Persistent Cutaneous Insulin Allergy Resulting From High-Molecular-Weight Insulin Aggregates

Cutaneous insulin allergy remains a clinical problem despite the use of highly purified human insulins. We used in vitro lymphocyte-transformation studies to examine the reactivity of various insulin formulations in diabetic patients with (n = 4) and without (n = 8) cutaneous allergies. Nonspecific response to concanavalin A demonstrated a >40-fold response in both groups. Control patients did not respond to the addition of commercial insulin preparations (stimulation index [SI] <4), whereas allergic patients had an 11-fold response to beef (P < 0.01), a 10-fold response to pork (P < 0.01), and a 6-fold response to human (P < 0.01) insulins. This response was limited to a single insulin manufacturer's preparations and was uniform in all three species tested. Efforts to identify the offending agent revealed no lymphoblast transformation when crystalline insulin was used or when commercial preparations were purified to a single peak by high-performance liquid chromatography (HPLC). Pure crystalline insulin dimers of beef, pork, and human species were tested; control subjects responded with mean Sis of 1.9,1.9, and 1.8, respectively, whereas allergic patients showed greater reactivity to beef (SI 7.3) and pork (S114.8). The lymphoblast-transformation response to crystalline human dimer was dose dependent with mean Sis of 0.9 at low concentration (2.8 ng/ml) and 19.2 at a higher concentration (20.4 ng/ml). The commercial insulin preparations were run on size-exclusion HPLC to determine high-molecular-weight aggregate content. Independent of species, a single manufacturer had products demonstrating aggregate levels 3- to 6-fold higher than those found in other manufacturers’ preparations. Our in vitro data suggest that insulin aggregates rather than native monomers cause cutaneous allergy and that antigenicity of commercial preparations correlates with the relative concentration of aggregates. In vivo testing is needed to confirm our results.