BY

G.MUTHULAKSHMI
M.Tech Biopharmaceutical technology
INTRODUCTION
 Type 1 diabetes mellites is a chronic disease results from
severe or absolute insulin deficiency that is pancreatic
cells fails to produce insulin.

 Two main causes:

1. Immune mediated- most prevalent
Destruction of -cells of pancreas by T-cells.
2. Idiopathic
No known cause and autoimmunity but patients
cannot produce insulin
FACTORS CONTRIBUTE TO TYPE I DIABETES
MELLITUS
Genetic factors: specific HLA allele type (HLA-
DR3/4, DQ2/8)
Viral infection: congenital rubella infection increase
the risk of type I diabetes mellitus
Environmental factors: Increased free radicals and
toxic doses of nitrosamines
From mother to young:
Onset of type I diabetes mellitus is seen in young
before 5yrs of age due to transfer of auto insulin
antibodies.
PANCREAS
Three types of cells
 cell - Glucagon,
Proglucagon
 cell – Insulin, C-
peptide, Proinsulin,
amylin
δ cell – Somatostatin
F cell – Pancreatic
polypeptide
 INSULIN
Insulin is a polypeptide hormone
formed, after elimination of C peptide
by hydrolysis. Its molecular weight is
58KDa consist of two chains of 21 and
30 amino acids, connected by two
disulfide bridges. Belongs to the group
of peptides called IGF (insulin like
growth factors) or somatomedins.
Insulin concentration in blood is 10-8
to 10-11 M
Exist as a monomer during biological
action
Exist as dimer in aqueous and neutral
solution and in the presence of zinc
 INSULIN SECRETION
 Secretion is biphasic
immediate effect of short
duration and sustained
effect
 Pancreatic cells releases
insulin into portal venous
system
 Basal insulin alues of 5-
15μU/ml (30-90 pmol/L)
are found in normal
humans, with a peak rise
to 60-90 μU/ml (360-540
pmol/L) during meals
 REGULATION OF INSULIN RELEASE
Nutrient secretagogues: Increase level of glucose in blood

 Non-nutrient secretagogues:
Neural stimuli - ‘cephalic phase’ of secretion. Stimulated
by vagus nerve, cholinergic and muscarinic receptors in 
cells activate phospholipase C with subsequent
intracellular events activating protein kinase A ,triggering
calcium release and insulin secretion.

Adrenergic pathway - Catecholamines inhibit insulin

release through 2-adreno receptors and stimulate insulin
synthesis by - adrenoreceptors.
Contd..
Peptide hormones
Somatostatin – inhibit insulin secretion
GLP-1 –combined with glucose it enhances proinsulin
biosynthesis. The mechanism of action is mediated by C-
AMP pathway.

 Amino acids
Arginine – cationic amino acid activates insulin
secretion by depolarizing the cell.
Leucine + glutamine- enhances insulin secretion.
Regulation of insulin release
EFFECT NUTRIENTS HORMONE NEURAL

Stimulatory Glucose GLP-1 β adregenic

Aminoacids Glucagon pathway
Growth hormone Vagal
GIP (para
Cholecystokinin sympathetic)
Gastrin
VIP
Gastrin releasing peptide
Inhibitory Adrenocorticosteroids α adrenergic
Somatostatin
Adrenalin
Noradrenalin
Galanin
Neuropeptide Y
Calcitonin gene-related
peptide (CGRP)
Prostaglandin E
Long term influences on insulin
secretion
Insulin secretion decreases during
Neonatal maturation
Exercise
Starvation
Aging
` Growth hormone deficiency
Excess thyroid hormone
Insulin secretion increases during pregnancy
 INSULIN RECEPTOR
Insulin exerts its action on various
tissues by binding with the receptor
present on the membrane surface
Insulin receptor consists of two
covalently linked heterodimers.
α subunit – extracellular and
consitutes recognition site
β subunit – spans the membrane,
contains tyrosine kinase
 ACTION OF INSULIN
Insulin action at cellular
level encompasses
carbohydrate, lipids,
aminoacid metabolism
and mRNA transcription
and translation.
 GLUCOSE TRANSPORTERS
TRANSPOR TISSUES GLUCOSE FUNCTION
TER Km
(mmol/L)
GLUT -1 All tissues, especially 1-2 Basal uptake of glucose,
red blood cells and brain transport across the blood
brain barrier.

GLUT -2 B cells of pancreas, Liver, 15-20 Regulation of insulin

kidney, gut release, other aspects of
glucose homeostasis
GLUT -3 Brain, Kidney, placenta, <1 Uptake into neurons,
other tissues other tissues

GLUT -4 Muscle, adipose tissue, 2.5 - 5 Insulin mediated uptake

heart of glucose
GLUT -5 Gut, Kidney 1-2 absorption of fructose
 PHYSIOLOGICAL ACTION OF INSULIN
MUSCLE: Insulin uptake is through GLUT-4 and muscle
accounts for 60%- 70% whole body insulin mediated
uptake.
Effects:
Increases glucose transport
Increased glycogen synthesis
Increased protein synthesis
Increased aminoacid synthesis
Supression of proteolysis
In case of insulin deficiency ,
there is an increase in protein break down leads
to increase level of nitrogen in the urine (honey-
urine-disease).
imbalance in protein turnover and protein
balance leads to muscle wastage.
decreased synthesis of intestinal mucosal
proteins results in gastrointestinal complications.
 ACTION ON LIVER
While glucose uptake in liver is not insulin- dependent, it
accounts for about 30% of total body insuin mediated
glucose disposal.
EFFECTS:
Promotes glucose storage as glycogen
Increases triglyceride synthesis and VLDL formation
Inhibits conversion of fatty acids and amino acids to keto
acids
inhibits glycogenolysis and gluconeogenesis
inhibits conversion of amino acids to glucose
 ACTION ON ADIPOSE TISSUE
Insulin uptake is through GLUT-4 and constitutes 10% of
total body insulin mediated glucose uptake

Effects: Increased triglyceride synthesis

Lipoprotein lipase is induced and activated by
insulin to hydrolyze triglycerides from lipoproteins
Glucose transport into cell provides glycerol
phosphate to esterification of fattyacids
 ACTION ON BRAIN
While the brain is not insulin-dependent, insulin
receptors are concentrated in the olfactory bulb,
hypothalamus, hippocampus,40 retina and vessels of the
choroid plexus, as well as in regions of the striatum and
cerebral cortex. Insulin is believed to act as a
neuropeptide, involved in satiety, appetite regulation,
olfaction, memory and cognition.

There is suggestion of a potential link to Alzheimer’s

disease.
CONTD..
PITUITARY: Insulin receptors present in anterior pituitary
gland stimulates production of growth hormone and in
turn stimulates IGF-1 production by liver

KIDNEY: The kidney does not require insulin for glucose

transport.Insulin receptors are found in the proximal
tubules; insulin regulates mineral transport and
gluconeogenesis in the kidney.Insulin is degraded in the
kidney.
 TYPE I DIABETES MELLITUS
In Normal individuals:
 Glucose concentration is in the range of 3.5-7mM throughout the day.
 Fasting or pre-prandial insulin concentration is 6.1mM
 2-hr Postprandial glucose level peaks within 30-60 min after meal and
does not exceed 7.8mM.
 HbA1c level in the range of 4%-6%

In treatment regimens for type I diabetes patients,

 Blood glucose concentration of 5.0-7.2 mM, and 6.1-8.3 mM at bedtime is
maintained.
 HbA1c level in the range less than 6.5% is maintained

In treatment regimens for type I diabetes patients,

TREATMENT OF DIABETES MELLITUS
The goal of therapy is
(a) elimination of ketosis;
(b) elimination of symptoms of
hyperglycemia such as polydipsia, polyuria, vaginitis,
fatigue, and visual blurring;
(c) restoration of normal blood chemistry
values;
(d) regaining of lost weight; and
(e) restoration of sense of well-being.
 COMPLICATIONS IN HYPERGLYCEMIA
Acute complications:
Thirst, polyuria, glucosuria, hunger, hyperosmolar
hyperglycemic nonketotic coma, an often fatal result of
osmotic diuresis and dehydration. Ketoacidosis due to
excessive lipolysis

Late complications:
microvascular-retinopathy, nephropathy and neuropathy,
and macrovascular-ischemic heart disease ,peripheral
vascular disease, and stroke.
INSULIN THERAPY
Insulin therapy is the only efficacious treatment for type I
diabetes mellites.
Ideally, insulin therapy should imitate the normal level of
insulin secretion.
In people who do not have diabetes, the release of insulin
follows 2 patterns:
A baseline secretion of insulin and intermittent pulses of insulin
release following each meal .Administration of exogenous
insulin in a basal-bolus regimen attempts to mimic the natural
release of insulin through multiple daily injections.
Contd..
Earlier, bovine and porcine insulin was used for
treatment of type I diabetes mellites

Production of human insulin

À Semisynthetic method: Porcine insulin is changed
enzymatically Ex: Velosulin, Hoechst insulin

Á Biosynthetical DNA-technology method

Production from baker’s yeast Novo-Nordisk insulin

Production from Ecoli-bacteria Eli-Lilly insulin

 STRUCTURE ACTIVITY RELATIONSHIP
 High binding affinity to insulin receptor is accomplished
by amino acids at positions
A chain: A1, A2, A3, A16, A19, A21, A13
B chain: B6, B12, B15, B23, B24, B25, B17
 B8, B9, B12, B13, B16, and B23- B28 are amino acid
positions required for dimer formation
 Hexamer formation is stabilised through association of six
His B10 with two zinc atoms
 Subsitutions at B26- B30 maintains high binding affinity.
This region is not critical for binding of insulin receptor
but binding affinity in IGF-1 receptor is altered.
Contd..
Aspartic acid subsituted at His B10 which is essential
for hexamer formation, is absorbed twice as rapidly as
regular insulin
3.5 fold increasing affinity for insulin receptor
compare to regular insulin
About twice as potent than regular insulin in
metabolic assays.
TYPES OF INSULIN
Insulin, classified based on the action profile as
Rapid/Short-acting insulin
Used for bolus (meal time) needs
Used for elevated BG
 Intermediate/Long-acting insulin
Used for basal (between meal) needs
Not intended to cover meals
Dosage unit
 One international unit of insulin (1 IU) is defined as the biological
equivalent of about 45.5 μg pure crystalline insulin (1/22 mg exactly).
This corresponds to the old USP insulin unit, where one unit (U) of
insulin was (arbitrarily) set equal to the amount required to reduce
the concentration of blood glucose in a fasting rabbit to 45 mg/dl (2.5
mmol/L).
REGULAR INSULIN
Onset of action Peak duration
30-60min 2-4hr 6-10hr
 It is a man made human insulin
structurally identical to that of human
insulin secreted by pancreas.

 Normal insulin concentration peaks at

30-45 min and returns to basal level
after 2-3 hr.

 Does not mimic endogenous insulin

level due to delayed absorption from
subcutaneous injection site

 Disparity in plasma glucose levels create

brief hyperglycemic period immediately
after meal and post hypoglycemic period
3-4 hr after a meal. Ex:Novolin®
RAPID ACTING INSULIN
Onset of action Peak Duration
10-15 min 1hr 3-5 hr
These are insulin analogs
of human insulin
Types: Insulin lispro
(Humalog ®), Insulin
aspart (Novolog ®), Insulin
glulisine (Apidra ®)
Best mimics normal
insulin secretion during
meals.
 INSULIN LISPRO(HUMALOG®)
First rapid acting analogue produced by Eli lilly
company. Its M.W is 5808.
Lysine in B29 and proline in B28 position is reversed
in insuline lispro . It does not alter insulin receptor
binding affinity.
Does not self associate because of changes in terminal
portion of B chain. Thus it dissociates rapidly from
SC injection site
It has high affinity towards IGF-1 receptor compared
with human insulin
INSULIN LISPRO
Pharmacokinetic properties
Absorption: Rapid absorption from injection site.
Absorption is more quickly from intenstinal site than
from deltoid or femoral sites. Heat increases
absorption.
Distribution: volume of distribution is 0.26-0.36L/Kg
Bioavailablity: 55%-77%
Elimination: dose dependent half life is 26-52 min.
Pharmacodynamic properties
Insulin lispro reach the maximum concentration
within 38 min compared to 101 min for human insulin
Mean residence time that is average time a drug
remains in the body for insulin lispro is half the time
of regular insulin.
Lower post prandial rise in serum glucose level and
lower occurrence of hypoglycemia due to its short
duration of action.
INSULIN ASPART ( NOVOLOG®)
Proline at B28 position is
replaced by aspartic acid
Introduction of charged
group prevents the
formation of dimers and
hexamers.
Affinity towards insulin
receptors and IGF-1
receptor is similar to that
of human insulin
 Pharmacokinetic properties
Absorption: choice of injection affects action profile.
Abdominal administration decreases plasma glucose
level in short duration compared to deltoid or thigh
muscle . Faster rate of absorption and narrow peak
Distribution: Low plasma protein binding <10%
Elimination: apparent half life is 81 min
Pharmacodynamic properties
Peak serum concentration reaches approximately
twice as that of human insulin
Mean residence time is 40-50 min
Occurrence of post prandial rise in glucose level after
4 hr of administration
Increase occurrence of hypoglycemia in night time.
INSULIN GLULISINE(APIDRA®)
Substitution of
asparagine by lysine in
B3 and Lysine at position
B29 by glutamine.
Isoelectric point shifted
from 5.1 to 5.5 increases
solublity .
Increased affinity
towards IGF-1 receptor
thus there is a risk of
tumorogenecity.
Pharmacokinetic properties
Absorption: Faster absorption due to its high
solublity
Bioavailablity: 70%
Elimination: 13-17min
Pharmacodynamic properties
Peak concentration occur within 6omin
Lower occurrence of hypoglycemia, nocturnal
hypoglycemia.
INTERMEDIATE ACTING INSULIN
Onset of Peak Duration
action
1-2 hr 4-14 hr 10-16 hr

•NPH insulin is an
intermediate acting insulin
wherein absorption
and onset of action is
delayed by appropriate
amount insulin and
protomine so that neither is
present is uncomplexed form
•Protamine is a mixture of
six major and some minor
compounds isolated from
the sperm of rainbow trout
Pharmacokinetic properties
Absoption: After subcutaneous injection, proteolytic
tissue enzymes degrade the protamine to permit
absorption of insulin.
It has onset of action 2-5 hr and duration of 4-12 hr
It cannot be injected intravenously and intravenously
It can be mixed with regular insulin.
LONG ACTING INSULIN
Onset of Peak Duration
action
1 -2 hr flat 23-24 hr

Mimics the basal insulin

secretion level
Given once in a day or
twice
INSULIN DETEMIR (LEVEMIR®)
It is an acylated
derivative of human
insulin
It binds to albumin
through fatty acid chain
attached to lysine at
residue at B29
Lower affinity for
insulin receptor
necessiates higher
doses.
Given twice daily for
smooth basal insulin
level
Pharmacokinetic properties
Absorption: Less variablity in absorption than NPH.
Maximum serum concentration reached between 6-8 hr
after administration. Slower and prolonged absorption is
seen
Distribution: 98% bound to albumin
Elimination: terminal half life is 5-7hr depend on dose.
renal impairment decreases elimination.
Pharmacodynamic properties
Flat action profile with duration 23hr at the highest dose
and 5.7hr at the lowest dose.
INSULINE GLARGINE(LANTUS®)
Subsitution of glycine for
asparagine at position A21
and addition of 2 arginine
rings at B30.
Isoelectric pH shifts from
5.4 to 6.7 makes it more
soluble at acidic solution
and less soluble at
physiological pH.
Increase in affinity towards
IGF-1 receptor
Pharmacokinetic properties
Absorption and Bioavailability: After subcutaneous
injection of insulin glargine , the insulin serum
concentrations indicated a slower, more prolonged
absorption and a relatively constant concentration/time
profile over 24 hours with no pronounced peak in
comparison to NPH insulin.
Metabolism: Insulin glargine is partly metabolized at the
carboxyl terminus of the B chain in the subcutaneous
depot to form two active metabolites with in vitro activity
similar to that of insulin, M1 (21A-Gly-insulin) and M2
(21A-Gly-des-30B-Thr-insulin). Unchanged drug and these
degradation products are also present in the circulation
Mixed insulin(Humulin®70/30)
Humulin 70/30 is a mixture of 70% Human Insulin
Isophane Suspension and 30% Human Insulin Injection
(rDNA origin). Itis an intermediate-acting insulin
combined with the more rapid onset of action of Regular
human insulin. The duration of activity may last up to 24
hours following injection.
As with all insulin preparations, the duration of action of
Humulin 70/30 is dependent on dose, site of injection,
blood supply, temperature, and physical activity
Insulin preparations
Modes of Administration
Subcutaneous administration:
Insulin is usually taken as subcutaneous
injections by single-use syringes with
needles, an insulin pump, or by
repeated-use insulin pens with needles.
Administration schedules often attempt
to mimic the physiologic secretion of
insulin by the pancreas. Hence, both a
long-acting insulin and a short-acting
insulin are typically used
Standard mode of subcutaneous
injection is by using disposable needles
and syringes.
Insulin pens
To facilitate multiple sub
cutaneous injections of
insulin.
Contain cartridges of
insulin and replaceable
needles.
Pen-like device with
cartridges holding 150-300
units of insulin
 Reusable or Disposable
 Dial insulin dosage and
inject (like a syringe)
INSULIN PUMPS
Insulin pumps are used to create an artificial insulin
secretion subcutaneously that can suppress excessive
blood glucose.
In general, it continuously delivers a basal rate of
exogenous insulin automatically and also has
capability of manually adding extra insulin to the
basal rate for the intensive insulin secretion by a
continuous subcutaneous insulin infusion (CSII)
technique.
Contd..
Most of the insulin pumps
consist of a small processing
module with a display, a
disposable insulin reservoir,
and an insulin syringe, and
are powered by batteries.
Advantages :
Better control over
background or 'basal' insulin
dosage.
Limitations: Cost, catheter
problems, and no "closed
loop" means of controlling
insulin delivery based on
current blood glucose levels.
Inhaled insulin
FDA has approved an inhaled insulin preparation of
finely powdered form and aerosolized human insulin

Insulin has been readily absorbed through alveolar

walls but the molecule should be small.
It has a rapid onset of action and peak action similar
to rapid acting insulins and duration of action similar
to regular insulin
Exubera , powder form of inhaled insulin was
introduced in Jan-06 by Pfizer and Nektar
therapeutics
Bio availablity is only 10% to 40%
It is discontinued due to patients uncompliance.
AIR insulin system introduced by Eli lilly in a dry
powder form was also discontinued due to its high
particle size of 5 to 50μm.
Buccal insulin
Drug delivery through the buccal mucosa has
following advantages,
Direct contact with mucosa
relatively large surface for absorption (100–200 cm²);
level of vascularization is very high in some areas;
weak variations of pH (≠GI);
buccal enzymatic activity is mainly intracellular and
less developed than in other mucosae.
Drawbacks
The buccal mucosa is not an absorptive organ.
Sublingual area is thin and nonkeratinized, highly
permeable (high drug input);
Cheek mucosa is thicker and nonkeratinized, fairly
permeable (low but sustained drug input); and
In summary, the continuous, but variable, saliva flow
and the robust multilayered structure of the oral
epithelium constitute an effective barrier to
penetration of drugs.
 oral-lyn
Generex US-based company develop oral-lyn
Oral-lyn is a liquid formulation of human regular
insulin with a spray propellant for prandial insulin
therapy.
The formulation results in an aerosol with relatively
large micelles (85% of that having a mean size >10
μm) and therefore cannot go into the lungs.
On administration with Rapid Mist device insulin
passes through superficial layers of mucosa into the
blood stream
Each 28 mlcanister contains 400 U of regular human
insulin.
The drug is under clinical trials
Oral insulin
Advantages:
Increased patients compliance
Direct absorption from the gut.
 Transfer of insulin directly to liver
 Control hepatic glucose production to the same
extent of natural insulin.
This more “physiological insulin delivery” would be
associated with reduced peripheral hyperinsulinemia
obstacles:
Enzymatic degradation in the gut.
Insulin has to pass through the high viscous mucous
layer of the gut
Disadvantages:
50% of insulin is degraded by liver thus peripheral
insulin concentration is low.
Biocon has taken over the oral insulin technology
developed by Nobex. IN-105 is a human insulin molecule
conjugated on position B29 with polyethylene glycol via an
acyl chain
IN-105 is declared to have the following characteristics:
improved half-life in the digestive tract and improved
absorption,
lower immunogenicity as compared to insulin,
lower mitogenicity as compared to insulin,
retains a similar pharmacological activity as insulin, and
conserves safety profile and good clearance profile as
compared to insulin. It is under phase III clinical trials
Adverse reactions
Hypoglycemia: Most common complication of insulin
therapy. They may result from delay in taking a meal,
inadequate carbohydrate intake, unusual physical
exertion or dose of insulin is too large.
Symptoms are palpitations, sweating, nausea, hunger.
In persons with persistent hypoglycemia
manifestations of insulin excess may develop.
Includes confusion, weakness, bizarre behavior, coma.
Mainfestations of hypoglycemia are relieved by
glucose administration.
Contd..
Lipodystrophy: It occurs in the subcutaneous fatty
tissue if injected repeatedly at the same site
Weight gain
Immune allergy: immediate type hypersensitivity
results from histamine release from mast cells
sensitized by anti-insulin IgE antibodies.
Immune insulin resistance: IgG anti-insulin
antibodies neutralizes the action of insulin
 Drug interactions
Drug increases glucose lowering effects of insulin
pramlintide, ACE inhibitors, disopyramide, fibrates,
fluoxetime, propoxyphene, pentoxyfyline, salicylates,

Drug decreases glucose lowering effects

corticosteroids, niacin, danazol, diuretics, isoniazid,
phenothiazin, somatropin, thyroid h0rmone, estrogen,
protease inhibitors, atypical antipsycotics.
 DIABETES EXPOSURE
Although conventional hypoglycemic therapies prevent
acute metabolic complications in patients with diabetes,
they do not restore metabolic homeostasis.
 The result of this imperfect treatment is a novel milieu
that includes various combinations of metabolic,
hormonal, and physiologic alterations.
These include hyperinsulinemia, hyperglycemia,
hyperlipidemia, abnormalities in blood flow, and the
formation of glycation products all of which constitute
diabetes exposure.
As a consequence of this exposure, diverse functional and
morphologic alterations develop that lead to severe
complications affecting the eyes, kidneys, and heart.
 DIABETIC RETINOPATHY
Structural changes:
thickening of the capillary basement membrane,
loss of retinal pericytes,
increased vessel permeability
and formation of capillary microaneurysms.
These structural changes are accompanied by
decreased retinal blood flow, capillary occlusion,
angiogenesis, hemorrhage, fibrotic tissue formation,
and tractional retinal detachment. Some of these
events, or all of them in combination, can ultimately
result in impairment or complete loss of vision
Incidence, progression and development of
proliferative diabetic retinopathy is highest in type I
diabetes patients compared to type II diabetes
patients
High glycemic control reduces the onset of disease
Hypertension, Higher ocular perfusion pressure and
myopia increases the risk of progression
DIABETIC NEUROPATHY
Diabetic neuropathy is common in diabetic patients,
with a prevalence of more than 50%
 The pathogenesis is multifactorial and considered to
be both hyperglycemia-induced pathologic changes
intrinsic to neurons and ischemia-induced neuronal
damage by decreased neurovascular blood flow .
 Histologically, increases in endothelial cell area and
luminar narrowing of capillaries are present in the
endoneurium of patients with diabetes.
DIABETIC NEPHROPATHY
Diabetic nephropathy is characterized by
thickening of the glomerular basement membrane,
increased fractional mesangial volume,
 Expansion of the glomerular mesangium, which
occurs at the expense of the glomerular capillary
lumen and filtration surface area, correlates most
closely with the decline in renal function and the
development of proteinuria