Contact

© dkfz.de

Jürg P.F. Nüesch, Ph D                           

Tel. +49 6221 424982
FAX +49 6221 424962

Summary

© dkfz.de

Protoparvoviruses were originally detected as contaminants in human tumor cell lines grown in nude rats and nude mice. This became the basis for their natural oncotropism and oncolytic activities. One of these agents, H-1PV is validated as a potential anti-cancer agent in clinical trials of patients suffering of currently untreatable cancers. My Group is interested to study parvovirus host cell interactions on the molecular level to fund the basis for the improvement of safety and efficacy of these viruses in cancer therapy. For this we are addressing the following issues:
  1. Identification of potential marker/functions promoting neoplastic transformation. In naturally permissive cells, rodent PVs stimulate the PDK1/PKB/PKC signalling cascade to counteract stress responses and to ensure productive infection and spreading. Upon host switch to human cells, PVs strongly depend on this signalling pathway to be activated. This makes cancer cells ideal hosts for PV. Complementarities between PV interference in natural host cells and factors promoting permissiveness in human cancer cells are thought to serve as markers for neoplastic transformation.
  2. By studying molecular mechanisms of PV induced egress and oncolysis we intend to design novel PV-derived oncotoxins to arm heterologous viruses for Virotherapy of Cancer and to improve the induction of a bystander effect by inducing an anti-tumor immune response.
  3. To generate a portfolio of self-propagating oncolytic (parvo)viruses for a virotherapy of cancer aiming to provide a panel of therapeutics for patient-tailored and to establish pre-treatment diagnostics.

Current and future projects

© dkfz.de

PV Interference with intracellular signalling cascades

Regulation of the PV non-structural protein NS1 is strongly dependent on the PDK1/PKC/PKB signalling cascade. Conversely PVs, particularly MVM was shown to interfere with this pathway, leading to activation of PDK1 and its downstream targets PKCλ the short-lived PKCη and PKB/Akt1 (Lachmann et al., 2008). In addition we found that ERM family proteins (mediators between actin cytoskeleton and membranes) play essential roles in the virus cycle. Particularly radixin, in conjunction with PKCη was shown to modulate NS1 and capsid phosphorylation (Nüesch et al., 2009). Recently, we were able to show that Rdx/PKCη-complex targets PDK1 for phosphorylation and activation not only in MVM-infected A9 cells but constitutes an internal loop-back activation mechanisms in highly aggressive cancer cells such as glioblastoma multiforma, rendering PDK1 activity independent of its cofactor PIP3 and, hence growth factor signalling through PI3-kinase (Bär et al., 2015). Currently we are aiming to determine PV-induced activation of the PDK1-downstream survival-kinase PKB/Akt1 and its action on PV-late functioning for progeny particle egress and spreading.

Cytotoxicity, Vesicular egress of progeny virions and oncolytic activity

In past, most oncotoxic/oncolytic functions could be assigned to the large parvoviral multifunctional NS1 protein. This regulatory protein targets a multitude of cellular components and pathways leading to cell death independent of previously acquired resistance towards known death inducers such as cisplatin or TRAIL (reviewed in Nüesch et al., 2012, Nüesch and Rommelaere, 2014). A major oncotoxic function of NS1 was identified as a complex with the catalytic subunit of casein kinase II (CKIIα) targeting a variety of illegitimate substrates with the result of cytoskeleton collapse and necrosis (Nüesch et al., 2005; Nüesch and Rommelaere, 2006). This oncotoxicity could be mimicked by composing CKIIα (binding) with the NS1-targeting domain in novel polypeptides leading to the specific death of permissive cells (Nüesch and Rommelaere, 2007). However, although NS1 is capable to kill target tumor cells alone, this is a rather inefficient process and could be improved.

During PV-infection oncolysis appears to be a tightly regulated process associated with a regulated transport of progeny virions from the nuclear periphery to the plasma membrane. This pathway through ER and Golgi involves COPII vesicles and is necessary to introduce post-assembly modification leading to the maturation of progeny particles and induction of cell lysis (Bär et al., 2008; Bär et al., 2013). Recent investigations identified a late produced non-structural protein called SAT as a key-protein for the permeabilization of the plasmamembrane and the release of progeny virions from infected cells (Bretscher et al., in preparation). We are currently investigating to further characterize particle release and spreading. This is of interest, since intracellular tumor-associated antigens (TAAs) are transported as “co-cargoes” to the cellular periphery and might serve, together with viral antigens as DAMPs and PAMPs in order to attract the host immune system after oncolysis. In addition, we are interested, in collaboration with Prof. Dr. Jörg Huwyler (Pharmaceutical Institute University Basel) to exploit this knowledge to design and generate new oncotoxins on the base PV-induced oncolysis to target Hepato Cell Carcinoma (HCC) with non-viral delivery systems (Grossen et al., in prep).

(Parvo)Virotherapy of Cancer

© dkfz.de

Improving Efficacy:  To improve a virotherapy of cancer with self-propagating oncolytic protoparvoviruses, we are analysing the properties of naturally occurring isolates (e.g. LuIII, TVX) and adapted variants aiming to identify determinants for their oncotropism and oncolytic activities. This will not only allow to characterize the genetic drift (Nüesch et al., in prep), but also to identify hot-spots to engineer new variants with enhanced tumor-suppressive potential (Collaboration with Prof. Dr. Antonio Marchini (LOVIT). The (originally) rodent Protoparvoviruses obtain their specificity to target human tumor cells through the special environment offered by the target cell upon transformation. This is slightly different in the tumors of every individual patient. To choose the right therapeutic agent it is therefore important to determine, whether the agent is suitable for an efficient treatment or not. Therefore, we are currently establishing tools for pre- and post-treatment diagnostics to allow in the future individual patient-tailored treatment possibilities.

Tackling Safety Issues: Recently, genetic screens have identified human Protoparvoviruses, close relatives to the rodent species currently under evaluation for cancer therapies. So far, little is known about these viruses. However, we might obtain valuable information from these potential human pathogens about their life cycle and persistence in humans. In collaboration with the In House Corefacility we are currently establishing new screening tools for mABs with distinct specificities. In Collaboration with Dr. Assia Angelova we will establish new diagnostic tools to tackle interactions of these novel human protoparvoviruses with the human host and to relate these to the application of the oncolytic agents in therapeutical approaches (e.g. crossneutralization, tropism and persistence, anti-tumor activities).

Significant Accomplishments

  • Identification of activated PDK1 as a mediator of PV (onco) tropism
  • Identification of phosphoPDK1 as a diagnostic/prognostic marker in human glioma
  • Generation of new oncotoxins on the base of NS1 induced cytolysis
  • Characterization of vesicular egress of PV through ER and Golgi
  • Generation of H-1PV variants surpassing the wild type virus in anti-cancer activity

External Collaborations

Jörg Huwyler, Pharmazeutisches Institut Universität Basel (Switzerland): “Non-viral delivery of Parvoviral Oncotoxins”

Markus Möhler, Universitätsklinikum Mainz (Deutschland): “Anti-tumor activities of oncolytic Parvoviruses”

Lars Redecke, University of Hamburg, Hamburg (Germany): “Determination of the NS1 crystal structure”

Uwe Truyen, Institut für Tierhygiene Universität Leipzig, (Deutschland): “Persisent Infections of porcine parvoviruses in pigs”

Peter Tatterall, Yale University, New Haven (USA): “Parvovirus non-structural protein functioning”

Peter Tatterall, Yale University, New Haven (USA): “Parvovirus small non-structural protein functioning”

Claytus Davis, Ben Gurion University of the Negev, Beer Sheva (Israel): “PV interaction with the host innate immune and stress responses”

to top