Insulin antibody production

M/sce/Zaneows-Allergic reactions. Sodium retention and edema may occur, particularly if previously poor metabolic control is improved by intensified insulin therapy. Antibody production. 

Preparation of Insulins. Until the early 1980s insulin for therapeutic purposes was produced almost exclusively by extraction from beef and pork pancreases. Between 100 and 400 mg of insulin can be obtained from each kg of pancreatic tissue, and it has been estimated that there would be sufficient supplies of animal insulin to meet the requirements of diabetic patients into the twenty-first century (2). Through modem purification procedures animal insulins can be prepared in essentially pure form, which eliminates the possibility of developing antibodies against impurities in the insulin preparations. However, patients treated with purified insulins still develop antibodies to insulin, suggesting that differences in the primary structures of these insulins might stimulate antibody production. Therefore, enzymatic and biosynthetic methods have been developed for the preparation of therapeutic insulin identical to human insulin. 

Switching from ordinary insulin to insulin lispro can reduce the production of insulin antibodies. 

Other problems that may be encountered are related to the immunologic effects of insulin use. Certain forms of insulin may evoke an immune reaction and stimulate antibody production. These anti-insulin antibodies may cause an allergic reaction in some individuals, as well as a resistance to the exogenous insulin molecule. As discussed previously, the incidence of these immunologic reactions seems to be greater when animal (i.e., pork) forms of insulin are used. Consequently, these problems are often resolved by switching the patient to another type of preparation, preferably biosynthetic human insulin. 

Where aiumal insulins are still in use, change to a highly purified pork or human insulin may be successful in reducing resistance. Responsiveness to insulin may sometimes be restored by immunosuppression, e.g. an adrenocortical steroid (prednisolone 20-40 mg/d) over weeks (or a few months), to suppress antibody production. Obviously, if this is successful, insulin dosage will have to be reduced in accordance with the impredictable reduction in cmtibodies. Patients need to be carefully monitored to avoid severe hypoglycaemia. Ketoacidosis also reduces the effect of insulin. 

Accurate measurement of proinsulin has been difhcult for several reasons the blood concentrations are low antibody production is difficult most antisera cross-react with insulin and C-peptide, which are present in much higher concentrations the assays measure intermediate cleavage forms of proinsulin and reference preparations of pure proinsulin are not readily available. However, a more sensitive nonequiUb-rium RIA method for measuring proinsiilin was developed by adsorbing the initial antiserum with biosynthetic human C-peptide coupled to agarose to eliminate cross-reactivity with C-peptide.An enzyme-linked immunosorbent assay (ELISA) has been described that employs an antibody to C-peptide as the coating antibody and antiinsulin antibody for detection. The detection limit is 0.25 pmol/L. ... 

The choice of animal is mostly one of convenience rather than necessity. A notable exception is the production of anti-insulin antibodies for which guinea pigs should be used. Generally, rabbits, goats and sheep are used for polyclonal antisera and mice and rats for monoclonal antisera. As a rule of thumb, about 15 pg of immunogen or 5 X 10 cells per kg of body weight suffices. 

Purity of insulin refers to the amount of proinsuhn and other impurities present in a given insuhn product. Prior to 1980, most insulin contained enough impurities (300 to 10,000 ppm) to cause local reactions upon injection, as well as systemic adverse effects from antibody production. Modem technology has provided less expensive techniques to purify insulin. As a result, aU insulin products contain 10 ppm of proinsulin, with purified preparations (all recombinant DNA human insulin and insulin analogs) containing < 1 ppm of proinsulin. 

The measurement of C-peptide provides a fully validated means of quantifying endogenous insulin secretion, preventing influence of exogenous insulin or insulin antibody. However, most C-peptide assay kits cannot differentiate C-peptide from proinsulin and proinsulin conversion products. The influence of proinsulin may be significant in cases where serum proinsulin is elevated, as in Type 2 diabetes, familial hyperproinsulinemia and in patients with proinsulin antibody. 

The majority of reactions observed, particularly the local ones, have been attributed to the impurities present in the injected products. However, we now know that insulin itself can be responsible for sensitization (Minars et al. 1974 Diem and Teuscher 1979). Insulin is composed of 51 amino acids divided into two chains chain A is composed of 21 amino acids and chain B of 30. It appears that the antigenic properties of insulin and the formation of anti-insulin antibodies are linked to chain A (Halpern and Bourdon 1966) but an epitope on Chain B is probably involved in cell-mediated reactions (Faulk et al. 1975 MacCuish et al. 1975). 

Jovanovic-Peterson, L., Sparks, S., Palmer, J. P., and Peterson, C. M., 1993, Jet-injected insulin is associated with decreased antibody production and postprandial glucose variability when compared with needle-injected insulin in gestational diabetic women. Diabetes Care 16 1479-1484. 

Insulin and human growth hormone, discussed above, are classic protein hormones produced with recombinant DNA technology. The immune system produces protein antibodies that attack disease causing-agents such bacteria and viruses. Genetic antibody production interferes with or attacks entities associated with diseases such as psoriatic arthritis and Crohn s disease. Recombinant DNA technology produces proteins binding with specialized white blood cells to reduce inflammation associated with rheumatoid arthritis. 

Arquilla, E. R., Finn, J. Genetic differences in antibody production to determinant groups on insulin. Science 142, 400-401 (1963). 

The highly purified insulins, such as the MC insulins, are becoming more widely used. Devlin (3 ) reported that of 42 newly diagnosed diabetic patients, treated with MC insulin for 1—4 years, only 19% had detectable antibodies as compared to 82% of 45 patients treated with mixed porcine/bovine insulin. The antibody level fluctuated during the observation period. This is in contrast to an earUer communication of Yue and Turtle (4 ) who reported a significant antibody production to MC insulin in 3 out of 7 patients. 

Bovine insulin differs from human insulin by three amino acids (alanine for threonine A-chain 8, isoleucine for valine A-chain 10, alanine for threonine B-chain 30 see Fig. 27.3). Bovine insulin does promote antibody production to some extent and doses may need to be adjusted to compensate for this. 

Porcine insulin differs from human insulin by one amino acid having alanine instead of threonine at B-chain 30 (see Fig. 27.3). The difference in structure from human insulin is not sufficient for antibody production to occur. 

Biotechnology era beginning First recombinant DNA products Human insulin Human growth hormone Interferons, etc. Monoclonal antibodies Nucleotide blockage Growth in use of natural products and neutraceuticals... 

After the approval of the first product, recombinant insulin, in 1982, progress in the development of new recombinant protein pharmaceuticals was slow ([10], Fig. 17.1). The number of biotechnology-derived drugs and vaccines approved by the US Food and Dmg Administration (FDA) has increased significantly only since 1995. More recently, sales of biologies have skyrocketed, e.g. from 900 million in 1999 to an estimated 3.5 billion in 2001 for monoclonal antibodies [11]. The annual global market for biopharmaceuticals is estimated to have increased from 12 billion US to 30 billion US in 2003 [12]. 500 candidate biopharmaceuticals are undergoing clinical evaluation and over one hundred protein-based therapeutics are in the... 

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