No abstract

Adiponectin, a hormone secreted from adipocytes, has been shown to protect against development of insulin resistance, ischemia-reperfusion injury, and inflammation. Adiponectin assembles into multiple oligomeric isoforms: trimers, hexamers and several higher molecular weight (HMW) species. Of these, the HMW species are selectively decreased during the onset of type 2 diabetes. Despite the critical role of HMW adiponectin in insulin responsiveness, its assembly process is poorly understood. In this report, we investigated the role of divalent cations in adiponectin assembly. Purified adiponectin 18mers, the largest HMW species, did not collapse to smaller oligomers after treatment with high concentrations of EDTA. However, treatment with EDTA or another chelator DTPA inhibited the oligomerization of 18mers from trimers in vitro. Zn(2+) specifically increased the formation of 18mers when compared with Cu(2+), Mg(2+), and Ca(2+). Distribution of adiponectin oligomers secreted from zinc chelator TPEN-treated rat adipocytes skewed toward increased proportions of hexamers and trimers. While we observed presence of zinc in adiponectin purified from calf serum, the role of zinc in disulfide bonding between oligomers was examined because the process is critical for 18mer assembly. Surprisingly, Zn(2+) inhibited disulfide bond formation early in the oligomerization process. We hypothesize that initial decreases in disulfide formation rates could allow adiponectin subunits to associate before becoming locked in fully oxidized conformations incapable of further oligomerization. These data demonstrate that zinc stimulates oligomerization of HMW adiponectin and possibly other disulfide-dependent protein assembly processes.

Abbreviations: GLUT4, glucose transporter 4; IL, interleukin; IRS2, insulin receptor substrate 2; PI3K, phosphoinositide 3-kinase; p38 MAPK, p38 mitogen-activated protein kinase; PPAR, peroxisome proliferator-activated receptor; TNF, tumor necrosis factor.

Programmed cell death ligand-1 (PD-L1), expressed on both tumor cells (TC) and tumor-associated immune cells (IC), has been shown to be a useful biomarker and predictive of response to anti-PD-L1 agents in certain tumor types. In recurrent or metastatic head and neck squamous cell carcinoma (R/M HNSCC), there is a growing interest in the role of PD-L1 expression on ICs, as well as TCs, for predicting response to immune checkpoint inhibitors. Using pooled data from the phase II HAWK and CONDOR studies, we investigated the association of baseline PD-L1 expression with durvalumab efficacy in patients with R/M HNSCC. To determine an optimal PD-L1 cut-off point for predicting survival, we assessed PD-L1 expression levels at different TC and IC cut-off points in patients treated with durvalumab. Longer survival was associated with higher TC membrane PD-L1 expression and IC staining. When the combined TC/IC algorithm was applied, a cut-off point for PD-L1 expression of ≥50% on TCs or ≥25% on ICs (TC ≥ 50%/IC ≥ 25%) showed a higher objective response rate (17.2% vs. 8.8%), longer median progression-free survival (2.8 vs. 1.9 months), and longer median overall survival (8.4 vs. 5.4 months) in the PD-L1–high versus PD-L1–low/negative patient populations, respectively. A scoring algorithm combining PD-L1 expression on TCs and ICs using the cut-off point TC ≥ 50%/IC ≥ 25% was optimal for identifying patients with HNSCC most likely to benefit from durvalumab treatment. The new algorithm is robust and can be reproducibly scored by trained pathologists. Significance: A novel algorithm for PD-L1 expression using the cut-off point TC ≥ 50%/IC ≥ 25% is robust for identifying patients with HNSCC most likely to benefit from durvalumab treatment and can be reproducibly scored by trained pathologists.