Among patients with type 1 diabetes mellitus (T1DM), fear of hypoglycemia is a very common problem and often leads to impaired quality of life.1, 2
Such a fear often leads to relaxation of tight glycemic control, which in turn predisposes to long-term micro- and macrovascular complications. The issue of hypoglycemia must also be dealt with in the setting of the closed-loop artificial endocrine pancreas in which insulin is delivered via the subcutaneous route.
Insulin delivered subcutaneously has a delayed action and a prolonged effect. This pharmacodynamic profile is very different from that of endogenous insulin, which is rapidly secreted by the beta cell in response to a rising glucose and rapidly delivered into the portal venous system. The advent of insulin analogs in the 1980s greatly improved the treatment of diabetes by shortening delay and duration of action. Nonetheless, there remains a large gap between commercially available insulin and the ideal insulin preparation for closed-loop control. One method of compensating for these delays during closed-loop control is the provision of a pre-meal bolus to avoid marked postprandial hyperglycemia. Another is the often-used insulin-on-board (IOB) calculation designed to avoid hypoglycemia after large amounts of insulin have been given. Though shorter-acting than regular insulin, the effect of rapid insulin analogs may persist for 8–9 h after subcutaneous delivery.3,4
Due to this prolonged effect, cessation of insulin at times of impending hypoglycemia is often insufficient to prevent overt hypo-glycemia. The shortcomings of utilizing insulin alone in a closed-loop system have led our group5,6
and a group in Boston7,8
to study the addition of glucagon, an idea first proposed by Kadish in the 1960s.9
Glucagon, a hormone normally synthesized and secreted by the pancreatic alpha cell, is the first line of defense among many counter-regulatory hormones that prevent hypoglycemia. The major site of glucagon action is the liver. Glucagon is secreted into the portal circulation, exposing the liver to levels that are two to three times higher than other organs.10
In the liver, glucagon functions primarily to raise blood glucose via glycogenolysis. In diabetes, glucagon secretion is dysregulated. Over time, people with T1DM lose the ability to secrete glucagon and epinephrine in response to hypoglycemia, contributing to the problem of severe hypoglycemia and hypoglycemic unawareness.11
The cause of the glucagon secretory defect is likely multi-factorial, relating in large part to loss of insulin production, which regulates glucagon release by a paracrine effect. Neural factors and structural islet changes likely contribute to the dysregulation.12
The drug glucagon is currently U.S. Food and Drug Administration-approved as a parenteral injection for treatment of severe hypoglycemia. Glucagon’s appeal in a bihormonal, closed-loop system stems from its favorable pharmacodynamic profile. In particular, due to its rapid absorption, the onset of glucagon action is much quicker than the offset of insulin action.13,14
For this reason, the subcutaneous delivery of glucagon can prevent hypo-glycemia more quickly and more effectively than the discontinuation of subcutaneous insulin delivery.
In our study of 13 adult subjects with T1DM, glucagon was very effective in reducing time spent in the hypo-glycemic range. When given in a brisk (front loaded or high gain) fashion, glucagon significantly reduced the time spent in the hypoglycemic range by 56% versus saline placebo (18 ± 11 vs 41 ± 13 min). In this study, both insulin and glucagon were delivered subcutaneously using a standard infusion set. The infusion rates were determined by glucose sensor values input into the fading memory proportional derivative algorithm (discussed in the next section). Glucagon prevented overt hypoglycemic events in approximately two-thirds of cases of incipient hypoglycemia. The probability of glucagon successfully preventing hypoglycemia was in part related to IOB and sensor accuracy. Higher IOB and delayed glucagon delivery (due to sensor overestimation of blood glucose) resulted in a higher probability of failure.5
Russell and colleagues7
found a similar rate of success as well as a correlation between higher insulin levels and glucagon failure. Rivera and colleagues15
made an important contribution by demonstrating that the ability of glucagon to stimulate glucose production is not only related to insulin level, but also to glucose level, with a threefold greater glucagon effect during hypoglycemia versus euglycemia.
The short-term side effects of high-dose glucagon are well known, with the most prominent being nausea and vomiting. However, the dose of glucagon delivered at any one time by a closed-loop system is quite small, approximately 50–100 mcg, compared to the 1 mg dose of a glucagon emergency kit. During chronic secretion of large amounts by tumors, glucagon is known to cause serious skin rashes, but in lower intermittent doses, it has been very well tolerated in our experience. Haymond and colleagues16
found that one to two mini-doses of glucagon were effective in preventing and treating hypoglycemia in children. However, there exists a potential risk of glycogen depletion after glucagon is given repetitively; such depletion could explain the failures to prevent hypoglycemia during closed-loop studies. Studies are underway at Oregon Health and Science University to assess liver glycogen before and after repeated doses of glucagon by the use of an ultra high- resolution technique using a 7 Tesla magnet. This magnetic resonance spectroscopy (MRS) technique is based on previous reports in which less powerful magnets were utilized.17
Two magnetic resonance spectra obtained in one normal volunteer are shown in
; the glycogen signature (dual peak centered at ~100 PPM) is substantially higher in the fed state (left panel) as compared to the fasting state (right panel).
Figure 1 Two magnetic resonance spectra of hepatic glycogen obtained in a normal, nondiabetic volunteer with the use of a Siemens Magnetom 7 Tesla instrument. Development and programming were carried out by Dr. Mark Woods and Dr. Yu Cai from the Oregon Health (more ...)
Stability of glucagon after reconstitution is also a major hurdle that must be overcome prior to its general appli- cation in a closed-loop glycemic control system. In standard commercially available systems, immediately after aqueous reconstitution at acid pH, glucagon begins to form fibrils. These fibrils, a form of beta sheet amyloid, are dependent on many factors, including glucagon concentration, heat, pH, and agitation. shows early minimal fibril formation soon after reconstitution of commercially available glucagon (left panel) and dense fibril formation after 7 days of aging the same preparation (right panel).
Figure 2 Transmission electron micrographs (TEM) of native glucagon reconstituted in sterile water. The left panel shows freshly reconstituted glucagon (1 mg/ml, Novo Gluca-Gen) and the right panel shows the same preparation after being aged for 7 days at 37 °C. (more ...)
Our group has demonstrated that the use of an alkaline preparation of native glucagon (pH 10) greatly increases the stability of glucagon, and eliminates the cytotoxicity seen with commercially available glucagon preparations that are formulated with lactose and hydrogen chloride at pH 2–3.18
Steiner and colleagues19
have addressed this issue by increasing solubility at neutral pH by using a lysolecithin surfactant, an alcohol, and a sugar to prevent glucagon self-association. A group led by DiMarchi and colleagues20
has created multiple glucagon analogs with excellent stability and retention of physiologic action.