confirmation in large prospective studies, they reinforce the concept that individual genetic variation may allow personalized selection of chemotherapy to optimize clinical outcomes.

Introduction. True malignant mixed tumor, also known as carcinosarcoma, is a rare tumor of the salivary gland composed of both malignant epithelial and malignant mesenchymal elements. Frequently carcinosarcoma arises in the background of a preexisting pleomorphic adenoma; however, if no evidence of benign mixed tumor is present, the lesion is known as carcinosarcoma “de novo.” We reported the first case of true malignant mixed tumor of the submandibular gland composed of high grade mucoepidermoid carcinoma associated with osteosarcoma. Case Presentation. A 69-year-old Caucasian male came to our department complaining of the appearance of an asymptomatic left submandibular neoformation progressively increasing in size over 3 months. We opted for surgical treatment. Histological examination confirmed the diagnosis of carcinosarcoma with the coexistence of high grade mucoepidermoid carcinoma and osteosarcoma. Conclusion. To the best of our knowledge, in the true malignant mixed tumor of the submandibular gland, mucoepidermoid carcinoma associated with osteosarcoma has never been previously reported.