Tumour inflammatory infiltrate predicts survival following curative resection for node-negative colorectal cancer

BackgroundA pronounced tumour inflammatory infiltrate is known to confer a good outcome in colorectal cancer. Klintrup and colleagues reported a structured assessment of the inflammatory reaction at the invasive margin scoring low grade or high grade. The aim of the present study was to examine the prognostic value of tumour inflammatory infiltrate in node-negative colorectal cancer.MethodsTwo hundred patients had undergone surgery for node-negative colorectal cancer between 1997 and 2004. Specimens were scored with Jass’ and Klintrup’s criteria for peritumoural infiltrate. Pathological data were taken from the reports at that time.ResultsLow-grade inflammatory infiltrate assessed using Klintrup’s criteria was an independent prognostic factor in node-negative disease. In patients with a low-risk Petersen Index (n = 179), low-grade infiltrate carried a threefold increased risk of cancer death. Low-grade infiltrate was related to increasing T stage and an infiltrating margin.ConclusionAssessment of inflammatory infiltrate using Klintrup’s criteria provides independent prognostic information on node-negative colorectal cancer. A high-grade local inflammatory response may represent effective host immune responses impeding tumour growth.     

The relationships between cellular components of the peritumoural inflammatory response, clinicopathological characteristics and survival in patients with primary operable colorectal cancer

Background:

The host inflammatory response is an important determinant of cancer outcome. We examined different methods of assessing the local inflammatory response in colorectal tumours and explored relationships with both clinicopathological characteristics and survival.

Methods:

Cohort study of patients (n=130) with primary operable colorectal cancer and mature follow-up. Local inflammatory response at the invasive margin was assessed with: (1) a semi-quantitative assessment of peritumoural inflammation using Klintrup–Makinen (K–M) grading and (2) an assessment of individual immune cell infiltration (lymphocytes, plasma cells, neutrophils, macrophages and eosinophils).

Results:

The peritumoural inflammatory response was K–M low grade in 48% and high grade in 52%. Inflammatory cells were primarily macrophages, lymphocytes and neutrophils with relatively few plasma cells or eosinophils. On univariate analysis, K–M grade, lymphocyte infiltration and plasma cell infiltration were associated with cancer-specific survival. On multivariate analysis, only systemic inflammatory response, TNM (tumour, node and metastases) stage, venous invasion, tumour necrosis and K–M grade were independently associated with cancer-specific survival. There was no relationship between local infiltration of inflammatory cells and a systemic inflammatory response. However, high K–M grade, lymphocyte infiltration and plasma cell infiltration were associated with a number of favourable pathological characteristics, including an absence of venous invasion.

Conclusion:

Infiltration of inflammatory cells in the invasive margin of colorectal tumours is beneficial to survival. The adaptive immune response appears to have a prominent role in the prevention of tumour progression in patients with colorectal cancer.     

The role of the in situ local inflammatory response in predicting recurrence and survival in patients with primary operable colorectal cancer

Colorectal cancer progression and survival is dependent on complex interactions between the tumour and the host. The pronounced local inflammatory response in and around the tumour is thought to represent the in situ host anti-tumour immune response. Since early reports, 40 years ago, there has been a continuing interest in establishing the cellular composition of immune cell infiltrates and their relationship with survival in colorectal cancer. In this review, we comprehensively examine the evidence for the local inflammatory cell reaction/in situ immune response in predicting outcome in primary operable colorectal cancer and make recommendations as to how such information may be incorporated into routine clinical assessment. Generally, an increasing number/density of immune cells in and around the tumour is associated with improved outcome in over 100 studies. Whilst the prognostic value of a generalized lymphocytic infiltrate or non-specific peritumoural inflammatory response is strongly related to survival based on 40 different studies, it is also apparent that most individual immune cell types relate to recurrence and cancer specific survival. The evidence is particularly robust for tumour infiltrating T lymphocytes and their subsets (CD3+, CD8+, CD45RO+, FOXP3+) in addition to tumour associated macrophages, dendritic cells and neutrophils. Taken together, the evidence suggests both adaptive and innate anti-tumour immune responses play key roles in determining cancer progression. In order to establish routine clinical utility there is a need to rationalise this prognostic information, published over a 40 years period, into a standardized assessment of tumour inflammatory cell infiltrate. Such standardization may also guide development of novel therapeutic interventions.     

The Role of Immune Cells in Breast Tissue and Immunotherapy for the Treatment of Breast Cancer

Immune cells are present in normal breast tissue and in breast carcinoma. The nature and distribution of the immune cell subtypes in these tissues are reviewed to promote a better understanding of their important role in breast cancer prevention and treatment. We conducted a review of the literature to define the type, location, distribution, and role of immune cells in normal breast tissue and in in situ and invasive breast cancer. Immune cells in normal breast tissue are located predominantly within the epithelial component in breast ductal lobules. Immune cell subtypes representing innate immunity (NK, CD68⁺, and CD11c⁺ cells) and adaptive immunity (most commonly CD8⁺, but CD4⁺ and CD20⁺ as well) are present; CD8⁺ cells are the most common subtype and are primarily effector memory cells. Immune cells may recognize neoantigens and endogenous and exogenous ligands and may serve in chronic inflammation and immunosurveillance. Progression to breast cancer is characterized by increased immune cell infiltrates in tumor parenchyma and stroma, including CD4⁺ and CD8⁺ granzyme B⁺ cytotoxic T cells, B cells, macrophages and dendritic cells. Tumor-infiltrating lymphocytes in breast cancer may serve as prognostic indicators for response to chemotherapy and for survival. Experimental strategies of adoptive transfer of breast tumor-infiltrating lymphocyte may allow regression of metastatic breast cancer and encourage development of innovative T-cell strategies for the immunotherapy of breast cancer. In conclusion, immune cells in breast tissues play an important role throughout breast carcinogenesis. An understanding of these roles has important implications for the prevention and the treatment of breast cancer.     

The local inflammatory response in colorectal cancer – Type,location or density? A systematic review and meta-analysis

IntroductionThe host anti-tumour inflammatory response is a strong prognostic indicator, and tumour infiltrating lymphocytes (TILs) are believed to have a complimentary role alongside TNM assessment in dictating future management. However, there is wide disagreement regarding the most efficacious and cost-effective method of assessment.MethodsA comprehensive literature search was performed of EMBASE, MedLine and PubMed as well as an assessment of references to identify all relevant studies relating to the assessment of the peri-tumoural inflammatory response or TILs and prognosis in colorectal cancer (CRC). A meta-analysis was performed of 67 studies meeting the REMARK criteria using RevMan software.ResultsIntratumoural assessment of both CD3 and CD8 in CRC were significant for disease-free survival (DFS) (combined HRs 0.46; 95%CI: 0.39–0.54 and 0.54; 95%CI: 0.45–0.65), as well as overall survival (OS) and disease-specific survival (DSS). The same was true for assessment of CD3 and CD8 at the invasive margin (DFS: combined HRs 0.45; 95%CI: 0.33–0.61 and 0.51; 95%CI: 0.41–0.62). However, similar fixed effects summaries were also observed for H&E-based methods, like Klintrup-Makinen grade (DFS: HR 0.62; 95%CI: 0.43–0.88). Furthermore, inflammatory assessments were independent of MSI status.ConclusionThe evidence suggests that it is the density of a co-ordinated local inflammatory infiltrate that confers survival benefit, rather than any individual immune cell subtype. Furthermore, the location of individual cells within the tumour microenvironment does not appear to influence survival. The authors advocate a standardised assessment of the local inflammatory response, but caution against emphasizing the importance of any individual immune cell subtype.     

Comparison of visual and automated assessment of tumour inflammatory infiltrates in patients with colorectal cancer

BackgroundCancer-associated inflammation is increasingly recognised to be an important determinant of oncological outcome. In colorectal cancer, the presence of peri-tumoural inflammatory/lymphocytic infiltrates predicts improved survival. To date, these infiltrates, assessed visually on haematoxylin and eosin (H&E) stained sections, have failed to enter routine clinical practice, partly due to their subjective assessment and considerable inter-observer variation. The present study aims to develop an automated scoring method to enable consistent and reproducible assessment of tumour inflammatory infiltrates in colorectal cancer.Methods154 colorectal cancer patients who underwent curative resection were included in the study. The local inflammatory infiltrate was assessed using the method described by Klintrup–Makinen. H&E tumour sections were uploaded to an image analysis programme (Slidepath, Leica Biosystems). An image analysis algorithm was developed to count the inflammatory cells at the invasive margin. The manual and automated assessments of the tumour inflammatory infiltrates were then compared.ResultsThe automated inflammatory cell counts assessed using the freehand annotation method (p < 0.001) and the rectangular box method (p < 0.001) were significantly associated with both K–M score (p < 0.001) and K–M grade (p < 0.001). The inflammatory cell counts were divided using quartiles to group tumours with similar inflammatory cell densities. There was good agreement between the manual and automated scoring methods (intraclass correlation coefficient (ICC) = 0.82). Similar to the visual K–M scoring system, the automated K–M classification of the inflammatory cell counts, using quartiles, was significantly associated with venous invasion (p < 0.05) and modified Glasgow Prognostic Score (mGPS) (p ⩽ 0.05). On univariate survival analysis, both automated K–M category (p < 0.05) and automated K–M grade (p < 0.005) were associated with cancer-specific survival.ConclusionThe results of the present study demonstrate that automated assessment effectively recapitulates the clinical value of visual assessment of the local inflammatory cell infiltrate at the invasive margin of colorectal tumours. In addition, it is possible to obtain an objective assessment of tumour inflammatory infiltrates using routinely stained H&E sections. An automated, computer-based scoring method is therefore a workable and cost-effective approach to clinical assessment of local immune cell infiltrates in colorectal cancer.     

CD8+ T cells inhibit metastasis and CXCL4 regulates its function

Background The mechanism by which immune cells regulate metastasis is unclear. Understanding the role of immune cells in metastasis will guide the development of treatments improving patient survival.Methods We used syngeneic orthotopic mouse tumour models (wild-type, NOD/scid and Nude), employed knockout (CD8 and CD4) models and administered CXCL4. Tumours and lungs were analysed for cancer cells by bioluminescence, and circulating tumour cells were isolated from blood. Immunohistochemistry on the mouse tumours was performed to confirm cell type, and on a tissue microarray with 180 TNBCs for human relevance. TCGA data from over 10,000 patients were analysed as well.Results We reveal that intratumoral immune infiltration differs between metastatic and non-metastatic tumours. The non-metastatic tumours harbour high levels of CD8⁺ T cells and low levels of platelets, which is reverse in metastatic tumours. During tumour progression, platelets and CXCL4 induce differentiation of monocytes into myeloid-derived suppressor cells (MDSCs), which inhibit CD8⁺ T-cell function. TCGA pan-cancer data confirmed that CD8^lowPlatelet^high patients have a significantly lower survival probability compared to CD8^highPlatelet^low.Conclusions CD8⁺ T cells inhibit metastasis. When the balance between CD8⁺ T cells and platelets is disrupted, platelets produce CXCL4, which induces MDSCs thereby inhibiting the CD8⁺ T-cell function.Subject terms: Tumour immunology, Metastasis     

The in situ local immune response,tumour senescence and proliferation in colorectal cancer

Background:

Immune cell infiltrates are important determinants of colorectal cancer (CRC) outcome. Their presence may be driven by tumour or host-specific factors. From previous studies in mice, senescence, a state of cell cycle arrest, may moderate tumour progression through upregulation of antitumour immune responses. The relationships between senescence and immune infiltrates have not previously been studied in humans. We explore whether a marker of senescence (p16^ink4a) in combination with low level expression of a proliferation marker (ki-67) relate to T cell infiltrates in CRC, and whether p16^ink4a, Ki-67 and immune infiltrates have similar prognostic value.

Methods:

Immunostaining of p16^inka and Ki-67 was performed within a CRC tissue microarray. Nuclear p16^inka and Ki-67 were categorised as high/low. T-cell markers, CD3, CD45RO, CD8 and FOXP3 were scored separately as high/low grade in three areas of the tumour: the invasive margin (IM), tumour stroma and cancer cell nests (CCNs).

Results:

Two hundred and thirty stage I–III cancers were studied. High nuclear p16^ink4a was expressed in 63% and high proliferation (Ki-67 >15%) in 61%. p16^ink4a expression was associated with reduced CD45RO+ cells at the IM (P<0.05) and within the stroma (P<0.05) and reduced CD8+ cells at the IM (P<0.01). A low Ki-67 proliferative index was associated with reduced density of CD3+ cells in CCNs (P<0.01), reduced CD45RO+ cells at the IM (P<0.05) and within the CCNs (P<0.001), reduced FOXP3+ cells at the IM (P<0.001), within the stroma (P=0.001) and within CCNs (P<0.001) and reduced CD8+ cells at the IM (P<0.05) and within the CCNs (P<0.05). Tumours with both a low proliferative index and expression of p16^ink4a demonstrated similar consistent relationships with reduced densities of T-cell infiltrates. On multivariate analysis, TNM stage (P<0.001), low CD3 cells at the IM (P=0.014), low CD8 cells at the IM (P=0.037), low proliferation (Ki-67; P=0.013) and low senescence (p16^ink4a; P=0.002) were independently associated with poorer cancer survival.

Conclusion:

Senescence, proliferation and immune cell infiltrates are independent prognostic factors in CRC. Although related to survival, p16^ink4a-associated senescence is not associated with an upregulation of antitumour T-cell responses.     

Body mass index and risk of colorectal cancer according to tumor lymphocytic infiltrate

Higher body mass index (BMI), higher body adiposity and obesity have been associated with increased risk of colorectal cancer. Evidence suggests that excess energy balance may influence systemic immune and inflammatory status. Thus, we hypothesized that the positive association between BMI and colorectal cancer risk might differ according to colorectal carcinoma subtypes according to levels of histopathological lymphocytic reaction to tumor. We collected biennial questionnaire data on weight and baseline height information in two prospective cohort studies, the Nurses' Health Study (1980‐2010) and the Health Professionals Follow‐up Study (1986‐2010). Utilizing duplication‐method Cox proportional hazards regression models, we prospectively assessed the association between BMI and risk of colorectal cancer subtypes according to the degree of Crohn's‐like lymphoid reaction, peritumoral lymphocytic reaction, intratumoral periglandular reaction, tumor‐infiltrating lymphocytes, the overall lymphocytic reaction score, or T‐cell [CD3⁺, CD8⁺, CD45RO (PTPRC)⁺ or FOXP3⁺] density in tumor tissue. Statistical significance level was adjusted for multiple hypotheses testing by Bonferroni correction. During follow up of 1,708,029 men and women (over 3,346,752 person‐years), we documented 1,436 incident rectal and colon cancer cases with available formalin‐fixed paraffin‐embedded tumor tissue materials and pathological immunity data. BMI was significantly associated with higher risk of overall colorectal cancer (P_trend < 0.001); however, the association of BMI with colorectal carcinoma risk did not significantly differ by the level of lymphocytic reaction or T‐cell infiltration in tumor tissue status (P_{heterogeneity} > 0.10). BMI may be associated with risk of colorectal cancer regardless of levels of lymphocytic response to tumor.     

TiaochangXiaoliu decoction inhibits azomethane (AOM)/dextran sulfate sodium (DSS)-induced colorectal cancer by regulating immune response

BackgroundTiaochangXiaoliu decoction (TXD) has an anti-tumor effect in clinical practice. We further investigated the role of TXD in colorectal cancer (CRC).MethodsMouse models of CRC were induced by azomethane (AOM)/dextran sulfate sodium (DSS), with sixty male C57BL/6 mice randomly divided into six groups (10 mice/group): a control group, AOM/DSS group, TXD at low dose (L-dose) group, middle dose (M-dose) group, high dose (H-dose) group, and Celecoxib (Cel) group. The colorectum, serum, and plasma of mice in each group was collected following sacrifice to record the number of tumors. HE staining was utilized for observing pathological damage to colorectal tissues, ELISA used for detecting INF-γ, IL-2, and TNF-α expression in serum, and flow cytometry used for measuring the proportion of CD4⁺, CD8⁺, CD4⁺/CD8⁺, and NK cells in plasma.ResultsCompared with the control group, the AOM/DSS group showed tumor masses in the colorectum and different degrees of pathological damage in the intestine. AOM/DSS induction also resulted in an increase in INF-γ, IL-2, and TNF-α expression in serum, and a decrease in the percentages of CD4⁺, CD8⁺, CD4⁺/CD8⁺, and NK cells(P<0.05). In comparison with the AOM/DSS group, with the increase of TXD dose, the number of tumors decreased significantly, and intestinal structure and mucosal inflammatory cell infiltration also improved. Further, in comparison with the AOM/DSS group, all three doses of TXD and celecoxib caused an increase in the contents of CD4⁺, CD8⁺, CD4⁺/CD8⁺, and NK cells in plasma. In addition, in the M-dose, H-dose, and Cel groups, INF-γ, IL-2, and TNF-α expression showed a marked decrease, and the reduction in these two groups treated with TXD was dose-dependent.ConclusionsTXD leads to a marked reduction in the number of tumors and inflammatory cell infiltration in CRC mice. This decoction significantly decreased the levels of INF-γ, IL-2, and TNF-α in serum, and increased the contents of CD4⁺, CD8⁺, CD4⁺/CD8⁺, and NK cell in the plasma of mice with AOM/DSS-induced CRC.     

Colibactin-positive Escherichia coli induce a procarcinogenic immune environment leading to immunotherapy resistance in colorectal cancer

Colibactin-producing E. coli (CoPEC) are frequently detected in colorectal cancer (CRC) and exhibit procarcinogenic properties. Because increasing evidence show the role of immune environment and especially of antitumor T-cells in CRC development, we investigated the impact of CoPEC on these cells in human CRC and in the APC^Min/+ mice colon. T-cell density was evaluated by immunohistochemistry in human tumors known for their CoPEC status. APC^min/+ mice were chronically infected with a CoPEC strain (11G5). Immune cells (neutrophils and T-cell populations) were then quantified by immunofluorescent staining of the colon. The quantification of lymphoid populations was also performed in the mesenteric lymph nodes (MLNs). Here, we show that the colonization of CRC patients by CoPEC is associated with a decrease of tumor-infiltrating T lymphocytes (CD3⁺ T-cells). Similarly, we demonstrated, in mice, that CoPEC chronic infection decreases CD3⁺ and CD8⁺ T-cells and increases colonic inflammation. In addition, we noticed a significant decrease in antitumor T-cells in the MLNs of CoPEC-infected mice compared to that of controls. Moreover, we show that CoPEC infection decreases the antimouse PD-1 immunotherapy efficacy in MC38 tumor model. Our findings suggest that CoPEC could promote a procarcinogenic immune environment through impairment of antitumor T-cell response, leading to tumoral resistance to immunotherapy. CoPEC could thus be a new biomarker predicting the anti-PD-1 response in CRC.     

Presence of circulating Her2-reactive CD8?+?T-cells is associated with lower frequencies of myeloid-derived suppressor cells and regulatory T cells,and better survival in older breast cancer patients

Introduction

Breast cancer is one of the most common cancers among women. Its incidence is increasing in many countries and a higher number of older women are now being diagnosed with the disease. Immune parameters are implicated in disease progression, and the frequencies of both myeloid-derived suppressor cells (MDSCs) and regulatory T cells (Tregs), associated with tumour burden, have been suggested to be indicators of poor prognosis in cases of metastatic breast cancer.

Methods

Here, we have assessed the frequency of peripheral Tregs and MDSCs in relation to in vitro T cell responses to Her2 antigen in 40 untreated breast cancer patients 65 to 87 years of age at diagnosis.

Results

The five-year survival rate of patients who mounted a CD8+ T cell response to Her2 peptides and had a lower frequency of Lin⁻CD14⁺HLA-DR⁻MDSCs was 100% compared to only 38% in patients without Her2-reactive CD8+ T cells and with higher frequencies of MDSCs (P = 0.03). Patients who lacked a CD8 response to Her2 tended to have higher frequencies of MDSCs. Similarly, patients who lacked a CD8 response to Her2 and had higher frequencies of CD4⁺Foxp3⁺CD127^lowCD25⁺ Tregs had only 50% survival compared to the 100% survival of patients who did mount a CD8 response and had lower frequencies of Tregs (P = 0.03). A similar trend was observed for activated (CD4⁺CD45RA⁻Foxp3^hi) but not resting Tregs (CD4⁺CD45RA⁺FoxP3⁺). This survival advantage was observed in both metastatic and non-metastatic patients.

Conclusions

Our data demonstrate a negative role of both MDSCs and Tregs in the prognosis of breast cancer patients, the mechanism of which might be through dampening favourable CD8+ T cell immune responses to tumour-associated antigens.

Electronic supplementary material

The online version of this article (doi:10.1186/s13058-015-0541-z) contains supplementary material, which is available to authorized users.     

Modulation of intratumoural myeloid cells,the hallmark of the anti-tumour efficacy induced by a triple combination: tumour-associated peptide,TLR-3 ligand and α-PD-1

Background Anti-programmed cell death 1 (PD-1)/programmed death-ligand 1 (PD-L1) monoclonal antibodies (mAbs) show remarkable clinical anti-tumour efficacy. However, rational combinations are needed to extend the clinical benefit to primary resistant tumours. The design of such combinations requires the identification of the kinetics of critical immune cell populations in the tumour microenvironment.Methods In this study, we compared the kinetics of immune cells in the tumour microenvironment upon treatment with immunotherapy combinations with different anti-tumour efficacies in the non-inflamed tumour model TC-1/A9. Tumour-bearing C57BL/6J mice were treated with all possible combinations of a human papillomavirus (HPV) E7 long peptide, polyinosinic–polycytidylic acid (PIC) and anti-PD-1 mAb. Tumour growth and kinetics of the relevant immune cell populations were assessed over time. The involvement of key immune cells was confirmed by depletion with mAbs and immunophenotyping with multiparametric flow cytometry.Results The maximum anti-tumour efficacy was achieved after intratumoural administration of HPV E7 long peptide and PIC combined with the systemic administration of anti-PD-1 mAb. The intratumoural immune cell kinetics of this combination was characterised by a biphasic immune response. An initial upsurge of proinflammatory myeloid cells led to a further rise in effector CD8⁺ T lymphocytes at day 8. Depletion of either myeloid cells or CD8⁺ T lymphocytes diminished the anti-tumour efficacy of the combination.Conclusions The anti-tumour efficacy of a successful immunotherapy combination in a non-inflamed tumour model relies on an early inflammatory process that remodels the myeloid cell compartment.Subject terms: Oncology, Cancer     

The relationships between serum cytokine levels and tumor infiltrating immune cells and their clinical significance in colorectal cancer

Increased inflammatory cell infiltration correlates to improved survival in colorectal cancer (CRC). Development and progression of CRC is associated with alterations in serum cytokine levels but their significance is not well defined. In this study, we investigated the relationships between the serum levels of 13 cytokines and the densities of eight types of tumor infiltrating inflammatory cells and their impact on disease‐free survival (DFS), cancer‐specific survival (CSS) and overall survival (OS) in a prospectively recruited group of 147 CRC patients. There were strong positive correlations between the serum concentrations of different cytokines, as well as between the different types of tumor infiltrating immune cells, whereas the associations between serum cytokines and tumor infiltrating immune cells were generally weak. High serum IL‐12 levels associated with increased densities of peritumoral CD8⁺ T cells, intraepithelial CD3⁺ T cells and intratumoral neutrophils, while high serum CCL4 levels associated with increased densities of peritumoral CD68⁺ cells. In multivariate survival models, increased infiltration of intraepithelial CD3⁺ T cells and increased serum CCL4 associated with improved DFS, whereas higher intratumoral CD83⁺ dendritic cell density and increased serum interferon gamma levels associated with improved CSS and OS. Also high density of peritumoral CD3⁺ T cells associated with improved CSS. In conclusion, serum cytokines and tumor infiltrating immune cells in CRC represent entities with high intragroup correlations but relatively weak intergroup correlations. The results suggest that tumor infiltrating CD3⁺ T cells, CD83⁺ dendritic cells, serum CCL4 and serum interferon gamma represent relevant markers of disease outcome.     

Determining circulating endothelial cells using CellSearch system during preoperative systemic chemotherapy in breast cancer patients

Background

Circulating endothelial cells (CECs) have been studied as a biomarker for tumour progression and monitoring therapeutic effects. The CellSearch system is a semi-automated system that allows standardised analysis of CECs. This study assessed the clinical implications of CECs determined by the CellSearch system in breast cancer patients.

Methods

Seventy-six consecutive breast cancer patients (53 operable and 23 metastatic or recurrent) were enrolled for the study. Thirty-five patients with operable breast cancer received preoperative chemotherapy with a regimen based on anthracycline and/or taxane. CECs are defined as CD146⁺CD105⁺CD45⁻DAPI⁺ cells in the system. CD34 expression was examined using the additional channel in the system.

Results

A majority (4539 of 5183 cells, 88%) of CECs from patients with operable breast cancer were CD34-positive. Triple-negative cancers showed higher baseline CEC and CD34⁺CEC counts than the other types (P = 0.0387 and 0.0377, respectively). Low baseline CEC and CD34⁺CEC counts, and a low CD34 positive rate were associated with pathological complete response (pCR) of preoperative chemotherapy in patients with primary breast cancer (P = 0.046, 0.027 and 0.01, respectively). In multivariate analyses, the CD34 positive rate was significant for pCR (P = 0.021). During preoperative chemotherapy, CEC and CD34⁺CEC counts before each cycle of chemotherapy increased with taxane-based regimens (P = 0.0018 and 0.0008, respectively) but not with anthracycline-based regimens.

Conclusions

Baseline CEC, in particular CD34⁺CEC, counts and the CD34 positive rate might be useful for the prediction of treatment response of preoperative chemotherapy in patients with operable breast cancer.     

The proportion of circulating CD45RO+CD8+ memory T cells is correlated with clinical response in melanoma patients treated with ipilimumab

BackgroundImmune checkpoint blockade (ICB) has been a breakthrough in the treatment of metastatic melanoma. But with only about 20–40% long-term responders and severe side-effects in about 12–17%, finding predictive markers for treatment response is of great interest.MethodsWe prospectively assessed clinical data, haematologic parameters and freshly isolated peripheral blood mononuclear cells of 30 patients treated with ipilimumab (n = 21) and pembrolizumab (n = 9) prior to the first 4 cycles with ICB and before the first tumour assessment.ResultsWe discovered that the baseline levels of CD45RO⁺CD8⁺ T cells significantly differed among the patients. Thirteen (43%) of our patients had normal baseline levels of CD45RO⁺CD8⁺ T cells, whereas 17 (57%) patients were low on CD45RO⁺CD8⁺ T cells. The baseline levels of CD45RO⁺CD8⁺ T cells correlated significantly with the response to ipilimumab but not pembrolizumab. Patients with baseline levels of lower/equal 25% of CD45RO⁺CD8⁺ T cells did not respond to treatment with ipilimumab. Phenotyping the CD8⁺ T cells in patients treated with ipilimumab revealed an activated HLA-DR⁺CD25⁻ phenotype, implying antigen non-specific stimulation. The levels of the HLA-DR⁺CD25⁻CD8⁺ T cells were significantly higher in patients with a normal baseline of CD45RO⁺CD8⁺ T cells and even increased significantly during treatment. Furthermore, proliferation of melanoma antigen recognized by T cells 1 (MART-1)-specific CD8⁺ T cells was not observed. Patients with normal baseline levels of CD45RO⁺CD8⁺ T cells showed a significant longer overall survival when treated with ipilimumab but not pembrolizumab.ConclusionPatients with normal baseline levels of CD45RO⁺CD8⁺ T cells respond significantly more frequently to treatment with ipilimumab and the CD8⁺ T cells appear to be antigen non-specifically activated. The baseline level of CD45RO⁺CD8⁺ T cells represents a promising factor as biomarker for the prediction of the response to ipilimumab.     

Improved therapeutic efficacy of unmodified anti-tumor antibodies by immune checkpoint blockade and kinase targeted therapy in mouse models of melanoma

The use of specific anti-tumor antibodies has transformed the solid cancer therapeutics landscape with the relative successes of therapies such as anti-HER2 in breast cancer, and anti-EGFR in HNSCC and colorectal cancer. However, these therapies result in toxicity and the emergence of resistant tumors. Here, we showed that removing immune suppression and enhancing stimulatory signals increased the anti-tumor activity of unmodified TA99 antibodies (anti-TYRP1) with a significant reduction of growth of solid tumors and lung metastases in mouse models of melanoma. Immune checkpoint blockade enhanced the efficacy of TA99, which was associated with greater CD8⁺/Foxp3⁺, NK1.1⁺ and dendritic cell infiltrates, suggestive of an increased anti-tumor innate and adaptive immune responses. Further, MEK inhibition in melanoma cell lines increased the expression of melanosomal antigens in vitro, and combining TA99 and MEKi in vivo resulted in enhanced tumor control. Moreover, we found an improved therapeutic effect when YUMM tumor-bearing mice were treated with TA99 combined with MEKi and immune checkpoint blockade (anti-PD1 and anti-CTLA4). Our findings suggest that MEKi induced an increased expression of tumor-associated antigens, which in combination with anti-tumor antibodies, generated a robust adaptive anti-tumor response that was sustained by immune checkpoint inhibition therapy. We postulate that combining anti-tumor antibodies with standard-of-care strategies such as immune checkpoint blockade or targeted therapy, will improve therapeutic outcomes in cancer.     

Impact of Anti-HER2 Treatments Combined With Atezolizumab on the Tumor Immune Microenvironment in Early or Metastatic Breast Cancer: Results From a Phase Ib Study

BackgroundDespite advances, there continues to be unmet need in breast cancer. Combining anti–programmed death-ligand 1 (PD-L1) cancer immunotherapy atezolizumab with other targeted therapies may enhance T-cell–dependent cytolytic antitumor activity.MethodsThis open-label, phase Ib study evaluated the safety of atezolizumab-based combinations with antibody-dependent cellular cytotoxicity or antibody-drug conjugate (ADC) agents. Patients with unresectable human epidermal growth factor receptor 2–positive (HER2⁺) locally advanced or metastatic breast cancer (mBC) received atezolizumab with trastuzumab/pertuzumab, atezolizumab with the ADC ado-trastuzumab emtansine (T-DM1), or atezolizumab with trastuzumab/pertuzumab and docetaxel. In an early–breast cancer (eBC) “window of opportunity” study, patients with operable HER2⁺ locally advanced or inflammatory eBC received neoadjuvant atezolizumab with trastuzumab/pertuzumab or atezolizumab/T-DM1, followed by docetaxel, carboplatin, and trastuzumab/pertuzumab. Exploratory outcomes included tumor response and biomarkers.ResultsBy March 15, 2019, 73 patients were enrolled. Safety findings were consistent with the treatment components’ individual profiles. Objective responses were observed in 2 of 6 and 5 of 14 patients in 2 mBC cohorts receiving atezolizumab/T-DM1 and in 6 of 6 patients with mBC receiving atezolizumab, trastuzumab/pertuzumab, and docetaxel. PD-L1 in immune cells was the only biomarker that increased with atezolizumab/T-DM1. In the window of opportunity cohorts, PD-L1 levels and CD8⁺ T-cell infiltration increased from baseline in HER2⁺ eBC tumors receiving atezolizumab with trastuzumab/pertuzumab or T-DM1, irrespective of response. Despite increases in T-cell and B-cell gene signatures with trastuzumab/pertuzumab, but not T-DM1, neither combination promoted T-cell receptor clonal expansion.ConclusionAtezolizumab with antibody-dependent cellular cytotoxicity or ADC agents appears safe and may activate the adaptive immune system of patients with HER2⁺ eBC tumors more than those with mBC tumors.     

Evaluation of Immune Reaction and PD-L1 Expression Using Multiplex Immunohistochemistry in HER2-Positive Breast Cancer: The Association With Response to Anti-HER2 Neoadjuvant Therapy

Background

Immune reaction with tumor-infiltrating lymphocytes (TILs) has been extensively investigated in breast cancer. Programmed cell death 1 and its ligand (PD-L1) are key physiologic suppressors of cytotoxic immune reaction. However, the combination of TILs with PD-L1 expression has not been well studied in breast cancer.