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Hormone-receptor positive (HR+) breast cancer (BC) (including the luminal A and the luminal B subtypes) is the most common type of tumor in women diagnosed with early-stage BC (EBC). It represents a highly heterogeneous subgroup that is characterized by different risks of relapse. The aim of this review is to discuss the possible role played by the immune response in predicting this risk, along with the most common clinical and pathological factors and molecular tools that have been developed and are already in use. As opposed to what has previously been observed in the most aggressive human epidermal growth factor receptor 2 (HER2)-positive and triple-negative breast cancer (TNBC) subtypes, a high proportion of tumor-infiltrating lymphocytes (TILs)-reflecting a spontaneous and pre-existing immune response to the tumor-has been linked to a worse prognosis in HR+ EBC. This work provides some immune biological rationale explaining these findings and provides the basics to understand the principal clinical trials that are testing immunotherapy in HR+ (luminal) BC.

Immunotherapy approaches boosting spontaneous and durable antitumor immune responses through immune checkpoint blockade are revolutionizing treatment and patient outcomes in solid tumors and hematological malignancies. Among the various inhibitory molecules employed by the immune system to regulate the adaptive immune responses, cytotoxic T lymphocyte antigen-4 (CTLA-4) is the first successfully targeted immune checkpoint molecule in the clinic, giving rise to significant but selective benefit either when targeted alone or in combination with anti-programmed cell death protein-1 (PD-1) antibodies (Abs). However, the use of anti-CTLA-4 Abs was associated with the incidence of autoimmune-like adverse events (AEs), which were particularly frequent and severe with the use of combinational strategies. Nevertheless, the higher incidence of AEs is associated with an improved clinical benefit indicating treatment response. A prompt recognition of AEs followed by early and adequate treatment with immunosuppressive agents allows the management of these potentially serious AEs. This narrative review aims to summarize CTLA-4 biology, the rationale for the use as a companion for anti-PD-1 Abs in humans with results from the most relevant Phase III clinical trials including anti-CTLA-4 Abs in combination with anti-PD-1 Abs in solid tumors.

Graphical abstract Physiology and pathophysiology of pulmonary alveoli during SARS-CoV-2 infection. (A) COVID-19 severity: frequency of degrees of clinical manifestations in a Chinese cohort (>44.000 patients) (ref: Wu Z. et al, 2020). (B) Phases of COVID-19 disease and timing for the use of glucocorticoids. Legend: ARDS: Adult Respiratory Distress Syndrome; COVID-19: CoronaVirus Disease-19; MOF: multi-organ failure; pts: patients; SIRS: systemic inflammatory response syndrome., Highlights • Dexamethasone reduces the risk of death in patients critically ill with COVID-19. • No evidence of benefit from other complementary and/or supportive treatments was seen in critically ill COVID-19 hospitalized patients. • Synthetic glucocorticoids switch off cytokine storm, and consequent severe respiratory and multi-organ failures in patients with COVID-19. • Effects of synthetic glucocorticoids might be confounded by the contemporary use of other complementary drugs., The viral infection by SARS-CoV-2 has irrevocably altered the life of the majority of human beings, challenging national health systems worldwide, and pushing researchers to rapidly find adequate preventive and treatment strategies. No therapies have been shown effective with the exception of dexamethasone, a glucocorticoid that was recently proved to be the first life-saving drug in this disease. Remarkably, around 20 % of infected people develop a severe form of COVID-19, giving rise to respiratory and multi-organ failures requiring subintensive and intensive care interventions. This phenomenon is due to an excessive immune response that damages pulmonary alveoli, leading to a cytokine and chemokine storm with systemic effects. Indeed glucocorticoids’ role in regulating this immune response is controversial, and they have been used in clinical practice in a variety of countries, even without a previous clear consensus on their evidence-based benefit.

Programmed cell death-1 (PD-1) receptor and its ligands physiologically regulate the activity of the adaptive immune system to limit excessive inflammatory processes, thus preventing normal tissue damage. Tumor cells escape from the host immune surveillance using this pathway, rendering it relevant therapeutic target. Despite the relevant clinical efficacy observed in patients with solid and hematological malignancies, the clinical benefit of these novel treatments is limited to a relatively restricted number of patients. A wide amount of genomic and immune related features is currently under investigation as potential predictive biomarkers for treatment selection. The results obtained so far are encouraging but still imperfect. Combination strategies using different immunotherapeutic agents or with other treatments (such as chemotherapy) are being investigated, showing promising but still not completely satisfactory results. This review aims to shed light on the main principles of targeting PD-1 in breast cancer, from biology through its functional and clinical implications.

The improved prognosis of breast cancer patients makes survivorship issues an area of crucial importance. In this regard, an increased attention is needed toward the development of potential anticancer treatment-related long-term side-effects, including gonadal failure and infertility in young women. Therefore, fertility preservation and family planning are crucial issues to be addressed in all young women of reproductive age with newly diagnosed cancer. Despite a growing availability of data on the efficacy and safety of fertility preservation options and the fact that conceiving after prior history of breast cancer has become more accepted over time, there are still several gray zones in this field so that many physicians remain uncomfortable to deal with these topics. The purpose of this review is to answer some of the most controversial questions frequently asked by patients during their oncofertility counseling, in order to provide a detailed and up-to-date overview on the evidence available in this field to physicians involved in the care of young women with breast cancer.

Background Venous (VTEs) and arterial thromboembolic events (ATEs) are causes of morbidity, disability, mortality, and increase in treatment costs in cancer patients. The risk associated with immune checkpoint inhibitors (ICIs) has not yet been clarified. The primary objective of this systematic review was to evaluate the incidence of VTEs and ATEs in patients treated with ICIs as single agents or in combination with other treatments. Material and methods Data from retrospective and prospective studies were selected from PubMed, EMBASE, SCOPUS, and The Cochrane Library from inception up to May up to 21st May 2020. All studies had to be in English and use human study participants. The studies were eligible if they provided a number (or rate) of VTEs and ATEs and the size of the population included. The PRISMA guidelines were followed. The data on the incidence of VTEs and ATEs were extracted for each arm, analyzed using random-effects models, and reported as weighted measures. Results A total of 20,273 patients from 68 studies were included (median follow-up ranged from a few months up to three years). Overall, there were 390 VTEs and 59 ATEs, with incidence rates of 2.7% (95%CI 1.8%–4%) and 1.1% (95%CI 0.5%–2.1%), respectively. The rate of pulmonary embolism was 1.6% (95%CI 0.7%–3.2%) and deep venous thrombosis was 2.7% (95%CI 1.4%–5.4%). In studies where ICIs were administered with chemotherapy, rates of VTEs were similar to ICI alone arms (2.8% vs 2.5%). The rate of stroke and myocardial infarction were 1.1% (95%CI 0.65%–1.45%) and 0.7% (95%CI 0.15%–1.15%), respectively. In randomized trials, compared with non-ICIs containing arms (e.g. chemotherapy), the relative risk (RR) of VTEs due to ICIs was similar (RR 1.08, 95%CI 0.6–1.9; P = .79). Conclusions Thromboembolic events associated with ICIs are relatively rare in cancer patients with an advanced stage of the disease. However, in randomized studies, their incidence is similar to control arms, suggesting that the contributory role of ICIs to the thromboembolic risk in many cancer settings is small.

Background: FOXP1, a transcriptional regulator of lymphocyte development, is abnormally expressed in some human tumors. This study investigated FOXP1-mediated regulation of tumor infiltrating lymphocytes (TIL) in untreated primary breast cancer (BC). Methods: FOXP1 expression was analyzed in tissues from primary untreated breast tumors, BC cell lines and the METABRIC gene expression BC dataset. Cytokine and chemokine expression and lymphocyte migration in response to primary tumor supernatants (SN) was compared between FOXP1hi and FOXP1lo primary BC. Finding: FOXP1 expression was higher in estrogen receptor positive compared to negative BC. FOXP1hi tumors were significantly associated with lower TIL and fewer tertiary lymphoid structures (TLS) compared to FOXP1lo BC. Silencing FOXP1 in BC cell lines positively impacted cytokine and chemokine expression with the inverse effect associated with overexpression. CXCL9, CXCL10, CXCL11, CXCL13, CX3CL, CCL20, IL2, IL21, GZMB and IFNG expression decreased while IL10 and TGFβ increased in FOXP1hi compared to FOXP1lo primary BC. Lymphocyte migration using primary BC supernatants detected decreased mobility toward FOXP1hi supernatants. FOXP1lo BC expresses higher levels of chemokines driving TIL migration. The METABRIC gene expression dataset analysis show FOXP1 expression is associated with unfavorable BC outcomes. Interpretation: These data identify FOXP1 as an important negative regulator of immune responses in BC via its regulation of cytokine and chemokine expression. Fund: Belgian Fund for Scientific Research (FNRS 3.4513.12F) and Opération Télévie (7.4636.13F and 7.4609.15F), Fonds J.C. Heuson and Fonds Lambeau-Marteaux., SCOPUS: ar.j, info:eu-repo/semantics/published

Background: We hypothesized that non-small cell lung cancer (NSCLC) patients with a tumor positive for single nucleotide polymorphisms (SNPs) of the Excision Repair Cross Complementation Group 1 (ERCC-1) gene could be more genetically instable and consequently more responsive to a programmed cell death-1 (PD-1) blockade. Methods: We evaluated the T19007C and C8092A ERCC-1 SNPs by pyrosequencing assay, on tumor specimens from two independent cohorts of patients who relapsed after one or more prior systemic treatments for advanced NSCLC and who received nivolumab (3 mg/kg intravenously every 2 weeks) as part of the Italian Expanded Access Program. We aimed to assess the outcome of enrolled subjects according to the ERCC-1 SNPs status, to evaluate the role of these polymorphisms as putative biomarkers associated with a response/clinical benefit to anti-PD-1 therapies. Results: Of the 45 patients included in the final analysis, 21 (47%) and 16 (36%) were positive for the T19007C and C8092A polymorphic genotype (PG), respectively. In univariate analyses, overall survival (OS) and progression free survival (PFS) were shorter in patients with the T19007C PG, but neither difference achieved statistical significance (P = 0.131 and P = 0.717, respectively). The presence of the C8092A PG was associated with a longer OS and PFS, although statistical significance was only reached for PFS (P = 0.112 and P = 0.025, respectively). These results were confirmed by multivariate analyses. The response rate was only significantly higher in patients with the C8092A PG vs. wild type ERCC-1 (62 vs. 7%, P < 0.001). Conclusions: Results from this hypothesis generating pilot study, provided suggestive evidence that a subgroup of NSCLC patients could benefit differently from nivolumab according to the C8092A ERCC-1 SNP status. However, these data warrant further investigation., SCOPUS: ar.j, info:eu-repo/semantics/published

The extent of tumor-infiltrating lymphocytes (TILs), along with immunomodulatory ligands, tumor-mutational burden and other biomarkers, has been demonstrated to be a marker of response to immune-checkpoint therapy in several cancers. Pathologists have therefore started to devise standardized visual approaches to quantify TILs for therapy prediction. However, despite successful standardization efforts visual TIL estimation is slow, with limited precision and lacks the ability to evaluate more complex properties such as TIL distribution patterns. Therefore, computational image analysis approaches are needed to provide standardized and efficient TIL quantification. Here, we discuss different automated TIL scoring approaches ranging from classical image segmentation, where cell boundaries are identified and the resulting objects classified according to shape properties, to machine learning-based approaches that directly classify cells without segmentation but rely on large amounts of training data. In contrast to conventional machine learning (ML) approaches that are often criticized for their “black-box” characteristics, we also discuss explainable machine learning. Such approaches render ML results interpretable and explain the computational decision-making process through high-resolution heatmaps that highlight TILs and cancer cells and therefore allow for quantification and plausibility checks in biomedical research and diagnostics., SCOPUS: re.j, info:eu-repo/semantics/published

Side effects of immune checkpoint blockade are often said to be infrequent and usually mild. The uniqueness of endocrine immune-related adverse events is their non-reversibility, with incidence and prevalence destined to increase in the coming years, particularly if immunotherapy is used at earlier stages of neoplastic disease. Immune-related hypophysitis is one of these observed endocrine adverse events. It is often difficult to diagnose, sometimes occurring without specific symptoms. It can lead to irreversibly altered functioning of diverse endocrine glands. Radiographically, the differential diagnosis of hypophysitis includes pituitary apoplexy and primary and secondary neoplastic lesions. Immune-related hypophysitis is most common with single-agent anti-CTLA-4, followed by the combination of anti-CTLA-4 and anti-PD-1, while occurs infrequently when anti-PD-1 or anti-PD-L1 agents are administered alone. Hypophysitis with immune checkpoint blockade requires early recognition, diagnosis, and treatment. Patients can present with headache, visual disturbances or other endocrine-related syndromes or they can be asymptomatic. The manifestation of symptoms should prompt blood analysis and magnetic resonance imaging of the brain. Imaging is important to exclude secondary meningeal or parenchymal lesions. Management should include discontinuation of the immune checkpoint blockade, initiation of corticosteroid therapy and eventually hormone replacement therapy. Hypophysitis impacts treatment of the disease and usually requires long-term management of this irreversible side effect. A multidisciplinary team approach is merited to insure the correct diagnosis and management of immune-related hypophysitis.

Radiotherapy is one of the main treatment strategies used in cancer. Aside from the local control of the disease, which is mediated by a direct cytotoxic effect on tumor cells, radiotherapy has also been shown to exert immune-mediated local and systemic effects. Radiotherapy can elicit anti-tumor responses in distant sites from the radiation field; this phenomenon is known as the abscopal effect and has been described in patients previously treated with immune checkpoint blockade (ICB). Considering that the efficacy of immunotherapy has been demonstrated only in a subset of patients-who often benefit with lasting responses-efforts are ongoing to potentiate its activity with the development of new combination strategies. Radiotherapy might represent a potential candidate for a synergistic combination with immunotherapy, by improving the immunogenicity of tumors and by enhancing local and systemic immune effects. This review aims to summarize the current pre-clinical and clinical data on the immune effects of radiotherapy and their potential implications for cancer immunotherapy.

Background A relationship between high baseline tumor-infiltrating lymphocytes (TIL) and better outcomes has been described in early-stage HER2-positive breast cancer. Nevertheless, the magnitude of this association and whether this effect could differ based on the type of anti-HER2 agent administered remain controversial. This meta-analysis investigated the association between baseline TIL and pathologic complete response (pCR) rates in HER2-positive breast cancer patients treated with neoadjuvant chemotherapy plus trastuzumab and lapatinib either alone or in combination. Methods A literature search covering PubMed, Embase and the Cochrane library up to October 31, 2016 identified randomized, controlled trials investigating neoadjuvant chemotherapy plus trastuzumab and lapatinib either alone or in combination where published data for pCR based on pre-treatment TIL scores was available. Two subgroups were considered: high baseline TIL vs. non-high TIL, according to each study definition. Summary risk estimates (odds ratio) and 95% confidence intervals (CI) were calculated for pCR using pre-treatment TIL levels for each trial. Pooled analyses were conducted using random and fixed effects models. Interaction P -values were computed using a Monte Carlo permutation test. Results A total of 5 studies ( N =1,256 patients) were included. Overall, high TIL subgroup was associated with a significantly increased pCR rate (OR 2.46; 95% CI 1.36-4.43; P =0.003). No interaction was observed between TIL subgroup (high vs. non-high TIL) and response to anti-HER2 agent(s) (trastuzumab vs. lapatinib vs. their combination; P =0.747) and chemotherapy (anthracycline and taxanes vs. taxanes only; P =0.201). A stronger association between high TIL subgroup and pCR rate was observed when examining only the 4 studies using neoadjuvant anthracycline- and taxane- based chemotherapy and the 60% cut-off for high TIL ( N =869, NeoALTTO excluded) with an OR of 2.88 (95% CI 2.03-4.08; P Conclusions In HER2-positive breast cancer, high baseline TIL are associated with increased pCR probability irrespective of anti-HER2 agent(s) and neoadjuvant chemotherapy regimen used.

The immunotherapy revolution in cancer treatment involves a variety of specialists, not only oncologists, but also internal medicine physicians, endocrinologists, dermatologists, gastroenterologists, rheumatologists, and radiologists, introducing new scenarios and novel challenges in the diagnosis and management of a number of novel immune-related adverse events. Among these, immune checkpoint inhibitor-induced pancreatic injury has been described (occurring in up to 4% of patients) and has been reported to be responsible for visits to the emergency departments in up to 1.9% of patients treated with immune checkpoint inhibitors. This side effect can be symptomatic or non-symptomatic, and can be associated with the development of long-term damage to the pancreas, requiring the involvement of different specialists, including radiologists and gastroenterologists in the multidisciplinary team that manages these patients. The aim of this narrative review is to provide a summary of the available literature related to immune checkpoint inhibitor-induced pancreatic injury including the epidemiology, the clinical findings, and the management algorithm for diagnosis with a detailed analysis of the differential diagnosis at imaging, and treatment. A more in-depth focus is dedicated to symptomatic acute pancreatitis with its peculiar findings at imaging (ultrasound, computed tomography, and magnetic resonance imaging).

Tumor-infiltrating B-cells (TIL-B) in breast cancer (BC) have previously been associated with improved clinical outcomes; however, their role(s) in tumor immunity is not currently well known. This study confirms and extends the correlation between higher TIL-B densities and positive outcomes through an analysis of HER2-positive and triple-negative BC patients from the BIG 02-98 clinical trial (10yr mean follow-up). Fresh tissue analyses identify an increase in TIL-B density in untreated primary BC compared to normal breast tissues, which is associated with global, CD4+ and CD8+ TIL, higher tumor grades, higher proliferation and hormone receptor negativity. All B-cell differentiation stages are detectable but significant increases in memory TIL-B are consistently present. BC with higher infiltrates are specifically characterized by germinal center TIL-B, which in turn are correlated with TFH TIL and antibody-secreting TIL-B principally located in tertiary lymphoid structures. Some TIL-B also interact directly with tumor cells. Functional analyses reveal TIL-B are responsive to BCR stimulation ex vivo, express activation markers and produce cytokines and immunoglobulins despite reduced expression of the antigen-presenting molecules HLA-DR and CD40. Overall, these data support the concept that ongoing humoral immune responses are generated by TIL-B and help to generate effective anti-tumor immunity at the tumor site. ispartof: JCI INSIGHT vol:4 issue:18 ispartof: location:United States status: published

The programmed cell death 1 (PD-1) pathway is an important regulator of immune responses in peripheral tissues, including abnormal situations such as the tumor microenvironment. This pathway is currently the principal target for immunotherapeutic compounds designed to block immune checkpoint pathways, with these drugs improving clinical outcomes in a number of solid and hematological tumors. Medical oncology is experiencing an immune revolution that has scientists and clinicians looking at alternative, non-redundant inhibitory pathways also involved in regulating immune responses in cancer. A variety of targets have emerged for combinatorial approaches in immune checkpoint blockade. The main purpose of this narrative review is to summarize the biological role of lymphocyte activation gene 3 (LAG3), an emerging targetable inhibitory immune checkpoint molecule. We briefly discuss its role in infection, autoimmune disease and cancer, with a more detailed analysis of current data on LAG3 expression in breast cancer. Current clinical trials testing soluble LAG3 immunoglobulin and LAG3 antagonists are also presented in this work., SCOPUS: ar.j, info:eu-repo/semantics/published

The broader use of immune checkpoint blockade in clinical routine challenges clinicians in the diagnosis and management of side effects which are caused by inflammation generated by the activation of the immune response. Nearly all organs can be affected by immune-related toxicities. However, the most frequently reported are: fatigue, rash, pruritus, diarrhea, nausea/vomiting, arthralgia, decreased appetite and abdominal pain. Although these adverse events are usually mild, reversible and not frequent, an early diagnosis is crucial. Immune-related pulmonary toxicity was most frequently observed in trials of lung cancer and of melanoma patients treated with the combination of the anti-cytotoxic T lymphocyte antigen (CTLA)-4 and the anti-programmed cell death-1 (PD-1) antibodies. The most frequent immune-related adverse event in the lung is represented by pneumonitis due to the development of infiltrates in the interstitium and in the alveoli. Clinical symptoms and radiological patterns are the key elements to be considered for an early diagnosis, rendering the differential diagnosis crucial. Diagnosis of immune-related pneumonitis may imply the temporary or definitive suspension of immunotherapy, along with the start of immuno-suppressive treatments. The aim of this work is to summarize the biological bases, clinical and radiological findings of lung toxicity under immune checkpoint blockade, underlining the importance of multidisciplinary teams for an optimal early diagnosis of this side effect, with the aim to reach an improved patient care., SCOPUS: re.j, info:eu-repo/semantics/published

Simple Summary This work aims to discuss how an anti- or pro-tumor immune response could be manipulated through immune checkpoint blockade in patients with early stage HER2-positive breast cancer. By summarizing previously published evidence in the field, authors present their personal view on how immune checkpoint blockade could be implemented in the neoadjuvant setting in this patient population. The hypothesis being presented is that an appropriate and effective administration of immune checkpoint blockade could assure a lasting control of the disease, by preventing relapses. One of the research priorities should be the identification of the patients who could benefit more by this strategy. Abstract The present commentary synthesizes the current evidence on the role of the immune response in HER2-positive breast cancer. It points out the strengths and weaknesses of the findings observed so far, particularly in the early setting, including the clinical significance of scoring tumor-infiltrating lymphocytes. A figure proposing research hypotheses for the implementation of immune checkpoint blockade use for patient candidates to neoadjuvant treatment is presented.

Chronic inflammation following infections, autoimmune diseases or exposure to environmental irritants plays a crucial role in tumor development and influences the host immune response to neoplastic cells. The presence of an anti-tumor immune infiltrate is often associated with better outcomes in gastro-intestinal primary cancers, particularly in those with high microsatellite instability (MSI-H). Immunotherapeutic drugs inhibiting the PD-1 and PD-L1 pathway showed promising results in the treatment of these patients in the metastatic setting. The aim of this review is to resume the role tumor infiltrating lymphocytes (TILs) play in gastrointestinal tumors, underlining their potential value as a prognostic and predictive biomarker. TILs assessment could identify subsets of patients with high extent of TILs and better prognosis, that could be spared from adjuvant systemic treatments. Immune infiltration parameters might be additional predictors of a greater benefit from the immunotherapy with the immune checkpoint blockade.

Anti-HER2 targeted therapy is one of the key advances in the treatment of breast cancer that have occurred in the last 20 years. In the adjuvant setting, the use of trastuzumab has led to prolonged and sustained survival benefit with very little toxicity as also confirmed by the 10-year follow-up results from the pivotal trials. Despite the survival improvement, several key issues are not entirely resolved in this field. These issues have led to multiple research efforts in de-escalating or escalating the standard treatment with chemotherapy and 1 year of adjuvant trastuzumab. Areas covered: In this paper, we present an in depth overview on the state of the art on these key issues of refining decision-making in adjuvant anti-HER2 therapy. Expert commentary: Despite many important research efforts in the field, chemotherapy plus trastuzumab for a total duration of 1 year remains the standard of care. However, recent data showed that besides standard anthracycline- and taxane-based cytotoxic therapy, alternative chemotherapy regimens can now be proposed to patients with small tumors without nodal involvement and to women at high-risk of developing cardiotoxicity. Of note, besides HER2 itself, biomarkers predicting patients who may truly benefit from anti-HER2 agents are still lacking.

In recent years the concept of precision medicine has become a popular topic particularly in medical oncology. Besides the identification of new molecular prognostic and predictive biomarkers and the development of new targeted and immunotherapeutic drugs, imaging has started to play a central role in this new era. Terms such as "radiomics", "radiogenomics" or "radi…-omics" are becoming increasingly common in the literature and soon they will represent an integral part of clinical practice. The use of artificial intelligence, imaging and "-omics" data can be used to develop models able to predict, for example, the features of the tumor immune microenvironment through imaging, and to monitor the therapeutic response beyond the standard radiological criteria. The aims of this narrative review are to provide a simplified guide for clinicians to these concepts, and to summarize the existing evidence on radiomics and "radi…-omics" in cancer immunotherapy.

The ability of cancer immunotherapy to generate lasting responses in a broad spectrum of tumors has generated great enthusiasm in medical oncology. A number of new immune-based compounds have now been approved based on the recent success of immune checkpoint blockade, either administered as monotherapy or in combination with other agents. Because clinical activity is limited only to subsets of patients, two major goals of cancer immunotherapy are (1) to reliably identify responders to these current treatments, and (2) to increase the number of patients who can respond to immunotherapy by developing new strategies. These goals are critically important since the hallmark of immune-based therapies is the induction of durable immunologic and clinical responses that result in overall survival benefit. Innovative combination strategies have great potential for bringing the benefit of immunotherapy to more patients. The use of cancer vaccines to actively induce immune effectors together with other drugs, which may include immune checkpoint blockade, chemotherapy, and/or molecularly targeted agents, is a particularly attractive strategy. Cancer vaccines have been tested both to prevent or intercept the development of cancer, and to decrease established tumor burdens. No vaccine has yet been approved for either breast cancer treatment or prevention. Here, we review the history of breast cancer vaccine development, and highlight near-term opportunities for moving forward.

Targeting inhibitory immune checkpoint molecules has dramatically changed treatment paradigms in medical oncology. Understanding the best strategies to unleash a pre-existing immune response or to induce an efficient immune response against tumors has emerged as a research priority. In this work, we focus on a novel target for cancer immunotherapy, the inhibitory receptor T-cell immunoglobulin and mucin domain 3 (TIM3). This narrative review describes TIM3 biology in different (tumor-infiltrating) immune cells, particularly in the immunosuppressive regulatory T cells and dysfunctional/exhausted cytotoxic T lymphocytes, but also in cells that confer innate immunity – natural killer and dendritic cells. We discuss the functional role of TIM3, its expression and its clinical significance in a variety of tumors, and confront the heterogeneous results emerging from different studies, including clinical trials of immunotherapy. Finally, this work summarizes the principal early-phase clinical trials exploring TIM3 blockade and discusses some future perspectives.

Cancer immunotherapy is demonstrating impressive clinical benefit in different malignancies and clinical oncologists are increasingly turning their attention to immune-oncology. It is now well recognized that innate and adaptive immune cells infiltrating tumors are associated with clinical outcomes and responses to treatments, and can be harnessed to patients' benefit. Considerable advances have also been made in understanding how cancers escape from immune attack. Targeting of immunological escape processes regulated by the expression of immune checkpoint receptors and ligands and the down-modulation of tumor antigen presentation is the basis of immuno-oncology treatments. Despite recent achievements, there remain a number of unresolved issues in order to successfully implement cancer immunotherapy in many cancers. Importantly, clinical biomarkers are still needed for better optimization of emerging combination immunotherapies and better treatment tailoring. In this review, we summarize the function of innate and adaptive immune cells in anti-tumor immunity and the general mechanisms exploited by tumor cells to escape and inhibit immune responses as well as therapeutic strategies developed to overcome these mechanisms and discuss emerging biomarkers in immuno-oncology.

This study investigated the prevalence of TIL subpopulations, TLS, PD-1 and PD-L1 in tumors from TNBC patients harboring wild-type or mutated BRCA1 or BRCA2 germline genes. This TNBC cohort included 85% TIL-positive (≥10%) tumors with 21% classified as TILhi (≥50%). Interestingly, the BRCAmut group had a significantly higher incidence of TILpos tumors compared to the BRCAwt group (P = 0.037). T cells were dominant in the infiltrate but no statistically significant differences were detected between BRCAwt and BRCAmut for CD3+, CD4+ and CD8+ T cells or CD20+ B cells. TLS were detected in 74% of tumors but again no significant differences between the BRCA groups. PD-1 expression was observed in 33% and PD-L1 in 53% (any cell, cut-off ≥1%) tumors for the entire TNBC cohort. PD-1 expression correlated with PD-L1 and both with TIL and TLS but was not associated with BRCA mutational status. Our analyses reveal that BRCAwt and BRCAmut TNBC are similar except for a significant increase of TILpos tumors in the BRCAmut group. While BRCA gene mutations may not directly drive immune infiltration, the greater number of TILpos tumors could signal greater immunogenicity in this group.

Tertiary lymphoid structures (TLS) have been detected in several types of human solid tumors. These structures are thought to regulate local adaptive immune responses that can promote or antagonize tumor progression. Despite positive prognostic values associated with a TLS presence in several studies, discrepancies still exist. TLS are structurally organized entities composed of varying numbers of multiple cell types making their assessment in tumor tissues, particularly biopsies, challenging. Immunohistochemical staining of TLS-related cell populations is the most frequently used method for identifying and scoring them; however, TLS-related gene expression has also been explored. The protocols described are detailed to allow the user to quantify TLS-related gene expression on formalin-fixed paraffin-embedded human breast tumor tissues.

There is an exponentially growing interest in targeting immune checkpoint molecules in breast cancer (BC), particularly in the triple-negative subtype where unmet treatment needs remain. This study was designed to analyze the expression, localization, and prognostic role of PD-1, PD-L1, PD-L2, CTLA-4, LAG3, and TIM3 in primary BC. Gene expression analysis using the METABRIC microarray dataset found that all six immune checkpoint molecules are highly expressed in basal-like and HER2-enriched compared to the other BC molecular subtypes. Flow cytometric analysis of fresh tissue homogenates from untreated primary tumors show that PD-1 is principally expressed on CD4+ or CD8+ T cells and CTLA-4 is expressed on CD4+ T cells. The global proportion of PD-L1+, PD-L2+, LAG3+, and TIM3+ tumor-infiltrating lymphocytes (TIL) was low and detectable in only a small number of tumors. Immunohistochemically staining fixed tissues from the same tumors was employed to score TIL and tertiary lymphoid structures (TLS). PD-L1+, PD-L2+, LAG3+, and TIM3+ cells were detected in some TLS in a pattern that resembles secondary lymphoid organs. This observation suggests that TLS are important sites of immune activation and regulation, particularly in tumors with extensive baseline immune infiltration. Significantly improved overall survival was correlated with PD-1 expression in the HER2-enriched and PD-L1 or CTLA-4 expression in basal-like BC. PD-1 and CTLA-4 proteins were most frequently detected on TIL, which supports the correlations observed between their gene expression and improved long-term outcome in basal-like and HER2-enriched BC. PD-L1 expression by tumor or immune cells is uncommon in BC. Overall, the data presented here distinguish PD-1 as a marker of T cell activity in both the T and B cell areas of BC associated TLS. We found that immune checkpoint molecule expression parallels the extent of TIL and TLS, although there is a noteworthy amount of heterogeneity between tumors even within the same molecular subtype. These data indicate that assessing the levels of immune checkpoint molecule expression in an individual patient has important implications for the success of therapeutically targeting them in BC., SCOPUS: ar.j, info:eu-repo/semantics/published

Assessment of tumor-infiltrating lymphocytes (TILs) in histopathologic specimens can provide important prognostic information in diverse solid tumor types, and may also be of value in predicting response to treatments. However, implementation as a routine clinical biomarker has not yet been achieved. As successful use of immune checkpoint inhibitors and other forms of immunotherapy become a clinical reality, the need for widely applicable, accessible, and reliable immunooncology biomarkers is clear. In part 1 of this review we briefly discuss the host immune response to tumors and different approaches to TIL assessment. We propose a standardized methodology to assess TILs in solid tumors on hematoxylin and eosin sections, in both primary and metastatic settings, based on the International Immuno-Oncology Biomarker Working Group guidelines for TIL assessment in invasive breast carcinoma. A review of the literature regarding the value of TIL assessment in different solid tumor types follows in part 2. The method we propose is reproducible, affordable, easily applied, and has demonstrated prognostic and predictive significance in invasive breast carcinoma. This standardized methodology may be used as a reference against which other methods are compared, and should be evaluated for clinical validity and utility. Standardization of TIL assessment will help to improve consistency and reproducibility in this field, enrich both the quality and quantity of comparable evidence, and help to thoroughly evaluate the utility of TILs assessment in this era of immunotherapy.

Assessment of the immune response to tumors is growing in importance as the prognostic implications of this response are increasingly recognized, and as immunotherapies are evaluated and implemented in different tumor types. However, many different approaches can be used to assess and describe the immune response, which limits efforts at implementation as a routine clinical biomarker. In part 1 of this review, we have proposed a standardized methodology to assess tumor infiltrating lymphocytes (TILs) in solid tumors, based on the International Immuno-Oncology Biomarkers Working Group guidelines for invasive breast carcinoma. In part 2 of this review, we discuss the available evidence for the prognostic and predictive value of TILs in common solid tumors, including carcinomas of the lung, gastrointestinal tract, genitourinary system, gynecological system, and head and neck, as well as primary brain tumors, mesothelioma and melanoma. The particularities and different emphases in TIL assessment in different tumor types are discussed. The standardized methodology we propose can be adapted to different tumor types and may be used as a standard against which other approaches can be compared. Standardization of TIL assessment will help clinicians, researchers and pathologists to conclusively evaluate the utility of this simple biomarker in the current era of immunotherapy.

Manipulating an individual's immune system through immune checkpoint blockade is revolutionizing the paradigms of cancer treatment. Peculiar patterns and kinetics of response have been observed with these new drugs, rendering the assessment of tumor burden particularly challenging in cancer immunotherapy. The mechanisms of action for immune checkpoint blockade, based upon engagement of the adaptive immune system, can generate unusual response patterns, including pseudoprogression, hyperprogression, atypical and delayed responses. In patients treated with immune checkpoint blockade and radiotherapy, a reduction in tumor burden at metastatic sites distant from the irradiation field (abscopal effect) has been observed, with synergistic systemic immune effects provoked by this combination. New toxicities have also been observed, due to excessive immune activity in several organs, including lung, colon, liver and endocrine glands. Efforts to standardize assessment of cancer immunotherapy responses include novel consensus guidelines derived by modifying World Health Organization (WHO) and Response Evaluation Criteria In Solid Tumors (RECIST) criteria. The aim of this review is to evaluate imaging techniques currently used routinely in the clinic and those being used as investigational tools in immunotherapy clinical trials.

The recent success of the immune checkpoint blockade in cancer immunotherapy has modified the treatment algorithms in a variety of aggressive neoplastic diseases. Nevertheless, optimal selection of ideal candidates to these drugs remains a challenge. The presence, location and composition of a pre-existing tumor immune infiltrate seem to impact on the benefit from these treatments. The association between the presence of baseline tumor-infiltrating lymphocytes (TIL) and patients' outcomes has been widely investigated in breast cancer, although immunotherapeutic strategies have historically been less successful with respect to other neoplastic diseases such as melanoma and kidney cancer. TIL extent varies and has different associations with outcomes in the various breast cancer subtypes. Furthermore, the presence of baseline high TIL has been associated with an increased benefit from some chemotherapeutic and targeted agents even though some conflicting results have been observed on this regard. This review aims to summarize the state of the art of TIL in breast cancer with a focus on their assessment, prevalence and clinical implications in the different subtypes.

Breast cancers arising in young women are biologically more aggressive, and most of these patients are candidates to receive aggressive treatments that include the use of chemotherapy. As most of these tumors express the hormone receptors (i.e., luminal disease), these patients are also candidates to adjuvant endocrine therapy. Chemotherapy-induced amenorrhea showed to be prognostic in young patients with luminal breast cancer. However, the role of ovarian function suppression (OFS) in addition to standard adjuvant treatments has been largely debated over the past years. Recently, several studies have provided important insights on the role of OFS. Currently, the use of tamoxifen alone without prior cytotoxic therapy can be considered a very effective treatment option in young patients with hormone receptor-positive breast cancer at low risk of relapse. On the other hand, for patients at higher risk of relapse as those who are candidates to (neo)adjuvant chemotherapy, OFS proved to be beneficial, and therefore luteinizing hormone-releasing hormone agonists (LHRHa) should be considered in addition to tamoxifen or aromatase inhibitors (AI). However, toxicity is considerable and patients should be actively engaged in decision-making. Finally, in young breast cancer patients who are candidates to (neo)adjuvant chemotherapy, loss of ovarian function and fertility may be a concern. Besides other techniques, recent results showed that temporary OFS with LHRHa during cytotoxic treatment can be considered a reliable strategy to preserve gonadal function and fertility. Despite the recent advances in the field, several gray zones remain unanswered: the role of OFS plus AI in women who remained premenopausal after 5 years of tamoxifen, the optimal extended approach in women treated with 5 years of OFS plus AI, and the role of temporary OFS with LHRHa during chemotherapy in the specific subgroup of patients with BRCA mutations and in women undergoing this strategy after prior embryo/oocyte cryopreservation.

In genito-urinary tumors immunotherapy has been administered for a long time: Calmette-Guerin Bacillus as adjuvant treatment in high risk patients with non muscle invasive urothelial bladder cancer and interleukin-2 and interferon-α in metastatic kidney cancer. The vaccine Sipuleucel-T has been approved by United States Food and Drug Administration for the treatment of castration resistant prostate cancer patients with asymptomatic or minimally symptomatic disease, given the 22% reduction of mortality risk in this group. Recently immunotherapeutic agents targeting inhibitory immune checkpoint molecules lead to improved outcomes and lasting anti-tumor effects in a variety of hematological and solid malignancies, including urogenital tumors. The benefit from these treatments has been observed only in a proportion of subjects, raising a need in optimizing patients' selection for immune checkpoint blockade. The composition and activity of a pre-existing immune infiltrate may aid in identifying ideal candidates to immunotherapy, with possible implications for the clinical management of neoplastic diseases from earlier to later stages.

The clinical relevance of tumor-infiltrating lymphocytes (TIL) in breast cancer (BC) has been clearly established by their demonstrated correlation with long-term positive outcomes. Nevertheless, the relationship between protective immunity, observed in some patients, and critical features of the infiltrate remains unresolved. This study examined TIL density, composition and organization together with PD-1 and PD-L1 expression in freshly collected and paraffin-embedded tissues from 125 patients with invasive primary BC. Tumor and normal breast tissues were analyzed using both flow cytometry and immunohistochemistry. TIL density distribution is a continuum with 25% of tumors identified as TIL-negative at a TIL density equivalent to normal breast tissues. TIL-positive tumors (75%) were equally divided into TIL-intermediate and TIL-high. Tumors had higher mean frequencies of CD4+ T cells and CD19+ B cells and a lower mean frequency of CD8+ T cells compare with normal tissues, increasing the CD4+/CD8+ T-cell ratio. Tertiary lymphoid structures (TLS), principally located in the peri-tumoral stroma, were detected in 60% of tumors and correlated with higher TIL infiltration. PD-1 and PD-L1 expression were also associated with higher TIL densities and TLS. TIL density, TLS and PD-L1 expression were correlated with more aggressive tumor characteristics, including higher proliferation and hormone receptor negativity. Our findings reveal an important relationship between PD-1/PD-L1 expression, increased CD4+ T and B-cell infiltration, TIL density and TLS, suggesting that evaluating not only the extent but also the nature and location of the immune infiltrate should be considered when evaluating antitumor immunity and the potential for benefit from immunotherapies.