Atezolizumab plus carboplatin-etoposide (ACE) represents the new first-line (1L) standard of care for extended stage (ES) Small Cell Lung Cancer (SCLC) patients (pts). This is a single-center retrospective-prospective translational study aiming at investigating the correlation of immune cell distribution and their spatial metrics in tumor samples of ES SCLC pts receiving ACE as 1L treatment, with response rate (RR), progression-free survival (PFS), time to treatment failure (TTF) and overall survival (OS). A 9-color multiplex immunofluorescence panel including primary antibodies (Abs) against CD68, CD163, CD8, FoxP3, CD4, CD20 and HLA-I and a mix of Abs against tumor markers has been performed. Preliminary data on the first 39 pts are reported. After a median follow-up of 7.2 months (mos), the estimated median PFS, TTF and OS were 5.4 (95% CI 4.5–6.3), 5.8 (95% CI 3.5–8.1), and 7.8 mos (95% CI 1.9–13.7), respectively. Lower CD163+ M2-polarized macrophages density and ratio on CD8+ cells in the total and tumoral areas were favorably associated with RR, PFS, TTF and OS (p < 0.05). High intra-tumoral CD4+FoxP3+ density correlated with better PFS (p = 0.004), TTF (p = 0.011) and OS (p = 0.026). CD8+ and CD20+ B lymphocyte infiltration in the total and tumoral areas correlated with longer OS. A positive role of CD20+ interaction with CD8+ on PFS (p = 0.038), TTF (intra-tumoral, p = 0.036) and OS (p = 0.032) has been observed. High percentage (%) of stromal CD163+ close to CD8+ cells and a low % of CD163+ cells close to tumor cells were correlated with longer PFS (p = 0.045) and TTF(p = 0.034). High % of CD4+ closed to CD8+ cells in the total area (p = 0.025) and in the stroma (p = 0.002), intra and peri-tumoral interaction between CD163+ (p = 0.020) and CD8+ cells (p = 0.008), CD8+ and tumor cells interaction (p = 0.012), correlated with longer OS. We identified that immune cell populations and cell-to-cell spatial metrics in ES-SCLC pts receiving ACE significantly correlated with outcome, highlighting the importance of tumor immune microenvironment and cell-to-cell interaction for tumor response and survival.

Immune checkpoint inhibitors (ICIs) are widely used in advanced non-small cell lung cancer (aNSCLC), but, even among patients expressing high levels of PDL-1 (>/=50%), heterogeneity in clinical benefit is observed. Preliminary data suggest potential role for longitudinal liquid biopsy (LB) in predicting clinical outcome and identifying patients likely to derive detrimental effects. Aim of the study is to define the role of LB in predicting differential clinical benefit in high PDL-1 expressor treated with first-line pembrolizumab.

Immune checkpoints inhibitors (ICIs) revolutionized aNSCLC treatment. Although ICIs are well tolerated, irAEs require prompt diagnosis and adequate treatment. The lack of predictive markers and criteria for ICIs re-administration after discontinuation for toxicity represent unmet needs in clinical practice. We retrospectively reviewed clinical and pathological data of aNSCLC pts consecutively treated with single-agent ICIs at Veneto Institute of Oncology (IOV) between August 2013 and September 2021. We evaluated the impact of clinical and pathological variables affecting the risk of irAEs development and relapse, by univariate and multivariate logistic regression model. We calculated NLR and platelet to lymphocyte ratio (PLR) before first ICI administration and at the onset of first irAE, dichotomizing in high (H-) and low (L-) NLR and H- and L-PLR, using prespecified cutoff values. Our analysis included 488 pts: 159 (35%) received ICI as first-line treatment. Median OS was 9.5 months (95% CI 7.8–11.3). 183 pts (40.8%) developed at least one irAE, mainly G1 (57%), 33 (18%) discontinued ICIs due to irAE, 14 (7.6%) required hospitalization. 90 pts (20%) experienced more than one irAE. Baseline L-NLR and L-PLR were associated with higher risk of irAE development (OR = 2.46, 95% CI 1.62–3.75, p < 0.001 and OR = 1.90, 95% CI 1.27–2.85, p = 0.002). Multivariate analysis confirmed NLR as independent predictor of irAEs (OR = 2.00, 95% CI 1.14–3.52; p = 0.015). Notably, L-NLR evaluated at the time of irAE onset demonstrated to be independently associated with the risk of toxicity recurrence or development of a different irAE in multivariate analysis (OR = 3.07, 95% CI 1.13–8.29, p = 0.027). NLR might be used to predict the risk for irAE-recurrence and planning a patient-tailored follow-up during and after ICI-treatment. Prospective validation of this approach can be helpful to personalize clinical decision-making concerning the reintroduction of ICIs after discontinuation for toxicity.

BackgroundICIs are widely used in aNSCLC, but reliable predictive markers are still missing. Preliminary data suggest potential role for longitudinal LB in predicting clinical outcome and identifying pts likely to derive detrimental effects. Aim of this work is to develop a model integrating clinico-pathological variables with longitudinal LB results.MethodsWe prospectively enrolled aNSCLC pts treated with ICIs at Veneto Institute of Oncology from 2017 to 2019. LB was performed at baseline (T1), after 3-4 weeks of ICI treatment (T2). Each sample was evaluated for cell free DNA (cfDNA) quantification and analysed with a 56 gene amplicon-based NGS panel. cfDNA quantification and variant allele fraction (VAF) of tumor-associated genetic alterations were evaluated as static and dynamic parameters. The genetic alteration with the highest VAF at baseline was considered as reference for NGS analysis. Variables independently associated with the clinical outcome (p<0.05) in a multiple Cox survival model were used to derive a risk score for predicting survival probabilities.ResultsOne hundred thirteen pts were included in the analysis, 57 of them were treated in first-line. cfDNA quantification at T1, VAF at T2, cfDNA change (T2-T1), PD-L1 expression and non squamous histology were significantly associated with progression-free survival (PFS); cfDNA at T2, VAF at T2, histology and PD-L1 >/=50% were associated with overall survival (OS). Pts with all favourable variables had 70% probability of 2 year-survival and 56% probability of 3 year-survival, while all unfavourable variables were associated with only 6% probability of 6 month-survival. LB at T2 is also able to identify pts at higher risk of death within 12 weeks since the start of ICIs and define prognostic groups in pts with equal radiological response.ConclusionsAssociation of clinical variables and LB performed early during ICI treatment is able to predict OS. This integrated model upon further validation could be implemented in clinical practice to personalize treatment strategies.Legal entity responsible for the studyIstituto Oncologico Veneto IOV IRCCS.FundingIstituto Oncologico Veneto IOV IRCCS.DisclosureL. Bonanno: Financial Interests, Personal, Advisory Board: AstraZeneca, MSD, BMS, Roche; Financial Interests, Personal, Invited Speaker: AstraZeneca, MSD, BMS, Roche; Financial Interests, Personal and Institutional, Invited Speaker: AstraZeneca. G. Pasello: Financial Interests, Institutional, Advisory Board: AstraZeneca, Boehringer Ing, MSD, Pfizer, Takeda; Financial Interests, Institutional, Invited Speaker: BMS, Eli Lilly, Novartis, Roche; Non-Financial Interests, Institutional, Principal Investigator: Amgen, AstraZeneca, Roche, Novartis, Eli Lilly. M. Fassan: Non-Financial Interests, Personal and Institutional, Research Grant: QED Therapeutics, Astellas Pharma; Financial Interests, Personal, Invited Speaker: Astellas Pharma, Tesaro, Roche, Diaceutics. V. Guarneri: Financial Interests, Personal, Advisory Board: Roche, Eli Lilly, Novartis, MSD, Gilead; Financial Interests, Personal, Invited Speaker: Eli Lilly, Novartis; Financial Interests, Institutional, Invited Speaker: Eli Lilly, Roche, BMS, Novartis, AstraZeneca, MSD, Synton Biopharmaceuticals, Merck. S. Indraccolo: Financial Interests, Personal, Invited Speaker: AstraZeneca. All other authors have declared no conflicts of interest. BackgroundICIs are widely used in aNSCLC, but reliable predictive markers are still missing. Preliminary data suggest potential role for longitudinal LB in predicting clinical outcome and identifying pts likely to derive detrimental effects. Aim of this work is to develop a model integrating clinico-pathological variables with longitudinal LB results. ICIs are widely used in aNSCLC, but reliable predictive markers are still missing. Preliminary data suggest potential role for longitudinal LB in predicting clinical outcome and identifying pts likely to derive detrimental effects. Aim of this work is to develop a model integrating clinico-pathological variables with longitudinal LB results. MethodsWe prospectively enrolled aNSCLC pts treated with ICIs at Veneto Institute of Oncology from 2017 to 2019. LB was performed at baseline (T1), after 3-4 weeks of ICI treatment (T2). Each sample was evaluated for cell free DNA (cfDNA) quantification and analysed with a 56 gene amplicon-based NGS panel. cfDNA quantification and variant allele fraction (VAF) of tumor-associated genetic alterations were evaluated as static and dynamic parameters. The genetic alteration with the highest VAF at baseline was considered as reference for NGS analysis. Variables independently associated with the clinical outcome (p<0.05) in a multiple Cox survival model were used to derive a risk score for predicting survival probabilities. We prospectively enrolled aNSCLC pts treated with ICIs at Veneto Institute of Oncology from 2017 to 2019. LB was performed at baseline (T1), after 3-4 weeks of ICI treatment (T2). Each sample was evaluated for cell free DNA (cfDNA) quantification and analysed with a 56 gene amplicon-based NGS panel. cfDNA quantification and variant allele fraction (VAF) of tumor-associated genetic alterations were evaluated as static and dynamic parameters. The genetic alteration with the highest VAF at baseline was considered as reference for NGS analysis. Variables independently associated with the clinical outcome (p<0.05) in a multiple Cox survival model were used to derive a risk score for predicting survival probabilities. ResultsOne hundred thirteen pts were included in the analysis, 57 of them were treated in first-line. cfDNA quantification at T1, VAF at T2, cfDNA change (T2-T1), PD-L1 expression and non squamous histology were significantly associated with progression-free survival (PFS); cfDNA at T2, VAF at T2, histology and PD-L1 >/=50% were associated with overall survival (OS). Pts with all favourable variables had 70% probability of 2 year-survival and 56% probability of 3 year-survival, while all unfavourable variables were associated with only 6% probability of 6 month-survival. LB at T2 is also able to identify pts at higher risk of death within 12 weeks since the start of ICIs and define prognostic groups in pts with equal radiological response. One hundred thirteen pts were included in the analysis, 57 of them were treated in first-line. cfDNA quantification at T1, VAF at T2, cfDNA change (T2-T1), PD-L1 expression and non squamous histology were significantly associated with progression-free survival (PFS); cfDNA at T2, VAF at T2, histology and PD-L1 >/=50% were associated with overall survival (OS). Pts with all favourable variables had 70% probability of 2 year-survival and 56% probability of 3 year-survival, while all unfavourable variables were associated with only 6% probability of 6 month-survival. LB at T2 is also able to identify pts at higher risk of death within 12 weeks since the start of ICIs and define prognostic groups in pts with equal radiological response. ConclusionsAssociation of clinical variables and LB performed early during ICI treatment is able to predict OS. This integrated model upon further validation could be implemented in clinical practice to personalize treatment strategies. Association of clinical variables and LB performed early during ICI treatment is able to predict OS. This integrated model upon further validation could be implemented in clinical practice to personalize treatment strategies.

The combination of immune checkpoints inhibitors (ICIs) and chemotherapy is the standard of care in untreated advanced non-small cell lung cancer (aNSCLC) patients with PD-L1 < 50% and no targetable mutations. In this work, we provide data about toxicity profile and outcome in a multicenter real-word setting.

IrAEs require prompt recognition and adequate treatment. Real-world data could help in optimizing clinical management of aNSCLC pts. We retrospectively analyzed clinical and pathological data of aNSCLC pts consecutively treated with single-agent ICIs at Veneto Institute of Oncology (IOV) between August 2013 and September 2021. We studied the association of clinical and pathological features with irAEs development by univariate and multivariable logistic regression models. 488 pts received ICIs as first (35%) or further line of treatment (75%). Median number of cycles was 6 (range 1–105). 183 pts (40.8%) developed at least one irAE; the most frequent irAEs were skin-related events (20%) and diarrhea (20%). 83 (45%) pts stopped ICIs due to irAE and 33 (18%) permanently discontinued ICIs, 14 (7.6%) required hospitalization. 90 pts (20%) developed two or more irAEs. At second irAE the rate of pts requiring ICIs interruption, discontinuation or hospitalization were similar. Improved outcome was confirmed in pts experiencing at least one irAE, regardless of line of treatment (HR for median overall survival (mOS) = .308, CI 95% .24-.39, p < .000; HR for median progression-free survival (mPFS) = .280, CI 95% .217-.360, p < .000). Multivariate analysis including clinical and pathologic variables showed low tumor burden (< 2 involved organ sites) as independent factor predicting the risk of irAE (OR = 1.85, CI 95% 1.13-3.04, p = .015). Pts experiencing skin-related events had a significantly longer mPFS and mOS than pts with other irAEs: 22.4 m vs 9.7 m (HR = .515, CI 95% .334-.796, p = .003) and 31.9 m vs 18.5 m (HR = .580, CI 95% .262-.703, p = .034). In real-world setting, the incidence of irAEs and their positive prognostic value were confirmed. Low tumor burden was associated with higher risk of irAEs. This could represent a starting point for the development of a nomogram capable to predict a global risk of irAEs and improve toxicity management.

Immune checkpoints inhibitors (ICIs) have revolutionized the treatment of advanced Non-small cell lung cancer (NSCLC). However, a proportion of patients could experience detrimental effects and no predictive markers are available for their early detection. Aim of the study is to use longitudinal liquid biopsy as a tool to identify patients experiencing hyper-progression (HPD) or death within 12 weeks (early death, ED) following ICIs.

The IM plays an important role in NSCLC development. C and R may modify the interplay between cancer and the immune system. Induction C-R is crucial in the management of SS-NSCLC. We investigated the effects of C-R on the IM. We retrospectively analyzed consecutive paired samples, pre and post C-R, of SS-NSCLC cases from 2015 to 2019. We performed PD-L1-TPS determination and definition of the rate of residual viable tumor cells (RVTC). CD3, 4, 8 and 68 tumor-infiltrating immune cells (TIICs) were evaluated and categorized using a semiquantitative score. Radiological and metabolic responses were reviewed. We calculated neutrophil-to-lymphocyte ratio (NLR) and platelet-to-lymphocyte ratio (PLR) before surgery (S). Preliminary analysis included 8 patients, one female and seven males, all smokers. They received a carboplatin-based regimen of C, concomitant to R (median dosage: 50 Gray in 28 fractions). Radiological responses were 6 disease stabilities (SD) and 2 partial responses (PR). Metabolic responses were 4 SD and 4 PR. A complete evaluation of IM in pre C-R tissue samples was not feasible because of insufficient tumor tissue. On surgical samples, no oncogene addiction was detected, PD-L1 TPS was < 1% in all cases and median rate of RVTC was 10%. All TIICs had a score equal or higher than 2+, except for CD4+ that scored 1+ in 88% of cases. No TIIC was absent at immunohistochemistry evaluation. Median NLR and PLR were 3.5 (1.7-4.6) and 199 (132-264) respectively. No correlation was observed between CD3, 4 and 8 TIICs and radiological or metabolic response, RVTC and NLR or PLR values. CD68+ TIICs was correlated with metabolic response (p=0.020, Χ2 test) and lower RVTC (Pearson's correlation coefficient, PCC=-0.7, p=0.042). CD68+ TIICs presence was associated with higher pre-surgery PLR (PCC=0.8, p=0.020). Interestingly, higher pre-S PLR values seemed linked with lower RVTC (PCC=-0.2, p=0.047). Pre-surgery PLR and CD68+ TIICs in surgical samples might be correlated to higher metabolic response to C-R and lower RVTC. These preliminary results could be useful for consolidation treatment selection and are going to be confirmed in a wider series of SS-NSCLC.

The introduction of ICIs in clinical practice has revolutionized aNSCLC treatment but a proportion of pts does not benefit from ICIs and could even derive detrimental effects. No predictive markers are currently available for their early detection. Aim of the study was to characterize pts experiencing HPD and ED with plasma NGS to analyze their modification at an early time-point during treatment. aNSCLC pts referring to Istituto Oncologico Veneto were prospectively enrolled in MAGIC-1 study: plasma was collected at baseline (T1) and after three/four weeks (w) of treatment, according to treatment schedule (T2). We selected pts experiencing ED and HPD following ICIs and randomly collected two control groups: having progressive disease (not HPD) as best radiological response and achieving clinical benefit. NGS was performed in plasma by using AVENIO ctDNA Expanded kit. Variant allele fraction (VAF) was used for relative quantification of tumor associated genetic alterations in plasma. From March 2017 to November 2019, 172 aNSCLCs receiving ICIs were enrolled. Median follow-up was 18.6 (IC95%: 9.3-26.8) months (m). Median overall survival (OS) was 12 (IC95%: 9.9-13.5) m. Median immune related PFS was 5.7 (IC95%: 4.9-6.9) m. Five cases matched criteria for HPD and 31 experienced ED; one overlapped. Median OS of HPD pts was 3.8 (95%CI: 3.6-N.A) m versus 12.1 (95%CI: 10.1-14.1) m of non-HPD pts (p:0.01). The presence of liver or brain metastases increased the risk for HPD/ED (OR: 3.01, p:0.026; OR: 3.03, p:0.033). Till now we analyzed plasma NGS results of all HPD pts, five ED pts and seven cases in each control group. At baseline we observed one RET/KIF5B fusion among HPD pts. Mean fold change (VAF T2/T1) was significantly different among the four groups (p:0.008). Mean fold change (VAF T2/T1) was significantly higher for pts experiencing HPD/ED versus control cases (p:0.037). In our study population longitudinal NGS analysis is able to discriminate pts potentially deriving detrimental effect (HPD, ED) from ICIs by highlighting VAF increase after three/four w of treatment.

Background: Immune checkpoint inhibitors (ICIs) have radically changed the treatment of aNSCLC patients (pts). IrAEs are mainly reversible but they require timely recognition and management and may be potentially life-threatening. No predictive markers are available to predict the onset of irAEs and their risk of recurrence. Aim of the study was to evaluate the potential role of circulating markers in predicting the development of irAEs. Methods: We retrospectively reviewed clinical data of aNSCLC pts consecutively treated with ICIs at Istituto Oncologico Veneto (Padua, Italy) and at San Bortolo General Hospital (Vicenza, Italy) between August 2013 and August 2018. We collected data on the type, grading (G) and timing of irAEs and calculated NLR and PLR before first ICI administration and at the onset of the irAEs. The values were dichotomized for analysis in two groups: high (H-) and low (L-) NLR and H- and L-PLR using pre-identified cut-offs of 3 and 180, respectively. Results: The analysis included 194 pts. ICIs were administered as first-line treatment in 29 (15%) pts. Median number of ICIs cycles was 6 (range: 1-66). Median PFS and OS were 4.6 months (m) (95% CI: 3.3-5.9) and 8.7m (95% CI: 6.1-11.4). Seventy-two pts (37%) developed irAEs, mainly G1-2 (68%), with permanent discontinuation of ICI in 23 (32%) cases; 15 pts (20.8%) experienced more than one irAE. Median time to irAE onset was 77 days (7-751 days). Pts with baseline L-NLR and L-PLR values had a higher risk of irAE development (OR = 1.98, 95% CI: 1.09-3.61, p = 0.025 and OR = 2.15, 95% CI: 1.16-3.98, p = 0.016). Multivariate analysis confirmed PLR as independent predictive marker (OR = 2.01, 95%CI: 1.02-3.96; p = 0.045). Among pts who experienced irAE, L-PLR at time of irAE onset was associated with the risk of irAE recurrence or second irAE development (OR = 4.5, 95% CI: 1.11-18.16, p = 0.035). Conclusions: Baseline evaluation of NLR and PLR may be a tool to predict the risk of irAE onset and to personalize clinical follow-up during and after treatment with ICIs. The evaluation of PLR at the time of irAE onset might predict the risk of further toxicity. If validated, PLR value may support the clinical decision of re-introducing ICIs after discontinuation. Legal entity responsible for the study: Istituto Oncologico Veneto (IOV), Padua, Italy. Funding: Has not received any funding. Disclosure: All authors have declared no conflicts of interest.

Background: The introduction of immune-checkpoint inhibitors (ICIs) in the management of aNSCLC has led to great outcome improvement, but reliable predictive biomarkers are still a need. Liquid biopsy has the potential to monitor biological effects of treatment. Aim of the study is to explore the potential predictive value of its dynamic analysis in aNSCLC treated with ICIs. Methods: aNSCLC patients consecutively treated with ICIs at Istituto Oncologico Veneto were prospectively enrolled and genotyped in tissue. Plasma samples were collected at baseline (T1), after three or four weeks according to the administration schedule (T2) and at the moment of the first radiological evaluation (T3). Patients carrying KRAS mutation in tissue were analyzed in plasma with droplet digital PCR (ddPCR). Semiquantitative index of fractional abundancy of mutated allele (MAFA) was used. Results: aNSCLC patients (N: 54) were prospectively enrolled and tissue genotyped, 24 of them carried KRAS mutation in tissue and 11 (46%) were positive in plasma at baseline. Positivity was not associated with tumor burden or other clinical features. We evaluated the impact of the presence and the quantitative variation of MAFA during treatment on outcome in terms of progression free-survival (PFS). After a median follow-up of 10.9 months, the presence of sentinel mutation at T1 did not affect PFS, whereas the MAFA increase from baseline to T2 and to T3 were associated with shorter PFS (HR: 5.9, 95%CI: 1.2-27.7, p:0.02 and HR: 12.1, 95%CI: 2.3-23.8, p:0.003, respectively). Median PFS was 5.2 months in the presence of MAFA increase T1-2, while it was not reached in case of decreased/stable MAFA. Conclusions: Increase in MAFA from baseline to three or four weeks after the start of ICI is associated with shorter PFS. Predictive value of dynamic analysis of sentinel mutations in plasma during ICIs treatment warrants further validation in aNSCLC. Legal entity responsible for the study: Istituto Oncologico Veneto. Funding: Istituto Oncologico Veneto. Disclosure: All authors have declared no conflicts of interest.

BACKGROUND:Targeted agents have improved the outcome of a subset of non-small cell lung cancer (NSCLC). Molecular profiling by next-generation sequencing (NGS) allows screening for multiple genetic alterations both in tissue and in plasma, but limited data are available concerning its feasibility and impact in real-world clinical practice.METHODS:Patients with advanced NSCLC consecutively referring to our Institution for potential eligibility to VISION trial (NCT02864992) were prospectively enrolled. They were already screened with standard method, and EGFR/ALK/ROS-1 positive cases were excluded. NGS was performed in plasma and tissue using the Guardant360 test covering 73 genes and the Oncomine Focus Assay covering 59 genes, respectively.RESULTS:The study included 235 patients. NGS was performed in plasma in 209 (88.9%) cases; 78 of these (37.3%) were evaluated also in tissue; tissue only was analyzed in 26 cases (11.1%). Half of the tissue samples were deemed not evaluable. Druggable alterations were detected in 13 (25%) out of 52 evaluable samples and 31 of 209 (14.8%) of plasma samples. Improved outcome was observed for patients with druggable alterations if treated with matched targeted agents: they had a longer median overall survival (not reached) compared with the ones who did not start any targeted therapy (9.1 months; 95% confidence interval, 4.6-13.6; p = .046). The results of NGS testing potentially also affected the outcome of patients treated with immunotherapy.CONCLUSION:Systematic real-life NGS testing showed the limit of tissue analysis in NSCLC and highlighted the potentiality of genetic characterization in plasma in increasing the number of patients who may benefit from NGS screening, both influencing the clinical decision-making process and affecting treatment outcome.IMPLICATIONS FOR PRACTICE:Genetic characterization of cancer has become more important with time, having had positive implications for treatment specificity and efficacy. Such analyses changed the natural history of advanced non-small cell lung cancer (aNSCLC) with the introduction of drugs targeted to specific gene alterations (e.g., EGFR mutations, ALK and ROS-1 rearrangements). In the field of cancer molecular characterization, the applicability of the analysis of a wide panel of genes using a high-throughput sequencing approach, such as next-generation sequencing (NGS), is still a matter of research. This study used NGS in a real-world setting to systematically and prospectively profile patients with aNSCLC. The aim was to evaluate its feasibility and reliability, as well as consequent access to targeted agents and impact on clinical outcome whenever a druggable alteration was detected either in tumor tissue samples or through liquid biopsy.

Background Immune checkpoint inhibitors (ICIs) have changed the treatment of pts with aNSCLC and mMel. No predictive markers of development of irAEs are available. Aim of the study is to evaluate the role of circulating markers in predicting irAE onset. Methods We reviewed clinical data of aNSCLC and mMel pts treated with ICIs at Istituto Oncologico Veneto (Padova, Italy) and San Bortolo Hospital (Vicenza, Italy) between January 2012 and January 2019. We collected data on type and grading (G) of irAEs and calculated neutrophil-to-lymphocyte ratio (NLR) and platelet-to-lymphocyte ratio (PLR) before first ICI administration and at irAE onset. Values were dichotomized in: high (H) and low (L) NLR and H- and L-PLR, using pre-identified cut-offs of 3 and 180 for aNSCLC, 3 and 120 for mMel. Results Analysis included 377 pts (252 aNSCLC and 125 mMel). In aNSCLC cohort, median PFS and OS were 4.9 months (m) (95% CI: 3.6-6.1) and 8.6m (95% CI: 6.3-10.8). Ninety-seven pts (38%) developed irAEs, mainly G1-2 (72%), with permanent ICI discontinuation in 29 (29.9%) cases; 26 pts (26.8%) experienced more than one irAE. Pts with baseline L-NLR or L-PLR had a higher risk of irAE (OR = 2.3, 95% CI: 1.3-3.9, p = 0.002 | OR = 2.4, 95% CI: 1.3-4.1, p = 0.02). Multivariate analysis confirmed NLR and PLR as independent predictive markers (OR = 1.8, 95%CI: 1.0-3.2, p = 0.04 | OR = 1.9, 95%CI: 1.0-3.4, p = 0.03). L-PLR at irAE onset was associated with risk of irAE recurrence or second irAE development (OR = 4.2, 95% CI: 1.4-12.9, p = 0.01). In mMel pts, median PFS and OS were 5.1m (95% CI: 3.6-6.5) and 18.1m (95% CI: 11-25.2). Fifty-four pts (43%) developed irAEs, mainly G1-2 (76%), with permanent ICI discontinuation in 10 (18.5%) cases; 14 pts (25.9%) had multiple irAEs. Pts with baseline L-NLR had higher risk of irAE (OR = 2.2, 95% CI: 1.1-4.6, p = 0.04). NLR and PLR at time of irAE onset were not associated with the risk of irAE recurrence or second irAE development. Conclusions Baseline NLR and PLR may be reliable and inexpensive predictive tools of irAE risk. For aNSCLC pts L-PLR at irAE onset correlates with risk of further toxicity. If validated, these biomarkers may help pts’ management during ICIs and treatment handling after a first irAE. Legal entity responsible for the study IRCCS Istituto Oncologico Veneto - IOV - Padua – Italy. Funding Has not received any funding. Disclosure All authors have declared no conflicts of interest.

Background: SCLC represents one of the most aggressive lung malignancies, characterized by a high growth fraction and early metastatic spread. New therapeutic options are badly needed and immunotherapy might represent a promising approach. Unfortunately, so far, no molecular prognostic markers have been validated for clinical practice and data on immune microenvironment are limited. Methods: We have retrospectively analyzed 104 SCLC cases. Immunohistochemistry evaluation of PD-L1 (22C3 clone, DAKO) was performed on tumor cells (TCs) and on tumor-infiltrating lymphocytes (TILs): positivity was defined as PD-L1 expression on 1% or more TCs or TILs. Immunohistochemistry for CD8 (C8/144B clone, DAKO) and FOXP3 (236A/E7 clone, ABCAM) was also performed. A semiquantitative score was used and CD8 and FOXP3 TILs categorized as positive versus negative. Results: The analysis included 104 patients: 48 surgically resected, 18 patients treated with radical-intent chemoradiotherapy and 38 metastatic. In overall study population, PD-L1 was expressed in TCs in 25% of cases. The expression of PD-L1 was significantly correlated with stage disease (32% stage I-III; 13% metastatic stage; p:0.034 for TCs and p:0.002 for TILs) and with outcome: median OS 46.8 months (m) (95% CI: 22.6 to 71.0) versus (vs) 10.9 m (95% CI: 6.2 to 15.7; p = 0.047) PD-L1 positive vs negative respectively; the relation with outcome however, was not confirmed in multivariate analysis. CD8-positive TILs and FOXP3-positive TILs were present in 59% and 72% of samples respectively. Neither the presence of CD8+ TILs nor that of FOXP3+ TILs was correlated to stage. The presence of FOXP3-positive TILs was associated with improved prognosis among non-metastatic patients: median OS 52.5 m (95% CI: 21.4 to 83.7) vs 20.5 m (95% CI: 0 to 49.2; p = 0.027) FOXP3-positive vs negative TILs, a relation confirmed in multivariate analysis. Conclusions: Expression of PD-L1 is reduced in advanced stage SCLC patients. Further studies are needed to understand if down-regulation of PD-L1 is linked to a more aggressive phenotype. The prognostic role of FOXP3 TILs in stage I-III SCLCs warrants further confirmation in larger series of patients. Legal entity responsible for the study: Istituto Oncologico Veneto (IOV), Padua, Italy Funding: Università degli Studi di Padova Disclosure: All authors have declared no conflicts of interest.

Background Tumor immune microenvironment (TME) plays a key role in malignant pleural mesothelioma (MPM) pathogenesis and treatment outcome, supporting a role of immune checkpoint inhibitors as anticancer approach. This study retrospectively investigated TME and programmed death ligand 1 (PD-L1) expression in naïve MPM cases and their change under chemotherapy. Patients and methods Diagnostic biopsies of MPM patients were collected from four Italian and one Slovenian cancer centers. Pathological assessment of necrosis, inflammation, grading, and mitosis was carried out. Ki-67, PD-L1 expression, and tumor infiltrating lymphocytes were detected by immunohistochemistry. When available, the same paired sample after chemotherapy was analyzed. Pathological features and clinical characteristics were correlated to overall survival. Results TME and PD-L1 expression were assessed in 93 and 65 chemonaive MPM samples, respectively. Twenty-eight samples have not sufficient tumor tissue for PD-L1 expression. Sarcomatoid/biphasic samples were characterized by higher CD8+ T lymphocytes and PD-L1 expression on tumor cells, while epithelioid showed higher peritumoral CD4+ T and CD20+ B lymphocytes. Higher CD8+ T lymphocytes, CD68+ macrophages, and PD-L1 expression were associated with pathological features of aggressiveness (necrosis, grading, Ki-67). MPM cases characterized by higher CD8+ T-infiltrate showed lower response to chemotherapy and worse survival at univariate analysis. Patients stratification according to a combined score including CD8+ T lymphocytes, necrosis, mitosis, and proliferation index showed median overall survival of 11.3 months compared with 16.4 months in cases with high versus low combined score (P < 0.003). Subgroup exploratory analysis of 15 paired samples before and after chemotherapy showed a significant increase in cytotoxic T lymphocytes in MPM samples and PD-L1 expression in immune cells. Conclusions TME enriched with cytotoxic T lymphocytes is associated with higher levels of macrophages and PD-L1 expression on tumor cells and with aggressive histopathological features, lower response to chemotherapy and shorter survival. The role of chemotherapy as a tumor immunogenicity inducer should be confirmed in a larger validation set.