The biology behind combining poly [ADP ribose] polymerase and androgen receptor inhibition for metastatic castration-resistant prostate cancer.

For about a decade, poly [ADP ribose] polymerases (PARP) inhibitors have been used almost exclusively to treat tumours that are deficient in one of the BRCA genes. In advanced prostate cancer, which is largely driven by the activity of the androgen receptor (AR), accumulating preclinical evidence has suggested an interplay between the AR and PARP, which could be therapeutically exploited independently of defects in the tumour's DNA homologous recombination repair (HRR) machinery. This includes the regulation of HRR genes by the AR, a mutual influence between the activities of PARP and the AR, and the co-localisation of BRCA2 to the retinoblastoma gene in the human genome. Based on these findings, randomised clinical trials have been initiated to study the addition of a PARP inhibitor to AR pathway inhibitor therapy. Three of four randomised studies demonstrated a significantly increased anti-tumour activity in men with metastatic prostate cancer, irrespective of HRR gene alterations. In this review, we summarise the available preclinical evidence that provides the rationale for the combination of inhibitors for PARP and the AR and discuss how it might contribute to the efficacy observed in the clinic.

Efficacy and safety of avelumab treatment in patients with metastatic Merkel cell carcinoma: experience from a global expanded access program.

BACKGROUND: Avelumab, a human anti-programmed death-ligand 1 immunoglobulin G1 monoclonal antibody, showed favorable efficacy and safety in patients with metastatic Merkel cell carcinoma (mMCC) in the phase II JAVELIN Merkel 200 trial, leading to approval in multiple countries. We describe real-world experience with avelumab in patients with mMCC from an expanded access program. METHODS: Eligible patients had mMCC and progressive disease during or after chemotherapy or were ineligible for chemotherapy or clinical trial participation. Patients received an initial 3-month supply of avelumab (administered as 10 mg/kg intravenously every 2 weeks until progressive disease or unacceptable toxicity); resupply was allowed following complete response, partial response, stable disease, or clinical benefit per physician assessment. RESULTS: Between December 15, 2015, and March 4, 2019, 558 of 620 requests from 38 countries were medically approved, and 494 patients received avelumab. Among 240 evaluable patients, the objective response rate was 46.7% (complete response in 22.9%, including 3 of 16 potentially immunocompromised patients), and the disease control rate was 71.2%. The median duration of treatment in evaluable patients with response was 7.9 months (range, 1.0-41.7) overall and 5.2 months (range, 3.0-13.9) in immunocompromised patients. No new safety signals were identified. The expanded access program closed for new requests on December 31, 2018, as required after regulatory approval; benefitting patients continued to receive avelumab. CONCLUSIONS: The avelumab expanded access program for patients with mMCC demonstrated efficacy and safety in a real-world setting, consistent with the results from JAVELIN Merkel 200, and provided a treatment for patients with limited options.

Management of crizotinib therapy for ALK-rearranged non-small cell lung carcinoma: an expert consensus.

Within 4 years of the discovery of anaplastic lymphoma kinase (ALK) rearrangements in non-small cell lung cancer (NSCLC), the ALK inhibitor crizotinib gained US and European approval for the treatment of advanced ALK-positive NSCLC. This was due to the striking response data observed with crizotinib in phase I and II trials in patients with ALK-positive NSCLC, as well as the favorable tolerability and safety profile observed. Recently published phase III data established crizotinib as a new standard of care for this NSCLC molecular subset. A consequence of such rapid approval, however, is the limited clinical experience and relative paucity of information concerning optimal therapy management. In this review, we discuss the development of crizotinib and the clinical relevance of its safety profile, examining crizotinib-associated adverse events in detail and making specific management recommendations. Crizotinib-associated adverse events were mostly mild to moderate in severity in clinical studies, and appropriate monitoring and supportive therapies are considered effective in avoiding the need for dose interruption or reduction in most cases. Therapy management of patients following disease progression on crizotinib is also discussed. Based on available clinical data, it is evident that patients may have prolonged benefit from crizotinib after Response Evaluation Criteria in Solid Tumors-defined disease progression, and crizotinib should be continued for as long as the patient derives benefit.

Different motifs regulate trafficking of SorCS1 isoforms.

The type I transmembrane protein SorCS1 is a member of the Vps10p-domain receptor family comprised of Sortilin, SorLA and SorCS1, -2 and -3. Current information indicates that Sortilin and SorLA mediate intracellular protein trafficking and sorting, but little is known about the cellular functions of the SorCS subgroup. SorCS1 binds platelet-derived growth factor-BB (PDGF-BB) and is expressed in isoforms differing only in their cytoplasmic domains. Here, we identify two novel isoforms of mouse SorCS1 designated m-SorCS1c and -d. In situ hybridization revealed a combinatorial expression pattern of the variants in brain and embryonic tissues. We demonstrate that among the mouse variants, only SorCS1c mediates internalization and that the highly conserved SorCS1c is internalized through a canonical tyrosine-based motif. In contrast, human SorCS1a, whose cytoplasmic domain is completely different from mouse SorCS1a, is internalized through a DXXLL motif. We report that the human SorCS1a cytoplasmic domain interacts with the alphaC/sigma2 subunits of the adaptor protein (AP)-2 complex, and internalization of human SorCS1a and -c is mediated by AP-2. Our results suggest that the endocytic isoforms target internalized cargo to lysosomes but are not engaged in Golgi-endosomal transport to a significant degree.

Arc/Arg3.1 is essential for the consolidation of synaptic plasticity and memories.

Arc/Arg3.1 is robustly induced by plasticity-producing stimulation and specifically targeted to stimulated synaptic areas. To investigate the role of Arc/Arg3.1 in synaptic plasticity and learning and memory, we generated Arc/Arg3.1 knockout mice. These animals fail to form long-lasting memories for implicit and explicit learning tasks, despite intact short-term memory. Moreover, they exhibit a biphasic alteration of hippocampal long-term potentiation in the dentate gyrus and area CA1 with an enhanced early and absent late phase. In addition, long-term depression is significantly impaired. Together, these results demonstrate a critical role for Arc/Arg3.1 in the consolidation of enduring synaptic plasticity and memory storage.

The N-terminus of the serum- and glucocorticoid-inducible kinase Sgk1 specifies mitochondrial localization and rapid turnover.

The serine/threonine protein kinase Sgk1 (serum- and glucocorticoid-inducible kinase 1) is characterized by a short half-life and has been implicated in the control of a large variety of functions in different subcellular compartments and tissues. Here, we analysed the influence of the N-terminus of Sgk1 on protein turnover and subcellular localization. Using green fluorescent protein-tagged Sgk1 deletion variants, we identified amino acids 17-32 to function as an anchor for the OMM (outer mitochondrial membrane). Subcellular fractionation of mouse tissue revealed a predominant localization of Sgk1 to the mitochondrial fraction. A cytosolic orientation of the kinase at the OMM was determined by in vitro import of Sgk1 and protease protection assays. Pulse-chase experiments showed that half-life and subcellular localization of Sgk1 are inseparable and determined by identical amino acids. Our results provide evidence that Sgk1 is primarily localized to the OMM and shed new light on the role of Sgk1 in the control of cellular function.

EBAG9 adds a new layer of control on large dense-core vesicle exocytosis via interaction with Snapin.

Regulated exocytosis is subject to several modulatory steps that include phosphorylation events and transient protein-protein interactions. The estrogen receptor-binding fragment-associated gene9 (EBAG9) gene product was recently identified as a modulator of tumor-associated O-linked glycan expression in nonneuronal cells; however, this molecule is expressed physiologically in essentially all mammalian tissues. Particular interest has developed toward this molecule because in some human tumor entities high expression levels correlated with clinical prognosis. To gain insight into the cellular function of EBAG9, we scored for interaction partners by using the yeast two-hybrid system. Here, we demonstrate that EBAG9 interacts with Snapin, which is likely to be a modulator of Synaptotagmin-associated regulated exocytosis. Strengthening of this interaction inhibited regulated secretion of neuropeptide Y from PC12 cells, whereas evoked neurotransmitter release from hippocampal neurons remained unaltered. Mechanistically, EBAG9 decreased phosphorylation of Snapin; subsequently, association of Snapin with synaptosome-associated protein of 25 kDa (SNAP25) and SNAP23 was diminished. We suggest that the occurrence of SNAP23, Snapin, and EBAG9 also in nonneuronal cells might extend the modulatory role of EBAG9 to a broad range of secretory cells. The conjunction between EBAG9 and Snapin adds an additional layer of control on exocytosis processes; in addition, mechanistic evidence is provided that inhibition of phosphorylation has a regulatory function in exocytosis.

The Golgi protein RCAS1 controls cell surface expression of tumor-associated O-linked glycan antigens.

Tumor immunology has received a large impetus from the identification of tumor-associated antigens. Among them, a monoclonal antibody, 22.1.1, was instrumental in defining a novel tumor-associated antigen that was termed "receptor binding cancer antigen expressed on SiSo cells" (RCAS1). RCAS1 was proposed to induce growth arrest and apoptosis on activated immune cells, mediated by a putative death receptor. Structurally, RCAS1 was predicted to exist as a type II transmembrane protein and in a soluble form. Here, we analyzed occurrence, membrane topology, and subcellular localization of the RCAS1-encoded gene product. RCAS1 was shown to be a ubiquitously expressed type III transmembrane protein with a Golgi-predominant localization. Monoclonal antibody 22.1.1 failed to recognize RCAS1, as demonstrated by confocal microscopy. Instead, we showed that the cognate 22.1.1 epitope is identical with the tumor-associated O-linked glycan Tn (N-acetyl-d-galactosamine, GalNAc). Overexpression of RCAS1 in cell lines that are negative for 22.1.1 surface staining led to the generation of Tn and the closely related TF (Thomsen-Friedenreich, Galbeta1-3GalNAc) antigen, thus providing a functional link to the generation of the 22.1.1 epitope. We suggest that RCAS1 modulates surface expression of tumor-associated, normally cryptic O-linked glycan structures and contributes indirectly to the antigenicity of tumor cells.

5-aza-2'-deoxycytidine-induced expression of functional cancer testis antigens in human renal cell carcinoma: immunotherapeutic implications.

PURPOSE: Limited therapeutic options are presently available for advanced renal cell carcinoma (RCC). This study was designed to define the clinical potential of the DNA hypomethylating agent 5-aza-2'-deoxycytidine (5-AZA-CdR) in human RCC, through its control of the expression of "therapeutic targets" of the cancer testis antigen (CTA) family, and of the tumor-associated antigen RAGE-1, in RCC cells. EXPERIMENTAL DESIGN: Reverse transcription (RT)-PCR assays of a panel of RCC cells treated with 5-AZA-CdR, investigated the induction of the expression of several CTAs and of RAGE-1. Immunoprecipitation and Western blotting assessed whether the expression of CTA-specific mRNA induced by 5-AZA-CdR resulted in a translated protein of appropriate molecular weight. The functional activity of de novo expressed CTA was evaluated using (51)Cr release cytotoxicity assays of 5-AZA-CdR-treated HLA-A2-positive RCC cells using HLA-A2-restricted NY-ESO-1-specific CTLs. RESULTS: Exposure to 5-AZA-CdR invariably induced the expression of the CTA MAGE-1, -2, -3, and -4, GAGE 1-6, and NY-ESO-1 in all of the RCC cells investigated. De novo expression of NY-ESO-1 was persistent, being still detectable 60 days after the end of treatment, and generated a functional protein efficiently recognized by HLA-A2-restricted NY-ESO-1-specific CTLs. 5-AZA-CdR also induced RAGE-1 expression in RAGE-1-negative RCC and sarcoma cells but not in neoplastic cells of different histology. CONCLUSIONS: This study provides the scientific rationale to establish new strategies of chemoimmunotherapy in RCC patients. The well-defined immunogenicity of the investigated CTAs and of RAGE-1 suggest that systemic administration of 5-AZA-CdR represents a promising strategy to enhance the constitutively poor immunogenic potential of RCC cells, and to propose that virtually all RCC patients receive active and/or adoptive CTA- or RAGE-1-based immunotherapy.

Human cytomegalovirus gene products US2 and US11 differ in their ability to attack major histocompatibility class I heavy chains in dendritic cells.

Human cytomegalovirus (HCMV) encodes several proteins that inhibit major histocompatibility complex (MHC) class I-dependent antigen presentation. The HCMV products US2 and US11 are each sufficient for causing the dislocation of human and murine MHC class I heavy chains from the lumen of the endoplasmic reticulum to the cytosol, where the heavy chains are readily degraded. The apparent redundancy of US2 and US11 has been probed predominantly in cultured cell lines, where differences in their specificities were shown for murine and human MHC class I locus products. Here, we expressed US11 and US2 via adenovirus vectors and show that US11 exhibits a superior ability to degrade MHC class I molecules in primary human dendritic cells. MHC class II complexes are unaffected by US2- and US11-mediated attack. We suggest that multiple HCMV-encoded immunoevasions have evolved complementary functions in response to diverse host cell types and tissues.