Tyrosine Kinases and Cancer Therapy
Prof. Carlo Gambacorti-Passerini
University of Milano-Bicocca, Italy
 

Tyrosine kinases (TKs) have been linked to various types of human cancer, from leukemias to solid tumors. Fundamental differences exists however in the evidence linking a certain TK to a particular type of cancer.

On one side are cancers in which a causal link between a certain TK and that particular tumor is well established, such as chronic myeloid leukemias (CML), gastrointestinal stromal tumors (GIST), lung adenocarcinomas carrying EGFR mutations. These tumors usually present a structural alteration of the TK, originating from a genomic mutation or translocation.

On the other side are instead tumors in which no causal link have been established and in which a certain TK is merely expressed or over expressed; examples in this group include glioblastomas and small cell lung cancer (SCLC).

An intermediate group is composed by tumors in which no structural alteration of the TK is present, but in which genetic events such as gene amplifications can be observed: an example for this group is presented by HER2/Neu+ breast carcinomas.
 

A separate group is constituted by TKs that are not expressed on the tumor cells but are present on normal cells such as endothelial cells, and could be relevant for tumor growth: an example is the vascular endothelial growth factor receptor (VEGFR).

Several TK inhibitors (TKI) have been developed and are now well established in the clinical arena. Examples are imatinib, dasatinib, gefitinib, erlotinib, sunitinib, bevacizumab, trastuzumab, cetuximab.

This presentation will deal with the use of TKI in various cancers; in particular it will discuss the clinical results obtained in relationship with the type of TKI and of cancer considered. It will also present and discuss the molecular mechanisms that can cause resistance to TKIs.

 

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The structural analysis of tyrosine kinases reveals a remarkable conformational plasticity. While in their active state (on-state) they adopt a strikingly similar structure, they adopt a wide range of conformations in their inactive state (off-state). On the one hand, this represents a challenge for the rational design of inhibitors of tyrosine kinases, on the other hand it offers the possibility of achieving selectivity and designing different types of inhibitors by exploiting binding sites formed along the conformational space covered over time by the protein. Several questions rose: are all possible conformations also targetable, does the ligand exploit the induced fit or/and conformational selection mechanism to bind, can we go beyond the structural information of crystal structures, can this challenge be addressed with predictive tools used in structure based design, can molecular probes help understanding the structural issues.

These issues will be discussed and exemplified by data taken from the literature and the anaplastic lymphoma kinase case study. Anaplastic lymphoma kinase (ALK) is aberrantly activated in cancer such as anaplastic large cell lymphoma (ALCL), inflammatory myofibroblastic tumors and diffuse large B cell lymphoma and thus a promising target for anti-cancer therapy. No crystal structure is publicly available up to now. Different homology models have been built to partially cover the conformational space and submitted to experimental validation using molecular probes and site directed mutagenesis. The assessed models were used for virtual screening and allowed the identification of hit compounds with inhibitory activity against ALK in the low micromolar range.
 

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ErbB Family of Tyrosine Kinases
Prof. Janos Szőllösi
Department of Biophysics and Cell Biology, University of Debrecen, Debrecen, Hungary
 

Members of the epidermal growth factor receptor (EGFR) family receptor tyrosine kinases play important roles in cell proliferation, differentiation, apoptosis and migration. The four known members of the family: ErbB1 (EGFR), ErbB2 (HER2/Neu), ErbB3 (HER3) and ErbB4 (HER4) may act as signal transducers at the cell membrane. Unlike other ErbB receptors, ErbB2 has no known ligand and amplification of this receptor can cause breast, ovarian, gastric and salivary cancers. Recent biochemical and biophysical evidence suggests that this protein operates as a shared receptor subunit with other ErbB proteins. Its medical importance stems form its frequent overexpression in breast and other cancers, resulting in various tumorigenic phenotypic changes, including higher transforming activity, metastatic potential, angiogenesis and drug resistance. Humanized antibodies against ErbB2 (i.e. Herceptin) have been introduced into clinical practice and were found to have cytostatic effect in ~40% of ErbB2 positive breast tumors. Our working hypothesis is that expression levels of ErbB kinases, their interactions and activity within multimolecular complexes and their lipid environment will determine the outcome of ErbB2 directed therapy.

We used Herceptin resistant (JIMT-1, MKN-7) and sensitive (SKBR-3, N-87) cell lines in order to demonstrate the importance of association pattern ErbB molecules with each other and with integrins, CD44 and lipid rafts. Herceptin-sensitive cell lines expressed more ErbB2 and fewer b1-integrin and CD44 molecules on their surface than their resistant counterparts, this finding probably does not explain the Herceptin resistant phenotype due to the weak interaction between b1-integrins and ErbB2 and between CD 44 and ErbB2. We have found that in the resistant cell lines active ErbB2 homodimers that bind Herceptin with high affinity are scarce, and we suggested that signaling that drives proliferation may originate from other ErbB kinase dimers such as the ErbB2-ErbB3 heterodimers.
 

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