Defining the critical hurdles in cancer immunotherapy.

Scientific discoveries that provide strong evidence of antitumor effects in preclinical models often encounter significant delays before being tested in patients with cancer. While some of these delays have a scientific basis, others do not. We need to do better.

New challenges in endpoints for drug development in advanced melanoma.

During the past 3 decades, the field of clinical research for the treatment of advanced melanoma lacked significant advances. Available drugs had low antitumor activity and no proven benefit in overall survival. Recently, new drugs developed based on an in-depth understanding of the biology of this disease have shown significant benefit, with ipilimumab and vemurafenib having recently shown a positive impact in overall survival in patients with metastatic melanoma leading to approval in this indication by the U.

Antitumor activity from antigen-specific CD8 T cells generated in vivo from genetically engineered human hematopoietic stem cells.

The goal of cancer immunotherapy is the generation of an effective, stable, and self-renewing antitumor T-cell population. One such approach involves the use of high-affinity cancer-specific T-cell receptors in gene-therapy protocols. Here, we present the generation of functional tumor-specific human T cells in vivo from genetically modified human hematopoietic stem cells (hHSC) using a human/mouse chimera model.

RAF inhibitor resistance is mediated by dimerization of aberrantly spliced BRAF(V600E).

Activated RAS promotes dimerization of members of the RAF kinase family. ATP-competitive RAF inhibitors activate ERK signalling by transactivating RAF dimers. In melanomas with mutant BRAF(V600E), levels of RAS activation are low and these drugs bind to BRAF(V600E) monomers and inhibit their activity.

Anti-CTLA4 monoclonal antibodies: the past and the future in clinical application.

Recently, two studies using ipilimumab, an anti-CTLA-4 monoclonal antibody (mab) demonstrated improvements in overall survival in the treatment of advanced melanoma. These studies utilized two different schedules of treatment in different patient categories (first and second line of treatment). However, the results were quite similar despite of different dosage used and the combination with dacarbazine in the first line treatment.

Decitabine immunosensitizes human gliomas to NY-ESO-1 specific T lymphocyte targeting through the Fas/Fas ligand pathway.

Background: The lack of effective treatments for gliomas makes them a significant health problem and highlights the need for the development of novel and innovative treatment approaches. Immunotherapy is an appealing strategy because of the potential ability for immune cells to traffic to and destroy infiltrating tumor cells. However, the absence of well-characterized, highly immunogenic tumor-rejection antigens (TRA) in gliomas has limited the implementation of targeted immune-based therapies.

Feeding behaviour after injection of α-adrenergic receptor agonists into the median raphe nucleus of food-deprived rats.

This study investigated the participation of median raphe nucleus (MnR) α1-adrenergic receptors in the control of feeding behaviour. The α1-adrenergic agonist phenylephrine (PHE) and α2-adrenergic agonist clonidine (CLON) (at equimolar doses of 0, 6 and 20 nmol) were injected into the MnR of: a) rats submitted to overnight fasting (18 h); or b) rats maintained with 15 g of lab chow/day for 7 days. Immediately after the drug injections, the animals were placed in the feeding chamber and feeding and non-ingestive behaviours such as grooming, rearing, resting, sniffing and locomotion were recorded for 30 min.

Pioneer GABA cells comprise a subpopulation of hub neurons in the developing hippocampus.

Connectivity in the developing hippocampus displays a functional organization particularly effective in supporting network synchronization, as it includes superconnected hub neurons. We have previously shown that hub network function is supported by a subpopulation of GABA neurons. However, it is unclear whether hub cells are only transiently present or later develop into distinctive subclasses of interneurons.

Immunomodulation by imiquimod in patients with high-risk primary melanoma.

Imiquimod is a synthetic Toll-like receptor 7 (TLR7) agonist approved for the topical treatment of actinic keratoses, superficial basal cell carcinoma, and genital warts. Imiquimod leads to an 80-100% cure rate of lentigo maligna; however, studies of invasive melanoma are lacking. We conducted a pilot study to characterize the local, regional, and systemic immune responses induced by imiquimod in patients with high-risk melanoma.

Combinatorial treatments that overcome PDGFRβ-driven resistance of melanoma cells to V600EB-RAF inhibition.

(V600E)B-RAF mutation is found in 50% to 60% of melanomas, and the novel agents PLX4032/vemurafenib and GSK2118436 that inhibit the (V600E)B-RAF kinase achieve a remarkable clinical response rate. However, as might be expected, acquired clinical resistance to these agents arises in most melanoma patients. PLX4032/vemurafenib resistance that arises in vivo in tumor matched short-term cultures or in vitro in melanoma cell lines is not caused by acquisition of secondary mutations in (V600E)B-RAF but rather is caused by upregulating platelet-derived growth factor receptor β (PDGFRβ) or N-RAS which results in resistance or sensitivity to mitogen-activated protein (MAP)/extracellular signal-regulated (ERK; MEK) kinase inhibitors, respectively.

A mechanistic proof-of-concept clinical trial with JX-594, a targeted multi-mechanistic oncolytic poxvirus, in patients with metastatic melanoma.

JX-594 is a targeted and granulocyte macrophage-colony stimulating factor (GM-CSF)-expressing oncolytic poxvirus designed to selectively replicate in and destroy cancer cells through viral oncolysis and tumor-specific immunity. In order to study the mechanisms-of-action (MOA) of JX-594 in humans, a mechanistic proof-of-concept clinical trial was performed at a low dose equivalent to ≤10% of the maximum-tolerated dose (MTD) in other clinical trials. Ten patients with previously treated stage IV melanoma were enrolled.

First report of Troglotrema acutum (Digenea, Troglotrematidae) in the Eurasian badger Meles meles in the Iberian Peninsula and presumptive lesions caused in the host.

A total of 109 badger Meles meles skulls from Catalonia (north-eastern Iberian Peninsula) were studied for helminths. The tremadode Troglotrema acutum is reported here for the first time in the Eurasian badger in the Iberian Peninsula and southern Europe. Three methodologies were used to detect this trematode: an examination for surface lesions, axial computed tomography and fresh skull dissection.

Improved survival with vemurafenib in melanoma with BRAF V600E mutation.

Background: Phase 1 and 2 clinical trials of the BRAF kinase inhibitor vemurafenib (PLX4032) have shown response rates of more than 50% in patients with metastatic melanoma with the BRAF V600E mutation.

Methods: We conducted a phase 3 randomized clinical trial comparing vemurafenib with dacarbazine in 675 patients with previously untreated, metastatic melanoma with the BRAF V600E mutation. Patients were randomly assigned to receive either vemurafenib (960 mg orally twice daily) or dacarbazine (1000 mg per square meter of body-surface area intravenously every 3 weeks).

Combination therapy with vemurafenib (PLX4032/RG7204) and metformin in melanoma cell lines with distinct driver mutations.

Background: A molecular linkage between the MAPK and the LKB1-AMPK energy sensor pathways suggests that combined MAPK oncogene inhibition and metabolic modulation of AMPK would be more effective than either manipulation alone in melanoma cell lines.

Materials And Methods: The combination of the BRAF inhibitor vemurafenib (formerly PLX4032) and metformin were tested against a panel of human melanoma cell lines with defined BRAF and NRAS mutations for effects on viability, cell cycle and apoptosis. Signaling molecules in the MAPK, PI3K-AKT and LKB1-AMPK pathways were studied by Western blot.

BRAF targeted therapy changes the treatment paradigm in melanoma.

After decades of stagnation, recent therapeutic advances in melanoma seem on the horizon. The discovery of the genetic underpinnings of this historically refractory disease has exposed potential targets for therapy, BRAF mutations being principal among them. In the 8 years following the discovery of BRAF mutations in 50-60% of advanced melanomas, only recently have potent and selective inhibitors of this intracellular signaling molecule shown efficacy from early clinical testing.

A clinical microchip for evaluation of single immune cells reveals high functional heterogeneity in phenotypically similar T cells.

Cellular immunity has an inherent high level of functional heterogeneity. Capturing the full spectrum of these functions requires analysis of large numbers of effector molecules from single cells. We report a microfluidic platform designed for highly multiplexed (more than ten proteins), reliable, sample-efficient (∼1 × 10(4) cells) and quantitative measurements of secreted proteins from single cells.

Agrobacterium-mediated genetic transformation of Coffea arabica (L.) is greatly enhanced by using established embryogenic callus cultures.

Background: Following genome sequencing of crop plants, one of the main challenges today is determining the function of all the predicted genes. When gene validation approaches are used for woody species, the main obstacle is the low recovery rate of transgenic plants from elite or commercial cultivars. Embryogenic calli have frequently been the target tissue for transformation, but the difficulty in producing or maintaining embryogenic tissues is one of the main problems encountered in genetic transformation of many woody plants, including Coffea arabica.

Organization and molecular evolution of a disease-resistance gene cluster in coffee trees.

Background: Most disease-resistance (R) genes in plants encode NBS-LRR proteins and belong to one of the largest and most variable gene families among plant genomes. However, the specific evolutionary routes of NBS-LRR encoding genes remain elusive. Recently in coffee tree (Coffea arabica), a region spanning the SH3 locus that confers resistance to coffee leaf rust, one of the most serious coffee diseases, was identified and characterized.

CTLA4 blockade induces frequent tumor infiltration by activated lymphocytes regardless of clinical responses in humans.

Background: CTLA4 blocking monoclonal antibodies provide durable clinical benefit in a subset of patients with advanced melanoma mediated by intratumoral lymphocytic infiltrates. A key question is defining whether the intratumoral infiltration (ITI) is a differentiating factor between patients with and without tumor responses.

Methods: Paired baseline and postdosing tumor biopsy specimens were prospectively collected from 19 patients with metastatic melanoma, including 3 patients with an objective tumor response, receiving the anti-CTLA4 antibody tremelimumab within a clinical trial with primary endpoint of quantitating CD8(+) cytotoxic T-lymphocyte (CTL) infiltration in tumors.

A beta-camera integrated with a microfluidic chip for radioassays based on real-time imaging of glycolysis in small cell populations.

Unlabelled: An integrated β-camera and microfluidic chip was developed that is capable of quantitative imaging of glycolysis radioassays using (18)F-FDG in small cell populations down to a single cell. This paper demonstrates that the integrated system enables digital control and quantitative measurements of glycolysis in B-Raf(V600E)-mutated melanoma cell lines in response to specific B-Raf inhibition.

Methods: The β-camera uses a position-sensitive avalanche photodiode to detect charged particle-emitting probes within a microfluidic chip.

Future perspectives in melanoma research. Meeting report from the "Melanoma Research: a bridge Naples-USA. Naples, December 6th-7th 2010".

Progress in understanding the molecular basis of melanoma has made possible the identification of molecular targets with important implications in clinical practice. In fact, new therapeutic approaches are emerging from basic science and it will be important to implement their rapid translation into clinical practice by active clinical investigation. The first meeting of Melanoma Research: a bridge Naples-USA, organized by Paolo A.

Notocotylus loeiensis N. sp. (Trematoda: Notocotylidae) from Rattus losea (Rodentia: Muridae) in Thailand.

Notocotylus loeiensis n. sp. (Trematoda: Notocotylidae) is described from the cecum of the lesser rice field rat (Rattus losea), from Loei Province in Thailand with a prevalence of 9.

Intra-lymph node prime-boost vaccination against Melan A and tyrosinase for the treatment of metastatic melanoma: results of a phase 1 clinical trial.

Purpose: The goal of this study was to test the safety and activity of a therapeutic vaccine, MKC1106-MT, in patients with metastatic melanoma.

Experimental Design: MKC1106-MT comprises a plasmid (pMEL-TYR) and two peptides (E-MEL and E-TYR), corresponding to Melan A and tyrosinase, administered by intra-lymph node injection in a prime-boost sequence. All 18 patients were HLA-A*0201 positive and received a fixed priming dose of plasmid and a low or a high peptide dose.

PTEN loss confers BRAF inhibitor resistance to melanoma cells through the suppression of BIM expression.

This study addresses the role of PTEN loss in intrinsic resistance to the BRAF inhibitor PLX4720. Immunohistochemical staining of a tissue array covering all stages of melanocytic neoplasia (n = 192) revealed PTEN expression to be lost in >10% of all melanoma cases. Although PTEN expression status did not predict for sensitivity to the growth inhibitory effects of PLX4720, it was predictive for apoptosis, with only limited cell death observed in melanomas lacking PTEN expression (PTEN-).