Rotterdam, The Netherlands

At the Josephine Nefkens Institute, Prof. Dr. Jan Trapman (head of the Department of Pathology) and Dr. Erik Jan Dubbink are using PamGene's Nuclear Hormone Receptor PamChip®4 Microarrays for their research on prostate cancer using androgen receptors.
Erik Jan explains,

Metastasized prostate cancer is a life-threatening disease for which no curative treatment is available. Because prostate cancer growth depends on activation of the androgen receptor (AR) by androgens, standard treatments of progressive disease aim at withdrawal of circulating androgens and direct blockade of AR function by anti-hormones. Unfortunately, virtually all patients will be confronted with a fatal remission of an endocrine therapy resistant tumor within a few years. AR still plays a central role in therapy-resistant cancer.

Our research program focuses on alternative or complementary strategies to block AR function. A candidate novel target for pharmacological intervention is the AR co-activator groove in the ligand-binding domain which is involved in cofactor binding and homodimerization via FXXLF-like motifs. In recent years we have defined many properties required for (specific) binding of FXXLF-like peptides to the AR groove.

The photo shows Eric Jan(left) working on the PamStation® together with Martijn Dankers of PamGene

Groningen, The Netherlands

Prof Dr W.A. Kamps' laboratory is using Tyrosine Kinase PamChip Microarrays as part of its focus on various aspects of modulating resistance in childhood cancer. The central theme of these investigations is the induction of cell death and/or decrease in tumor growth. As Dr. Eveline de Bont explains,

"We have specifically focused on the role of angiogenesis and/or postnatal vasculogenesis in induction of tumor cell death and decrease in tumor growth in hematological malignancies as well as more recently, pediatric brain tumors. The objective is to translate the results in new treatment strategies in addition to cytostatic drug therapy, increasing the survival rate and reducing therapy-associated side effects of children".

Groningen, The Netherlands

The research groups of Prof. Maikel Peppelenbosch of the Immunology Section are also using Tyrosine Kinase PamChip(R) Microarrays as part of their focus on the cell's biological basis of pathophysiology working on two assumptions.

"Our first assumption", as Dr Sander Diks describes, "is that morphogens remain important in adult organisms, especially for pathophysiology. Secondly the knowledge of cellular signal transduction will generate novel avenues for therapeutic treatment of disease. Important successes obtained hitherto were the development of kinome profiling as a tool for generating comprehensive descriptions of cellular kinase activity. Additionally, the identification of the role of hedgehog signaling for maintaining histostability in face of the relentless proliferation-differentiation-apoptosis sequence operative in organs".

Utrecht, The Netherlands

The Netherlands Proteomics Centre is a platform for hypothesis-driven research that will develop technologies and tools for researchers and product developers in knowledge institutes and industry. The NPC will improve and widen fundamental technological tools in proteomics research, and through well-organized knowledge transfer will make them available to the biological and biomedical research communities at large. With the help of these new tools proteomics research will contribute to a better understanding of processes of health and disease, and pave the way for the generation of new means for intervention and prevention of undesired life-processes. The infrastructure within the NPC is already such that top researchers work together. The NPC also includes a proteomics "hotel facility" intended for visiting academic and industrial researchers, both within and outside the centre, including European partners.
The hotel facility is equipped with PamGene instrumentation that is currently applied in several research projects

Amsterdam, The Netherlands

Dr. Connie R. Jimenez is head of the recently established OncoProteomics Laboratory (OPL), Cancer Center Amsterdam, VU University Medical Center, and she coordinates the VUMC-clinical proteomics activities in the context of the Centre for Medical Systems Biology, a national genomics center of excellence. The OPL proteomics infrastructure consists of a 4800 MALDI-TOF/TOF mass spectrometer and 2DLC instrumentation with the nanoLC part coupled to the Advion NanoMate and a LTQ-FT mass spectrometer.

The mission of the OPL is to develop and implement innovative proteomics technologies and data analysis methods to improve diagnostics and treatment of cancer. To this end, a major focus is on biomarker discovery in biofluids such as blood-serum/plasma, CSF and nipple aspirate fluid that can be collected non-invasively. Biofluids are profiled using two complementary approaches for the discovery of diagnostic, predictive and drug response patterns and biomarkers:
1. A high-throughput MALDI-TOF mass spectrometry-based method for proteomic pattern analysis of large cohorts of human serum and discovery of diagnostic, predictive and drug-response signatures.
2. Label-free (nano)LC-LTQ-FT mass spectrometry of subfractionated body fluids for in-depth profiling and discovery of predictive and drug-response biomarkers.

Other research lines include the analysis of drug-induced alterations in cancer cells and cancer cell conditioned media, organelle proteomics of tumor tissue (eg., the cell nucleus) and tumor-derived exosomes, as well as analysis of cell-surface markers in vitro and in vivo and (phospho)proteomics of protein complexes.

We are exploring new avenues of phosphoproteomics by means of kinase-activity profiling using the PamChip® Arrays. To this end, in collaboration with PamGene, we analyze model cell lines after growth factor treatment, cancer cell lines after drug treatment and colon tumour tissue biopsies in different stages of adenoma-to-carcinoma progression.

Amsterdam, The Netherlands

Prof. Jaap M.Middeldorp, Department of Pathology, VU University Medical Centre, Amsterdam is using our platform in his studies on "Kinase Profiling in EBV-Mediated Tumour Immune Escape." As he explains,

"The Epstein-Barr Virus (EBV) is a ubiquitous human tumour virus, etiologically linked to the genesis of both lymphomas and carcinomas. In EBV+ tumour cells viral proteins are expressed, yet these cells escape immune elimination by T-cells. Virus-driven immune escape involves multiple mechanisms, one being direct T-cell silencing by the viral oncogen LMP1. LMP1 is expressed in most EBV+ tumors and is secreted into the tumour micro-environment in the form of MHC-II rich exosomes. Purified recombinant LMP1 and LMP1 containing exosomes inhibit T-cell activation by antigens, mitogens and CD#+CD28 receptor cross-linking. This effect is mediated by an evolutionary conserved 6 amino acid domain in the first transmembrane helix of LMP1 (TM1).

Our current work aims to resolve molecular pathway(s) leading to T-cell silencing by LMP1. We use the PamGene Kinase platform for kinome profiling in Jurkat T-cells activated for different times (minutes) by anti-CD3/CD28 in the presence or absence of purified LMP1 or LMP1 TM1 deletion mutants."

Leiden, The Netherlands

The collaboration between Dr. Pieter van der Velden, Dr. Willem Maat and Prof. Martine Jager of the LUMC's department of Opthalmology and Dermatology is investigating the pathways that stimulate the proliferation of uveal melanoma. As Pieter van der Velden describes,

"Our primary approach is looking at the genetics of oncogenes but we also approach proliferation in uveal melanoma with functional analysis of signalling pathways that are known to be activated in uveal melanoma*.

In order to explore signalling pathways not yet known to be involved we are using the Tyrosine Kinase PamChip(R) Microarrays. Initial analysis of uveal melanoma is based on cell lines of primary uveal melanoma and metastatic melanoma". The validation is being performed on primary tumour tisue.

*Zuidervaart W, van Nieuwpoort F, Stark M, Dijkman R, Packer L, Borgstein AM, Pavey S, van der Velden P, Out C, Jager MJ, Hayward NK, Gruis NA. Activation of the MAPK pathway is a common event in uveal melanomas although it rarely occurs through mutation of BRAF or RAS. Br J Cancer. 2005 Jun 6;92(11):2032-8.

Oslo, Norway

Dr. Anne Hansen Ree is using PamGene's Kinase PamChip® Microarrays for looking at kinase activity pathways associated with prostate cancer metastasis to bone. As she explains below,

"Prostate cancer patients commonly develop painful and debilitating bone metastases, but regulatory mechanisms involved in the formation of osteoblastic bone metastasis in prostate cancer are inadequately understood. Androgens are critical regulators of prostate cancer growth and progression, and most patients, also those with metastatic disease, respond temporarily to androgen ablation therapy. To study regulatory mechanisms in the interaction between prostate carcinoma cells and bone osteoblasts, we have co-cultured the two cell types in the absence and presence of androgen, and analyzed kinase activity pathways (by means of a peptide substrate phosphorylation array; PamGene International B.V.) within each cell type after they have been segregated by immunomagnetic target cell selection. This experimental approach may provide information that might translate into therapeutic strategies in prostate cancer metastasis to bone."

Utrecht, The Netherlands

The collaboration with Eric Kalkhoven's group was set up in January 2007. Eric has a great interest in Nuclear Receptors (NRs) and co-activators and his research group focuses on PPARγ in fat tissue. PamGene's Nuclear Receptor PamChip® Microarrays, with their wide selection of different LXXLL en LXXXIXXXL peptides will be used to make "interaction profiles". In a first series of experiments, post-doc Arjen Koppen is going to look at the 3 different PPAR proteins, in combination with different ligands. An important part of the project will be to prove the predictive value of the NR-peptides. Does the intact, LXXLL-containing protein interact with the NR under the influence of the agonist or antagonist in a living cell too? If correlations between the PamChip® results and the cell-based assay results are robust, then PamGene's technology will be an important new step in the development of new medicines that target NRs

Eric's group (L-R): Ellen Jeninga (PhD student), Yuan Gao (PhD student), Eric Kalkhoven, Olivier van Beekum (PhD student), Anneke Post (student), Nicole Hamers (analyst), Veerle Fleskens (student), Arjen Koppen (post-doc). Missing from photo: Brenda Hendriks-Stegeman (analist).