Breast cancer is the main malignant tumor that harms women's health. Early detection is of great significance for improving the survival rate and prognosis of patients. MRI is the main method for detecting breast cancer. We plan to produce a contrast agent that can specifically targets breast cancer cells. This contrast agent is based on magnetosomes produced by magnetotropic bacteria. With appropriate modifications, magnetosomes linked with anti-Her2 antibodies. As a result, the magnetosomes are concentrated in tumor areas, showing a special pattern on the image.
Breast cancer is a major malignant tumor that harms women's health. According to the statistics of The American Cancer Society (ACS) in 2020, the incidence rate of breast cancer ranks the first among female tumors worldwide, and the mortality rate ranks the second, second only to lung cancer.Under the guidance of the principle of early screening and early treatment, patients should have radical mastectomy as soon as possible, otherwise the removal of the whole breast will increase the difficulty of surgery, and as time goes by, the probability of lesion metastasis rises sharply. Therefore, early detection of breast cancer is of great significance.
At present, the conventional imaging examination of breast cancer mainly includes ultrasound imaging mammography and breast magnetic resonance imaging. Compared with the other two methods, MRI has a high resolution to detect soft tissue and can perform multi-directional and multi-sequence imaging. MRI detection without ionizing radiation, however, there are still some shortcomings of conventional MRI diagnostic. Standard is not sensitive to tiny calcified points and more artifacts, and the lack of specificity target contrast agent, seriously affected the MRI detection accuracy, therefore, we plan to use synthetic biology methods to produce a specific targeted contrast agent of breast cancer, further improve the early detection of breast cancer.
An simple introduction to the protagonist of this project.
Magnetotactic bacteria are a major categories can along the direction of the magnetic field on the floorboard of the bacteria of magnetotactic movement, its characteristic is to produce a called organelles of magnetic body, this is a kind of phospholipid bilayer bag by the ferromagnetic oxide nanoparticles compared with chemical synthesis of magnetic nanoparticles, the biological source of the magnetic body with high purity and strong magnetic crystal granularity unique uniform and has good biocompatibility and genetic control.
Magnetic Resonance Imaging
MRI contrast agents usually by detectable change near the hydrogen nuclei of longitudinal (T1) or transverse relaxation time (T2) to play a role, magnetic corpuscle can by changing the T2 relaxation time for MRI imaging, on the other hand, magnetic corpuscle membrane is rich in protein. MamC is one in which content is one of the most abundant, so we plan on the MamC connection appropriate antibodies, to targeting breast cancer cells.
Single chain antibody fragment(scFv)
About scFvIt is well known that complete antibodies are Y-shaped, and the lack of complex organelles in prokaryotes makes it difficult to form spatially structured antibodies. Therefore, we chose a single chain antibody fragment (scFv) that is expressed in prokaryotes.
How toWe chose Trastuzumab as the source of single-stranded antibody, a monoclonal antibody used to treat Her2-positive breast cancer and the most widely used breast cancer treatment.We selected the variable region of trastuzumab and linked it with linker as our scFv.
Our original idea was to directly fuse MamC and scFv for expression, so that the single chain antibody fragment could be displayed on the surface of the magnetosome.
After a literature review, we found that the cytoplasm of magnetotactic bacteria is a reducing environment, which will reduce the disulfide bond of single antibody into thiol. SHuffle® is a kind of E.coli possesses mutation of gor and trxB, which result in SHuffle® can form disulfide bond correctly. Next, we need to connect the scFv expressed in SHuffle® to magnetosome, a protein interaction system, Fc – ZZ will be used.
ZZ is a synthetic analogue protein of Protein A. Protein A is a 42 kDa surface protein originally found in the cell wall of the bacteria Staphylococcus aureus and composed of five homologous Ig-binding domains that fold into a three-helix bundle. Each domain is able to bind proteins from many mammalian species, most notably IgGs. It binds the heavy chain within the Fc region of most immunoglobulins and also within the Fab region.
So we design as follow, fusion proteins of MamC and ZZ are expressed in magnetoblast bacteria, while fusion proteins of scFv and Fc are expressed in SHuffle®, and assembled in vitro to achieve the purpose of expressing antibodies on magnetosomes.
By injecting the magnetosome carrying anti-Her2-scFv into body, scFv can combine with Her2 to enrich the magnetosome near the breast cancer region, and then present obvious shadow areas in the imaging.
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