Restore fingerprint molecule dependent sensibility using inhibitors packed in pH dependent nanostructures

Project facts

Project promoter:
Iasi Regional Institute of Oncology(RO)
Project Number:
RO-RESEARCH-0037
Status:
Completed
Final project cost:
€1,127,325
Donor Project Partners:
University of Oslo(NO)
Other Project Partners
"Petru Poni" Institute of Macromolecular Chemistry(RO)
Programme:

Description

A specific subgroup of breast cancer patients carry a fingerprint molecule named Her2New receptor which is used for many years as a target for specific anticancer therapy. Her2 antibodies effectively block tumour cells development and prevent tumour growth. While effective initially, cancer cells develop metabolic mechanisms to escape therapeutic inhibition and a very valuable therapeutic tool is as such lost. We identified two goals: to manipulate the cancer cells such as to become again responsive to the biological therapy targeting Her2New  receptor; to augment distribution of intended drug such as to limit drug delivery to tumour tissue and limit systemic toxicity associated with non-targeted distribution. The team will produce and test polymeric chains that are capable to change their spatial configuration in relation with the pH of the medium. As such these chains will form a complex mycelia in neutral pH and will act like sponge absorbing the chemical compound needed to be delivered. We will use the specific behaviour of tumour tissue that will induce an acidic environment. .The new approach we develop will also be available for other types of tumours, as all of them share the acidic environment generated by rapid, uncontrolled multiplication of cancer cells.The group has also experience in the introduction of new drugs on the market and they will benefit from the cooperation with the Romanian partners by using a mechanism of feedback in production and in vitro studies that will allow rapid adjustments of nano-carriers structure. 

Summary of project results

Cancer  is a  major contributor to disease  burden worldwide.  Cancer  incidence  and mortality is rapidly  growing, and  female  breast  cancer  surpasses  lung  cancer  as  the  most  commonly diagnosed  cancer.  New  data  supports  the  urgent  need for  comprehensive  cancer  prevention and new treatment to effectively reduce the global burden of breast cancer. One breast cancer subtype,  overexpressing  Her2  receptors - called  Her2  positive,  is frequently  associated  with invasion   and   metastasis.  Trastuzumab   is  the  first   clinically   used  humanized   monoclonal antibody  used in targeted  treatment  of this subtype.  The  toxicity  of  Her2 targeted  therapies and the development  of resistance to treatment  challenge clinicians and researchers to search for  novel and efficient treatment  options. The  resistance to trastuzumab  involves modulation of signaling pathways that prevent biological effects of trastuzumab.  Modifying these pathways should   prevent   the  appearance  of   resistance   and  allow  for   restoration   of   response  to treatment,  allowing patients to further  benefit from trastuzumab treatment.  While  numerous alternatives  are  researched,  our  challenge  was  to  restore the efficacy  of  long term treatment with   trasutzumab   using   functionalized    nanocarriers.   These   nanosize   particles   will   use trastuzumab  for  targeting  cancer  cells  overexpressi ng  Her2  receptor,  and  should  be  loaded with an inhibitor of the Axl receptor expressed on the cancer cell surface.

The  project started  by identifying a very  efficient  pH-dependent  release nanostructure.  We have tested three  type of  micelles  and we  have selected·.for further  investigations  the one that is most stable at the physiological pH {7.6 -7:4) but in the same time with most efficient release at  pH 7.2. We  have focused  our  research on pH 7:''l sensitive  nanostructures  due to the  fact   that   primary  tumors   {where   pH · is  very   acidic)   are· surgically   removed   while metastases  and  the  remaining tissue  adjacent  to  primary  tumors  have a  pH  closer  to  the physiological  one.  Up  to  40%  of  women  develop  local  recurrence  after  surgery,  despite apparently  tumor-free  margins.  In addition  to ''the  physical-chemically  characterization,  the release have been also  investigated in cancer cell cultures.  The selected  micelles have been functionalized  by linking trastuzumab  for targeting the cancer cells. The  biological  effects of functionalized  micelles have been tested  by various techniques  in several  Her2 positive  and negative cancer  cell lines and in laboratory mice. Animal  experiments  have been performed according  with  the  European  legislation  on  the  protection  of  animals  used  for  scientific research.   Functionalized   micelles  have  been  loaded  with   Axl   inhibitor   and   investigate biological effect using sensitive and induced trastuzumab  resistant clones.

Using a Her2 positive cancer cell line, linking trastuzumab to micelles enhance therapeutical effects compared to free trastuzumab. Extending investigations to demonstrate the potentiated therapeutical effect of linked trastuzumab to micelles, we have used additional cell lines, both human Her2 positive and human and murine Her2 negative, as negative controls. Her2 negative cell lines have a low expression of Her2 receptor, while Her2 positive overexpress this receptor. As expected, free trastuzumab impaired cell viability only in the Her2 positive and responsive to treatment cell lines while did not reduce cell viability of tested Her2 negative cell lines. To our surprise, linked trastuzumab to micelles significantly impaired cell viability of particularly Her2 negative selected cell lines. Our overall results, demonstrates that trastuzumab functionalized micelles reduce metastasis, the process of spreading cancer by forming metastases in distal organs. This may open the possibility that these particular patients which have chemotherapy as the only systemic therapeutic alternative to benefit from the new therapy option. The results on Her2 positive investigations revealed a double mechanism of resistance, by increasing Axl expression and by decreasing Her2 expression. The release of Axl inhibitor loaded into functionalized micelles restores the therapeutically effects of trastuzumab in resistant clone.

Summary of bilateral results

Working in partnership is a complex and challenging activity requiring new skills for adapting new and controversial ideas and being able to negotiate and compromise in the group. Numerous meetings and brainstorming sessions made us better in recognizing how to make a step forward and to be able to identify a good solution and a dead end in research. We wrote together 3 more application for research and better understood how to divide work between partners and join together in assessing results. This project made us able to better estimate each partner''s strong points and how to mix these qualities together in a major project. We were exposed to criticism from our partners and by doing it we were all able to better evaluate our work and discuss steps in developing new experiments and analyse the results.

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