Probes for INS

Abstract

Background

Surrogate biomarkers of efficacy are needed for anti-PD1/PD-L1 therapy, given the existence of delayed responses and pseudo-progressions. We evaluated changes in serum IL-8 levels as a biomarker of response to anti-PD-1 blockade in melanoma and non-small cell lung cancer (NSCLC) patients.

Patients and methods

Metastatic melanoma and NSCLC patients treated with nivolumab or pembrolizumab alone or nivolumab plus ipilimumab, were studied. Serum was collected at baseline; at 2-4 weeks after the first dose; and at the time-points of response evaluation. Serum IL-8 levels were determined by sandwich ELISA. Changes in serum IL-8 levels were compared with the Wilcoxon test and their strength of association with response was assessed with the Mann-Whitney test. Accuracy of changes in IL-8 levels to predict response was estimated using receiver operation characteristics (ROC) curves.

Results

Twenty-nine melanoma patients treated with nivolumab or pembrolizumab were studied. In responding patients, serum IL-8 levels significantly decreased between baseline and best response (P < .001), and significantly increased upon progression (P = .004). In non-responders, IL-8 levels significantly increased between baseline and progression (P = .013). Early changes in serum IL-8 levels (2-4 weeks after treatment initiation) were strongly associated with response (P < .001). These observations were validated in 19 NSCLC patients treated with nivolumab or pembrolizumab (P = .001), and in 15 melanoma patients treated with nivolumab plus ipilimumab (P < .001). Early decreases in serum IL-8 levels were associated with longer overall survival in melanoma (P = .001) and NSCLC (P = .015) patients. Serum IL-8 levels also correctly reflected true response in 3 cancer patients presenting pseudoprogression.

Conclusions

Changes in serum IL-8 levels could be used to monitor and predict clinical benefit from immune checkpoint blockade in melanoma and NSCLC patients.

Pseudomonas aeruginosa colonization, prominent inflammation with massive expression of the neutrophil chemokine IL-8, and luminal infiltrates of neutrophils are hallmarks of chronic lung disease in patients with cystic fibrosis (CF). The nociceptive transient receptor potential ankyrin (TRPA) 1 calcium channels have been recently found to be involved in nonneurogenic inflammation. Here, we investigate the role of TRPA1 in CF respiratory inflammatory models in vitro. Expression of TRPA1 was evaluated in CF lung tissue sections and cells by immunohistochemistry and immunofluorescence. Epithelial cell lines (A549, IB3-1, CuFi-1, CFBE41o-) and primary cells from patients with CF were used to: (1) check TRPA1 function modulation, by Fura-2 calcium imaging; (2) down-modulate TRPA1 function and expression, by pharmacological inhibitors (HC-030031 and A-967079) and small interfering RNA silencing; and (3) assess the effect of TRPA1 down-modulation on expression and release of cytokines upon exposure to proinflammatory challenges, by quantitative RT-PCR and 27-protein Bioplex assay. TRPA1 channels are expressed in the CF pseudostratified columnar epithelium facing the bronchial lumina exposed to bacteria, where IL-8 is coexpressed. Inhibition of TRPA1 expression results in a relevant reduction of release of several cytokines, including IL-8 and the proinflammatory cytokines IL-1β and TNF-α, in CF primary bronchial epithelial cells exposed to P. aeruginosa and to the supernatant of mucopurulent material derived from the chronically infected airways of patients with CF. In conclusion, TRPA1 channels are involved in regulating the extent of airway inflammation driven by CF bronchial epithelial cells.

Severe influenza is often associated with disease manifestations outside the respiratory tract. Whilst pro-inflammatory cytokines can be detected in the lungs and blood of infected patients, the role of extra-respiratory organs in the production of pro-inflammatory cytokines is unknown. Here, we show that both pandemic H1N1 and highly pathogenic H5N1 virus induce expression of TNFα, IL-6 and IL-8 in the respiratory tract and central nervous system. In addition, H5N1 virus induced cytokines in the heart, pancreas, spleen, liver and jejunum. Together, these data suggest that extra-respiratory tissues contribute to systemic cytokine responses which may increase the severity of influenza.

Intestinal CD4+ T cell depletion is rapid and profound during early HIV-1 infection. This leads to a compromised mucosal barrier that prompts chronic systemic inflammation. The preferential loss of intestinal T helper 17 (Th17) cells in HIV-1 disease is a driver of the damage within the mucosal barrier and of disease progression. Thus, understanding the effects of new therapeutic strategies in the intestines has high priority. Histone deacetylase (HDAC) inhibitors (e.g., panobinostat) are actively under investigation as potential latency reversing agents in HIV eradication studies. These drugs have broad effects that go beyond reactivating virus, including modulation of immune pathways. We examined colonic biopsies from ART suppressed HIV-1 infected individuals (clinicaltrials.gov: NCT01680094) for the effects of panobinostat on intestinal T cell activation and on inflammatory cytokine production. We compared biopsy samples that were collected before and during oral panobinostat treatment and observed that panobinostat had a clear biological impact in this anatomical compartment. Specifically, we observed a decrease in CD69+ intestinal lamina propria T cell frequency and increased IL-17A mRNA expression in the intestinal epithelium. These results suggest that panobinostat therapy may influence the restoration of mucosal barrier function in these patients.

​STAT3 is considered to play an oncogenic role in several malignancies including lung cancer; consequently, targeting ​STAT3 is currently proposed as therapeutic intervention. Here we demonstrate that ​STAT3 plays an unexpected tumour-suppressive role in ​KRAS mutant lung adenocarcinoma (AC). Indeed, lung tissue-specific inactivation of ​Stat3 in mice results in increased ​KrasG12D-driven AC initiation and malignant progression leading to markedly reduced survival. Knockdown of ​STAT3 in xenografted human AC cells increases tumour growth. Clinically, low ​STAT3 expression levels correlate with poor survival and advanced malignancy in human lung AC patients with smoking history, which are prone to ​KRAS mutations. Consistently, ​KRAS mutant lung tumours exhibit reduced ​STAT3 levels. Mechanistically, we demonstrate that ​STAT3 controls NF-κB-induced ​IL-8 expression by sequestering NF-κB within the cytoplasm, thereby inhibiting ​IL-8-mediated myeloid tumour infiltration and tumour vascularization and hence tumour progression. These results elucidate a novel ​STAT3–NF-κB–​IL-8 axis in ​KRAS mutant AC with therapeutic and prognostic relevance.

Abstract

The existence of rare cancer cells that sporadically acquire drug-tolerance through epigenetic mechanisms is proposed as one mechanism that drives cancer therapy failure. Here we provide evidence that specific microenvironments impose non-sporadic expression of proteins related to epithelial plasticity and drug resistance. Microarrays of robotically printed combinatorial microenvironments of known composition were used to make cell-based functional associations between microenvironments, which were design-inspired by normal and tumor-burdened breast tissues, and cell phenotypes. We hypothesized that specific combinations of microenvironment constituents non-sporadically impose the induction of the AXL and cKIT receptor tyrosine kinase proteins, which are known to be involved in epithelial plasticity and drug-tolerance, in an isogenic human mammary epithelial cell (HMEC) malignant progression series. Dimension reduction analysis reveals type I collagen as a dominant feature, inducing expression of both markers in pre-stasis finite lifespan HMECs, and transformed non-malignant and malignant immortal cell lines. Basement membrane-associated matrix proteins, laminin-111 and type IV collagen, suppress AXL and cKIT expression in pre-stasis and non-malignant cells. However, AXL and cKIT are not suppressed by laminin-111 in malignant cells. General linear models identified key factors, osteopontin, IL-8, and type VIα3 collagen, which significantly upregulated AXL and cKIT, as well as a plasticity-related gene expression program that is often observed in stem cells and in epithelial-to-mesenchymal-transition. These factors are co-located with AXL-expressing cells in situ in normal and breast cancer tissues, and associated with resistance to paclitaxel. A greater diversity of microenvironments induced AXL and cKIT expression consistent with plasticity and drug-tolerant phenotypes in tumorigenic cells compared to normal or immortal cells, suggesting a reduced perception of microenvironment specificity in malignant cells. Microenvironment-imposed reprogramming could explain why resistant cells are seemingly persistent and rapidly adaptable to multiple classes of drugs. These results support the notion that specific microenvironments drive drug-tolerant cellular phenotypes and suggest a novel interventional avenue for preventing acquired therapy resistance.

Idiopathic Pulmonary Fibrosis (IPF) is a progressive fibrotic lung disease, but the mechanisms driving progression remain incompletely defined. We previously reported that the IPF lung harbors fibrogenic mesenchymal progenitor cells (MPCs), which serve as a cell-of-origin for IPF fibroblasts. Proliferating IPF MPCs are located at the periphery of fibroblastic foci in an active cellular front at the interface between the myofibroblast rich focus core and adjacent normal alveolar structures. Among a large set of genes that distinguish IPF MPCs from their control counterparts we identified IL-8 as a candidate mediator of IPF MPC fibrogenicity and driver of fibrotic progression. IPF MPCs and their progeny displayed increased steady state levels of IL-8 and its cognate receptor CXCR1 and secreted more IL-8 than did controls. IL-8 functioned in an autocrine manner promoting IPF MPC self-renewal and the proliferation and motility of IPF MPC progeny. Secreted IL-8 also functioned in a paracrine manner stimulating macrophage migration. Analysis of IPF lung tissue demonstrated co-distribution of IPF MPCs with activated macrophages in the active cellular front of the fibroblastic focus. These findings indicate that IPF MPC derived IL-8 is capable of expanding the mesenchymal cell population and recruiting activated macrophages cells to actively evolving fibrotic lesions.