Trastuzumab-DM1 causes tumour growth inhibition by mitotic catastrophe in trastuzumab-resistant breast cancer cells in vivo

Introduction Trastuzumab is widely used for the treatment of HER2-positive breast cancer. Despite encouraging clinical results, a significant fraction of patients are, or become, refractory to the drug. To overcome this, trastuzumab-DM1 (T-DM1), a newer, more potent drug has been introduced. We tested the efficacy and mechanisms of action of T-DM1 in nine HER2-positive breast cancer cell lines in vitro and in vivo. The nine cell lines studied included UACC-893, MDA-453 and JIMT-1, which are resistant to both trastuzumab and lapatinib. Methods AlamarBlue cell-proliferation assay was used to determine the growth response of breast cancer cell lines to trastuzumab and T-DM1 in vitro. Trastuzumab- and T-DM1-mediated antibody-dependent cellular cytotoxicity (ADCC) was analysed by measuring the lactate dehydrogenase released from the cancer cells as a result of ADCC activity of peripheral blood mononuclear cells. Severe Combined Immunodeficient (SCID) mice were inoculated with trastuzumab-resistant JIMT-1 cells to investigate the tumour inhibitory effect of T-DM1 in vivo. The xenograft samples were investigated using histology and immunohistochemistry. Results T-DM1 was strongly growth inhibitory on all investigated HER2-positive breast cancer cell lines in vitro. T-DM1 also evoked antibody-dependent cellular cytotoxicity (ADCC) similar to that of trastuzumab. Outgrowth of JIMT-1 xenograft tumours in SCID mice was significantly inhibited by T-DM1. Histologically, the cellular response to T-DM1 consisted of apoptosis and mitotic catastrophe, the latter evidenced by an increased number of cells with aberrant mitotic figures and giant multinucleated cells. Conclusions Our results suggest mitotic catastrophe as a previously undescribed mechanism of action of T-DM1. T-DM1 was found effective even on breast cancer cell lines with moderate HER2 expression levels and cross-resistance to trastuzumab and lapatinib (MDA-453 and JIMT-1).BioMed Central Open acces

Trastuzumab emtansine: mechanisms of action and drug resistance

Trastuzumab emtansine (T-DM1) is an antibody-drug conjugate that is effective and generally well tolerated when administered as a single agent to treat advanced breast cancer. Efficacy has now been demonstrated in randomized trials as first line, second line, and later than the second line treatment of advanced breast cancer. T-DM1 is currently being evaluated as adjuvant treatment for early breast cancer. It has several mechanisms of action consisting of the anti-tumor effects of trastuzumab and those of DM1, a cytotoxic anti-microtubule agent released within the target cells upon degradation of the human epidermal growth factor receptor-2 (HER2)-T-DM1 complex in lysosomes. The cytotoxic effect of T-DM1 likely varies depending on the intracellular concentration of DM1 accumulated in cancer cells, high intracellular levels resulting in rapid apoptosis, somewhat lower levels in impaired cellular trafficking and mitotic catastrophe, while the lowest levels lead to poor response to T-DM1. Primary resistance of HER2-positive metastatic breast cancer to T-DM1 appears to be relatively infrequent, but most patients treated with T-DM1 develop acquired drug resistance. The mechanisms of resistance are incompletely understood, but mechanisms limiting the binding of trastuzumab to cancer cells may be involved. The cytotoxic effect of T-DM1 may be impaired by inefficient internalization or enhanced recycling of the HER2-T-DM1 complex in cancer cells, or impaired lysosomal degradation of trastuzumab or intracellular trafficking of HER2. The effect of T-DM1 may also be compromised by multidrug resistance proteins that pump DM1 out of cancer cells. In this review we discuss the mechanism of action of T-DM1 and the key clinical results obtained with it, the combinations of T-DM1 with other cytotoxic agents and anti-HER drugs, and the potential resistance mechanisms and the strategies to overcome resistance to T-DM1.BioMed Central open acces

Overexpression of CD44 accompanies acquired tamoxifen resistance in MCF7 cells and augments their sensitivity to the stromal factors, heregulin and hyaluronan

Background: Acquired resistance to endocrine therapy in breast cancer is a significant problem with relapse being associated with local and/or regional recurrence and frequent distant metastases. Breast cancer cell models reveal that endocrine resistance is accompanied by a gain in aggressive behaviour driven in part through altered growth factor receptor signalling, particularly involving erbB family receptors. Recently we identified that CD44, a transmembrane cell adhesion receptor known to interact with growth factor receptors, is upregulated in tamoxifen-resistant (TamR) MCF7 breast cancer cells. The purpose of this study was to explore the consequences of CD44 upregulation in an MCF7 cell model of acquired tamoxifen resistance, specifically with respect to the hypothesis that CD44 may influence erbB activity to promote an adverse phenotype. Methods: CD44 expression in MCF7 and TamR cells was assessed by RT-PCR, Western blotting and immunocytochemistry. Immunofluorescence and immunoprecipitation studies revealed CD44-erbB associations. TamR cells (± siRNA-mediated CD44 suppression) or MCF7 cells (± transfection with the CD44 gene) were treated with the CD44 ligand, hyaluronon (HA), or heregulin and their in vitro growth (MTT), migration (Boyden chamber and wound healing) and invasion (Matrigel transwell migration) determined. erbB signalling was assessed using Western blotting. The effect of HA on erbB family dimerisation in TamR cells was determined by immunoprecipitation in the presence or absence of CD44 siRNA. Results: TamR cells overexpressed CD44 where it was seen to associate with erbB2 at the cell surface. siRNA-mediated suppression of CD44 in TamR cells significantly attenuated their response to heregulin, inhibiting heregulin-induced cell migration and invasion. Furthermore, TamR cells exhibited enhanced sensitivity to HA, with HA treatment resulting in modulation of erbB dimerisation, ligand-independent activation of erbB2 and EGFR and induction of cell migration. Overexpression of CD44 in MCF7 cells, which lack endogenous CD44, generated an HA-sensitive phenotype, with HA-stimulation promoting erbB/EGFR activation and migration. Conclusions: These data suggest an important role for CD44 in the context of tamoxifen-resistance where it may augment cellular response to erbB ligands and HA, factors that are reported to be present within the tumour microenvironment in vivo. Thus CD44 may present an important determinant of breast cancer progression in the setting of endocrine resistance

Surface organometallic chemistry : facile mu2-carbene to mu3-carbyne transformation of organocobalt carbonyls on silica surface

mu2-[ROC(O)C(R)]-mu2-(CO)Co2(CO)5 (R = Me, i-Pr, t-Bu, Ph) bridging carbene-type dinuclear Co carbonyls (1) undergo a facile surface-mediated clusterification while chromatographed on silica gel, yielding [mu3-RC(O)OC]Co3(CO)9 (2a) complexes. These latter are then transformed to the corresponding (mu3-RC)Co3(CO)9 (2b) derivatives by silica-mediated decarboxylation. The x-ray diffraction structure of 2a (R = t-Bu) was determined and that of 2b (R = Ph) was confirmed

Molecular elements of ion permeation and selectivity within calcium channels

Voltage-dependent calcium channels are located in the plasma membrane and form a highly selective conduit by which Ca2+ ions enter all excitable cells and some nonexcitable cells. Extensive characterization studies have revealed the existence of one low (T) and five high-voltage-activated calcium channel types (L, N, P, Q, and R). The high voltage-activated calcium channels have been found to exist as heteromultimers, consisting of an alpha(1), beta, alpha(2)/delta, and gamma subunit. Molecular cloning has revealed the existence of 10 channel transcripts, and expression of these cloned calcium channel genes has shown that basic voltage-activated calcium channel function is strictly carried by the corresponding a, subunits. In turn, the auxiliary subunits serve to modulate calcium channel function by altering the voltage dependence of channel gating, kinetics, and current amplitude, thereby creating a likelihood for calcium channels with multiple properties. Although for calcium channels to be effective, Ca2+ ions must enter selectively through the pore of the alpha(1)-subunit, bypassing competition with other extracellular ions. The structural determinants of this highly selective Ca2+ filter reside within the four glutamic acid residues located at homologous positions within each of the four pore-forming segments. Together, these residues form a single or multiple Ca2+ affinity site(s) that entrap calcium ions, which are then electrostatically repulsed through the intracellular opening of the pore. This mechanism of high-selectivity calcium filtration, the spatial arrangement of pore glutamic acid residues; and the coordination chemistry of calcium binding are discussed in this review

Violation of Distribution Symmetry in Statistical Evaluation of Absolute Enantioselective Synthesis

Enantiometric excesses obtained in absolute enantioselective synthesis by chiral autocatalysis (Soai-reaction) were statistically analyzed. Two sets of parallel experiments, which were performed under chemically different conditions, are available. The former group shows some interesting tendencies, but does not give conclusive statistical results. The sample of 84 parallel experiments, providing 39 R- and 45 S-excesses have shown that these data represent two distinct, non-symmetric sets with different non-Gaussian distributions. Clear S preference was found

O-SILYLATION OF ACYLCOBALT TETRACARBONYLS - SYNTHESIS OF A NEW TYPE OF DINUCLEAR MU-2-HYDROXYCARBENE COBALT CARBONYL DERIVATIVES

O-Silylation of acylcobalt tetracarbonyls, RC(O)Co(CO)4, gives the dinuclear carbene complexes [{mu-2-RC(OSiR'3)}(mu-2-CO)Co2(CO)6], of which one derivative (R = Me, R' = Ph) was characterized by X-ray diffraction