novel monoclonal antibody for a novel target (cancer therapy)

 

                                     

 

 

 

 

 

 

 

 

 

                                      Development of a novel monoclonal antibody

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Introduction

Cells differentiate and proliferate in multicellular organisms. These cells either survive in certain organs or die after their lifetime in the body. The cells that do not survive result in the activation that cause apoptosis of the cell. The inception of molecularly targeted treatments has lead to the diversification of treatment of colorectal cancer. This is with integrations of novel monoclonal antibodies with cetuximab and bevacizumab. The ever increasing development of agents for the treatment of the cancer gives much hope in the clinical treatment. The integration into standard and meaningful clinical treatments has resulted to better results in treatment (Zouhairi, Aline, & Michael, 2011). In this paper, I will discuss the development of monoclonal antibodies for therapy of metastatic colorectal cancer.

Development of monoclonal antibodies

Tumors have been studied to show the need for angiogenesis, a process that recruits blood vessels. This does not occur in healthy people except in remodeling of tissues and wound repair. Angiogenesis calls for improved vessel permeability, cell proliferation and migration causing long capillaries. These process results to vessels that are highly branching and leaky. The circulation at the micro level is inefficiently causing the place to be acidotic thus creating pressure to the stroma. The control of this process is thus inevitable in the fight against the cancer. The control has mainly been studied through the use of the growth factors of the vascular endothelial VEGFs and the receptors VEGFRs (Zouhairi, Aline, & Michael, 2011).

The development of these antibodies through from the production to purification method takes approximately five months. There are five phases that have exit points if results are not met. Phase o entails the preparation of the protein for the antibody production. This protein is expressed in either mammalian cells or E.coli using bacterial vectors (Adams & Weiner, 2005). In phase 2, the mice needed are between five and ten are test bled and immunized for 5 weeks. This is followed with a test bled with Elisa. The composition of the IgM and IgG are determined through screening.

The choice of the two or three best bled animals are med out of the mouse were used. These animals that have responded well, their spleen cells, are harvested for hybridoma fusion. The Elisa test is then used to screen the positive supernatants from the cells to make 2ml, then preservation of the cell lines as frozen. In the third phase is made through the subcloning in the limitation of dilution to attain the cell lines. There are two versions of cloning that is through a single round and multiple rounds of dilution at the very same concentration to have the same isotype.

The production of monoclonal antibodies can be either by cell or in mouse abdominal culture to produce ascites containing antibodies. The culture in mouse will effect to up to 10mg in 3ml of ascites’ fluid. The monoclonal antibody is finally extracted through the purification from the ascites. Purification is achieved through protein G/A resin, ion exchange or antibodies that are of immunoglobulin. Thawing of the cell lines occurs after which the viability of the cells are tested through T-25 and their expression put onto test via isotype matching (Lavande, Itekar, & Agnihotri, 2011). The viability tested through T-25 is affirmed through cell isostrip and density irrespectively.

In a mouse, once the hybridoma viability has been assessed the immunization of a number of cells primarily takes effect. The optimization of cells and mouse strain also occurs in relation to the immunization. After the success in the infection process, the ascites’ fluid is collected to produce an approximate of between 3-10mg from each of the mouse. The purification of the antibody is performed under salt concentrations that are considerable. This allows for better and stringent binding to the antigen including the isotype.

 

The mechanism of action

The most predominant form of VEGF is VEGF-A, which is a common mediator of angiogenesis. They are growth factors that are produced by stromal cells. They act as attachment sites for extracellular domains of their receptors. Intracellular domains in these receptors contain tyrosine kinases that are catalytic. The binding of the endothelial growth factors results to the initiation of cascades that ensure cell survival. A monoclonal antibody, Bevacizumab, is targeted to VEGF-A to inhibit the process of angiogenesis (Zouhairi, Aline, & Michael, 2011).

The use of the antibody only produces partial results, but in conjunction with cytotoxic chemotherapy leads to suppression of the tumor. The combination induces apoptosis and normalizes the architecture of blood vessels. This in effect causes the reduction of the hydrostatic pressure thus enabling the release correctional cancer agents to the tumor.

The combination of IFL regime with bevacizumab in patience that had previously untreated tumors showed high levels of response and free survival progression. In comparison of 32 months to 20 months in patients who use the antibody progressively. Those who progressively used the antibody had a high OS count. The use of bevacizumab has got side effects that come with it. It results to myocardial infarctions and perforation of gastro intestines. Hypertension can also result in about 20% of the patients.  

Bispecific antibodies

 

The antibodies can recognize two antigens. They stimulate the cells responsible for cytotoxic effects of tumors. The antibodies will directly result to the binding to a specific cell resulting to immunity. The specificity of the antibodies can be altered by changing the specificities of the anti-target and anti-effectors components. The specificity of the antibody is directly to the target cell during the production (Lavande, Itekar, & Agnihotri, 2011). The mAbs in cancer therapy are directed against certain cell surface receptors and begin a physiological function. Bio-mimetic antibodies are the name for these, which are used as for effects to modify a biological response.

Screening

The achievement of the knowledge of the role of VEGF as a factor in antibody therapy of colorectal cancer is important. This is done through its activation using the novel monoclonal antibody targeted on its receptor. Research is done on patients with tumors with bevacizumab directly as agents of disease.

The standard monoclonal antibody development includes the Elisa test. This is determined the cells to use for the production of the antibodies. Elisa matched pair is one of the methods used for screening of the antibodies. In the case of needed immunoassay, suitably matched pairs are developed. More than 50 cytokine and other Elisa products are used. Biacore is used to screen the supernatant accrued from the positive fusions. This is by injection of the supernatant at a concentration of about 0.25mg/ml, over the IgGFc surface and subsequent addition of recombinant antigen.

Qualification taste is made through the antibodies being tested against the standard provided cell lines. These cell lines have a stipulated preparatory procedural condition for formalin fixation (Maine biotechnology services, 2013). Simple western blotting the cell lines qualification is affirmed, through the antibody reactivity pattern as well as the cells under standard conditions of preparation.

Specific examination

This occurs through an advanced method of western blotting. These samples are enhanced for locations with cellular specificity before the phoresis. The process accords for as many as five samples to be used in testing for the specific monoclonal antibody. This method is significant for the low concentration of the antibody (Booy, Johar, & Maddika, 2006). The cell lines are used in the direct targeting of tumors through their receptor factors. The antibodies can only be used for a specific cell thus increasing its specificity.

Clinical trials

A study done on two trials, 20 patients suffering from colorectal cancer at different levels of the disease are used. Phase II clinical trials the antibody in combination with irinotecan demonstrates an increase in RR of up to 77%. In relation to this use of bevacizumab with chemotherapy showed a bad outcome in patients who previously were on other treatments. It is still not very clear on the essence of VEGF in colorectal cancer treatment, but studies are being done to address this.

The role of bevacizumab in the management of this cancer is done in 2 phase III trials. The PETACC-8 and NCCTG, these are clinical studies of handling of the tumor in EXPERT-C trial, in patients. The VEGF monoclonal antibodies have well been absorbed, but they have resulted in the development of rash in most patients. The rates off RR and OS significantly increased with an advent rise in toxicity levels in the skin. The development of rash is a true show of efficacy in the patients (Zouhairi, Aline, & Michael, 2011).

Conclusion

The use of monoclonal antibodies in inducing proliferation of tumor cells, as well as its apoptosis, is rapidly developing. The remarkably high affinity of the monoclonal antibodies for their intended targets is the basis of action against cancer. The future clinical use of these immunotherapeutic presents rather powerful approach to the treatment of cancer. Not only are the antibodies used in therapy, but also on a daily basis to identify and detect the presence of tumor antigens. This means that they are of so much significance to any biological laboratory.

 

 

References

Adams, G. P., & Weiner, M. L. (2005). Nature biotechnology. Monoclonal antibody therapy of cancer, 1147-1157.

Booy, E. P., Johar, D., & Maddika, S. (2006). Therapeutic antibodies. Monoclonal and bispecific antibodies, 85-101.

Lavande, J. P., Itekar, P. B., & Agnihotri, A. A. (2011). American journal of pharmtech research. Novel Monoclonal Antibodies for Cancer Treatment.

Maine biotechnology services. (2013). Retrieved 2014, from Monoclonal Antibody Development: http://www.mainebiotechnology.com/monoclonal/

Seimetz, D. (2011). Journal of Cancer. Novel Monoclonal Antibodies for Cancer Treatment: The Trifunctional An-tibody Catumaxomab (Removab®), 309-316.

Zouhairi, M. E., Aline, C., & Michael, P. j. (2011). Retrieved 2014, from Molecularly Targeted Therapy for Metastatic Colon Cancer: Proven Treatments and Promising New Agents: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3070284/

 

 

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