1. Targeting HER2-positive breast cancer with trastuzumab-DM1, an antibody-cytotoxic drug conjugate
Ralph H Schwall, Elaine Mai, Ravi V J Chari, Susan D Spencer, Walter A Blättler, Debra L Dugger, Sara R Kenkare-Mitra, John M Lambert, Gail D Lewis Phillips, Lisa M Crocker, Mark X Sliwkowski, Guangmin Li, Robert J Lutz, Hartmut Koeppen, Kathryn L Parsons, Wai Lee T Wong, Frederic S Jacobson Cancer Res . 2008 Nov 15;68(22):9280-90. doi: 10.1158/0008-5472.CAN-08-1776.
HER2 is a validated target in breast cancer therapy. Two drugs are currently approved for HER2-positive breast cancer: trastuzumab (Herceptin), introduced in 1998, and lapatinib (Tykerb), in 2007. Despite these advances, some patients progress through therapy and succumb to their disease. A variation on antibody-targeted therapy is utilization of antibodies to deliver cytotoxic agents specifically to antigen-expressing tumors. We determined in vitro and in vivo efficacy, pharmacokinetics, and toxicity of trastuzumab-maytansinoid (microtubule-depolymerizing agents) conjugates using disulfide and thioether linkers. Antiproliferative effects of trastuzumab-maytansinoid conjugates were evaluated on cultured normal and tumor cells. In vivo activity was determined in mouse breast cancer models, and toxicity was assessed in rats as measured by body weight loss. Surprisingly, trastuzumab linked to DM1 through a nonreducible thioether linkage (SMCC), displayed superior activity compared with unconjugated trastuzumab or trastuzumab linked to other maytansinoids through disulfide linkers. Serum concentrations of trastuzumab-MCC-DM1 remained elevated compared with other conjugates, and toxicity in rats was negligible compared with free DM1 or trastuzumab linked to DM1 through a reducible linker. Potent activity was observed on all HER2-overexpressing tumor cells, whereas nontransformed cells and tumor cell lines with normal HER2 expression were unaffected. In addition, trastuzumab-DM1 was active on HER2-overexpressing, trastuzumab-refractory tumors. In summary, trastuzumab-DM1 shows greater activity compared with nonconjugated trastuzumab while maintaining selectivity for HER2-overexpressing tumor cells. Because trastuzumab linked to DM1 through a nonreducible linker offers improved efficacy and pharmacokinetics and reduced toxicity over the reducible disulfide linkers evaluated, trastuzumab-MCC-DM1 was selected for clinical development.
2. Effect of trypsin concentration on living SMCC-7721 cells studied by atomic force microscopy
Li Li, Jin Yan, Zuobin Wang, Chenchen Xie, Zhengxun Song, Jiajing Zhu J Microsc . 2021 Dec;284(3):203-213. doi: 10.1111/jmi.13053.
Trypsin is playing an important role in the processes of cancer proliferation, invasion and metastasis which require the precise information of morphology and mechanical properties on the nano-scale for the related research. In this work, living human hepatoma (SMCC-7721) cells were treated with different concentrations of trypsin solution. The morphology and mechanical properties of the cells were measured via atomic force microscope (AFM). Statistical analyses of measurement data indicated that with the increase of trypsin concentration, the average cell height and the surface roughness were both increased, but the cell viability, the cell surface adhesion and the elasticity modulus were decreased significantly. The force required to puncture the cells was also gradually reduced. It indicates that trypsin not only hydrolyses the proteins between the cell and the substrate but also the membrane proteins. The results offer valuable clues for the cancerous process study, pathological analysis and trypsin inhibitor drug development. And this work provides an effective way for overcoming the cell membrane in drug injection for cell-targeted therapy.
3. TRAP/SMCC/mediator-dependent transcriptional activation from DNA and chromatin templates by orphan nuclear receptor hepatocyte nuclear factor 4
Sohail Malik, Robert G Roeder, Yun Kyoung Kang, Annika E Wallberg Mol Cell Biol . 2002 Aug;22(15):5626-37. doi: 10.1128/MCB.22.15.5626-5637.2002.
The orphan nuclear receptor hepatocyte nuclear factor 4 (HNF-4) regulates the expression of many liver-specific genes both during development and in the adult animal. Towards understanding the molecular mechanisms by which HNF-4 functions, we have established in vitro transcription systems that faithfully recapitulate HNF-4 activity. Here we have focused on the coactivator requirements for HNF-4, especially for the multicomponent TRAP/SMCC/Mediator complex that has emerged as the central regulatory module of the transcription apparatus. Using a system that has been reconstituted from purified transcription factors, as well as one consisting of unfractionated nuclear extract from which TRAP/SMCC/Mediator has been depleted by specific antibodies, we demonstrate a strong dependence of HNF-4 function on this coactivator. Importantly, we further show a TRAP/SMCC/Mediator-dependence for HNF-4 transcriptional activation from chromatin templates. The latter involves cooperation with the histone acetyltransferase-containing coactivator p300, in accord with a synergistic mode of action of the two divergent coactivators. We also show that HNF-4 and TRAP/SMCC/Mediator can interact physically. This interaction likely involves primary HNF-4 activation function 2 (AF-2)-dependent interactions with the TRAP220 subunit of TRAP/SMCC/Mediator and secondary (AF-2-independent) interactions with TRAP170/RGR1. Finally, recruitment experiments using immobilized templates strongly suggest that the functional consequences of the physical interaction probably are manifested at a postrecruitment step in the activation pathway.
