Insulin-dependent Type 1 Diabetes Mellitus (IDDM): Carbon Nanotubes delivering Interleukin-10 to Targeted Beta-Cells

Brandon, Dharma, Andrew

Background/Description of problem

• Insulin deficiency secondary to pancreatic beta-cell destruction

o T-cell mediated autoimmune attack

• Chronic disorder that often begins early in life

• Susceptibility is largely inherited

o Depends mainly on HLA genotype

• Incidence increasing at 2-5% per year worldwide[11]

• Trend toward decreasing age at presentation

Analysis of currently available treatments

• Insulin Injections

o Most common treatment, requires daily insulin injections to regulate blood-glucose levels

• Pancreas Transplantation

o Reserved for severe cases, essentially a ‘cure’ although has many side effects associated with transplantation

• Islet Beta-Cell Transplantation

o Injecting cells through catheter, still considered to be experimental since not therapeutic

o Biomaterial microencapsulation of islet cells - must be replaced every 5-9 months, not consistently successful in large animal models

Current Immunomodulators

• Immunomodulators

o Azathioprine

Acts as an immunosuppressive, inhibiting T-cell response to antigens

Paired with glucocorticoids in study of 46 patients, only 3 patients remained in remission at one year, it should be noted equally discouraging results were produced in a second study

o Mycophenolate Mofetil

Inhibits proliferation of both T/B-lymphocytes

Paired with daclizumab, 126 patient study showed neither MMF alone or in combination with daclizumab slowed Beta-cell destruction in T1D patients

Description of proposed design

• Single-walled carbon nanotubes (SWCNTs)

• Functionalized with:

o Antibodies to beta-cell specific surface receptors

o PEG

o Active peptide fragment of IL-10

• IL-10 linked to PEG via acid-labile bond

Schematic

Anti-GPR44 antibody

IL-10

Justification of proposed design

• SWCNTs - large surface area to volume ratio for functionalization[14]

• Functionalized with:

o Antibodies - for specific targeting of β-cells

o PEG - for solubility in aqueous conditions and preventing cellular uptake

o IL-10 - promote M2 macrophage polarization[5]

• IL-10 release is self-regulatory

Synthesis & acid-hydrolysis of PEG-IL-10 conjugate[12]

1. Hydroxyl on PEG can be converted to active ester using chloroformate

2. Primary amine groups on Interleukin-10 react with the activated PEG

3. PEG-IL-10 conjugate degrades in acidic pH to release IL-10[13]

The reaction mechanism above was proposed for IL-2[12] and we assume a similar chemical reaction with IL-

10 is plausible (IL-2 and IL-10 are proteins and consist of primary amine groups).

Mathematical model• When CNTs enter the bloodstream, they circulate before getting cleared.

• The fraction of CNTs that reach the pancreatic vasculature and bind to the β-cells is assumed to be ‘f’. f can be found through fluid transport simulations and experiments, which is not dealt here.

• Once bound, IL-10 is released via acid-hydrolysis of the amide bond. This can be modeled as follows.

a. Assumption 1: pH (or [H+]) linearly related to degree of inflammation and doesn’t change/changes very slowly with time

b. Assumption 2: all CNTs present in the environment are bound (zero order)

Rate of dissociation/IL-10 release rate is given by:

(‘a’ can be determined by measuring kH at two different pHs)

Release Profile

time time

[IL

-10]

[IL-1

0]

When [H+] (inflammation) decreases with

increase in [IL-10]

When [H+] doesn’t change with time

(Assumption 1) – the slope increases with

increase in inflammation (given by [H+]) from

Assumption 2

Improvement over existing technologies

• Azathioprine is non-specific, making the individual susceptible to

infection from lowered immune response

• Much less invasive/complicated than pancreatic/islet transplants

respectively

• Lower frequency dosage compared to daily injections

References1. Yu et al. Targeted Drug Delivery in Pancreatic Cancer. Biochim Biophys Acta. 2010 January; 1805(1): 97.

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3. Lindskog et al. Novel pancreatic beta cell-specific proteins: Antibody-based proteomics for identification of new biomarker candidates. Journal of Proteomics. Volume 75, Issue 9, 17 May 2012, Pages 2611–2620

4. Lu et al. Discrete functions of M2a and M2c macrophage subsets determine their relative efficacy in treating chronic kidney disease. Kidney Int. 2013 Oct;84(4):745-55. doi: 10.1038/ki.2013.135.

5. Biomed K. Spiller class recording Winter 2013_2/11/2013

6. "Fast Facts." Data And Statistics About Diabetes. American Diabetes Association, Mar. 2013. Web. Feb. 2014.

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8. Roep et al. Antigen Targets of Type I Diabetes Autoimmunity. Cold Spring Harbor Perspectives in Medicine, 2012, vol. 2.

9. Ann Lardner. The effects of extracellular pH on immune function. Journ. of Leukocyte Biol., 2001, vol. 69 (4), 522-530.

10. Atkinson et al. Type 1 Diabetes (Seminar). Lancet, 2014, vol. 383, 69-82.

11. Daneman, Denis. Type 1 Diabetes. The Lancet. Volume 367, Issue 9513, 11–17 March 2006, Pages 847–858

12. Aldwin, L., Goodson, R., Katre, N., Nitecki, D. E. “Preparation of a polymer/interleukin-2 conjugate”. US4902502A. Feb 20, 1990.

13. Electron-pushing mechanisms: <http://www.chem.wisc.edu/areas/reich/handouts/elecpush/ep-mechanisms.htm>

14. Madani, S. Y., Naderi, N., Dissanayake, O., Tan, A., Seifalian, A. M..A new era of cancer treatment:: carbon nanotubes asdrug delivery tools. Int J Nanomedicine. 2011; 6: 2963–2979.

Q&A session