PHARMACEUTICAL COMBINATION FOR USE IN THE TREATMENT AND/OR PREVENTION OF DIABETES

Abstract

The present invention relates to pharmaceutical combination for use in the treatment and/or prevention of diabetes, comprising a calcium channel blocking agent and an incretin.

Claims

1. A pharmaceutical combination for use in the treatment and/or prevention of diabetes, comprising: a) A calcium channel blocking agent or a pharmaceutically acceptable salt thereof, b) An incretin mimetic and/or an incretin enhancer or a pharmaceutically acceptable salt thereof.

2. A pharmaceutical combination of claim 1 for use in the treatment and/or prevention of Type-II diabetes

3. A pharmaceutical combination of claim 1 or 2 for use in the treatment and/or prevention of Type-II diabetes in patients not suffering from prehypertension and/or hypertension.

4. A pharmaceutical combination for increasing glucose-stimulated insulin secretion (GSIS) in a patient, comprising a) A calcium channel blocking agent or a pharmaceutically acceptable salt thereof, b) An incretin or a pharmaceutically acceptable salt thereof.

5. A pharmaceutical combination according to any of the claims 1 to 4, whereby the calcium-channel-blocking agent is selected from the group comprising dihydropyridines, phenylalkylamines, benzothiazepines or mixtures thereof.

6. A pharmaceutical combination according to any of the claims 1 to 5, whereby the calcium channel blocking agent is selected from the group comprising amlodipine, aranidipine, azelnidipine, barnidipine, benidipine, cilnidipine, clevidipine, cronidipine, darodipine, dexniguldipine, efonidipine, elgodipine, elnadipine, felodipine, flordipine, furnidipine, iganidipine, isradipine, lacidipine, lemildipine, lercanidipine, levamlodipine, levniguldipine, manidipine, nicardipine, nifedipine, niguldipine, niludipine, nilvadipine, nimodipine, nisoldipine, nitrendipine, olradipine, oxodipine, palonidipine, pranidipine, ryodipine, sagandipine, sornidipine, teludipine, tiamdipine, trombodipine, bepridil, devapamil, fendiline, gallopamil, verapamil, diltiazem, mibefradil, bepridil, flunarizine, fluspirilene, gabapentin, pregabalin, and ziconotide or mixtures thereof.

7. A pharmaceutical combination according to any of the claims 1 to 6 whereby the calcium channel blocking agent is nifedipine and/or verapamil.

8. A pharmaceutical combination according to any of the claims 1 to 7, whereby the incretin mimetic and/or an incretin enhancer is selected from liraglutide, semaglutide, [D-Ala2]-GIP, exenatide, lixisenatide, dulaglutide, albiglutide or mixtures thereof.

9. A pharmaceutical combination according to any of the claims 1 to 8, whereby the incretin mimetic and/or an incretin enhancer is selected from sitagliptin, vildagliptin, saxagliptin, linagliptin, gemigliptin, anagliptin, teneligliptin, alogliptin, trelagliptin, omarigliptin, evogliptin, gosogliptin, dutogliptin or mixtures thereof.

10. A pharmaceutical combination according to any of the claims 1 to 9, whereby the incretin mimetic and/or an incretin enhancer is liraglutide and/or semaglutide.

11. A pharmaceutical composition comprising a pharmaceutical combination according to any of the claims 1 to 10.

12. The pharmaceutical composition according to claim 11 for use in the treatment and/or prevention of diabetes.

13. The pharmaceutical composition according to claim 11 or 12 for use in the treatment and/or prevention of Type-II diabetes.

14. The pharmaceutical composition according to any of the claims 11 to 13 for use in the treatment and/or prevention of Type-II diabetes in patients not suffering from prehypertension and/or hypertension.

15. Use of a pharmaceutical combination according to any of the claims 1 to 10 and/or the pharmaceutical composition according to any of the claims claim 11 to 14 for increasing or re-potentiating of incretin response of pancreatic β-cell in T2D patients.

Description

[0136] Additional details, characteristics and advantages of the object of the invention are disclosed in the subclaims and the following description of the respective figures—which in an exemplary fashion—show several inventive examples according to the invention. Such examples do not necessarily represent the full scope of the invention, however, and reference is made therefore to the claims and herein for interpreting the scope of the invention. It is to be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are intended to provide further explanation of the present invention as claimed.

[0137] FIG. 1 shows the glucose-stimulated insulin secretion (GSIS) for four comparative examples (i.e. one control sample, one sample with GLTX medium, one with liraglutide alone and one with nifedipine alone) and one inventive example (liraglutide and nifedipine in combination).

[0138] FIG. 2 shows the glucose-stimulated insulin secretion (GSIS) for four comparative examples (i.e. one control sample, one sample with GLTX medium, one with semaglutide alone and one with nifedipine alone) and one inventive example (semaglutide and nifedipine in combination).

[0139] FIG. 3 shows the glucose-stimulated insulin secretion (GSIS) for four comparative examples (i.e. one control sample, one sample with GLTX medium, one with liraglutide alone and one with verapamil) and one inventive example (liraglutide and verapamil in combination).

[0140] General method for measuring the glucose-stimulated insulin secretion (GSIS): 3D InSight™ Islet Microtissues were produced by optimized dissociation and hanging-drop based scaffold free reaggregation of isolated primary human islet cells. This process allows for a precise control over the newly forming islet microtissue while ensuring homogeneous and native-like distribution of endocrine cells within each microtissue (Misun and Yesildag, 2020). The resulting uniform islets display long-term (>28 days) and robust pancreatic β-cell function enabling high-throughput and longitudinal study of pancreatic islet function, regeneration, and preservation (Misun and Yesildag, 2020). The expanded life span of 3D InSight™ Human Islet Microtissues additionally enables physiologically relevant disease modeling. The glucolipotoxicity (GLTX) induced islet injury assay makes use of the detrimental effects of prolonged exposure to high glucose and high free fatty acids on pancreatic islets to mimic T2D relevant β-cell dysfunction. The toxicity of this interventions can be assessed by measuring robust indicators of islet function (glucose stimulated insulin secretion, GSIS). 3D InSight™ Human Islet Microtissues were produced from isolated cadaveric islets that was donated by the next of kin with informed consent (MT-04-002-01, InSphero AG, Switzerland, lot hIsMT_151, UNOS ID #AHHY337, 42 years old, female, Caucasian, BMI 23.5, EBV and CMV IgG positive serology, HbA1c 5.4%, death by stroke, no history of type 2 diabetes, no hypertension). The islet microtissues were cultured for 13 days (in a humidified incubator at 37° C. in 5% CO2, 95% air) in either standard 3D InSight™ Human Islet Maintenance Medium (CS-07-005-01, InSphero AG, Switzerland) or 3D InSight™ Human Islet Glucolipotox Medium (CS-07-510-04 InSphero AG, Switzerland).

[0141] Human islet microtissues cultured in standard islet maintenance medium, which contains 5.5 mM glucose, were used as healthy controls. Human islet microtissues cultured in glucolipotox medium, which contains 11 mM glucose and an additional 300 μM of free fatty acids (200 μM of Oleic acid and 100 μM palmitic acid) were used as stressed controls. FFAs were bound to bovine serum albumin in 1:5 ratio, therefore matching BSA concentration was used for the healthy control groups. The compound treatments were performed in glucolipotox medium for nifedipine (HY-B0284-1ML, Lucerna-Chem AG, Switzerland, at 0.2 μM), verapamil (HY-A0064-1ML, Lucerna-Chem AG, Switzerland, at 10 μM), semaglutide (NNC0113-0217, Novo Nordisk A/S, Denmark, at 1 μM) and liraglutide (NNC0090-1170, Novo Nordisk A/S, Denmark, at 0.1 or 1 μM), separately or combined, as illustrated in the description for FIGS. 1 to 3 (see below).

[0142] Media and compounds were renewed every 2 or 3 days, with 70 μl of culture medium per well. DMSO was used as solvent for nifedipine and verapamil and its concentration was equalized in all the remaining wells. 6 technical replicates per group were used for all samples and each technical replicate represents one uniform islet microtissue cultured in an individual well of a Akura™ 96 well microplate (InSphero AG, Switzerland). As the starting material is homogenous for all data points, data normalization is not required.

[0143] After 13 days of incubation, conditioned culture media were removed and the glucose stimulated insulin secretion was performed in the absence of the compounds. Islet microtissues were washed twice with 70 μl of 3D InSight™ Krebs Ringer HEPES Buffer (KRHB, CS-07-051-01, InSphero AG, Switzerland) prepared following manufacturer's instructions and supplemented with 2.8 mM glucose. Next, 3D InSight™ Human Islet Microtissues were equilibrated in 70 μl of this solution for 1 hour. Following the equilibration, 3D InSight™ Human Islet Microtissues were washed twice with 70 μl of KRHB supplemented with 2.8 mM glucose and incubated with 55 μl of KRHB supplemented with 2.8 mM glucose for an additional 2 hours of static incubation. KRHB was removed and islet microtissues were washed twice with 70 μl of KRHB supplemented with 2.8 mM glucose. Next, 55 μl of KRHB supplemented with 16.7 mM glucose was added to each well to perform a glucose-stimulated insulin secretion (GSIS) assay. After 2 hours of static incubation, conditioned KRHB was collected for the analysis of stimulated insulin secretion. All the incubations were performed in a humidified incubator at 37° C. in 5% CO2, 95% air. Insulin was quantified using Insulin Ultra-Sensitive HTRF Assay (62IN2PEH, Cisbio, France).

[0144] Outliers were detected with ROUT outlier test (Q=5%). Statistical significance against the glucolipotox group (GLTX) was determined with One-way ANOVA and Dunnett's multiple comparisons test, rejecting the null hypothesis at p<0.05. “*” “**” and “***” represents p values smaller or equal to 0.05, 0.01 and 0.001 respectively. Statistical significance between the mono and combination therapies were determined with Student's t-test, rejecting the null hypothesis at p<0.05. “#”, “##” and “## #” represents p values smaller or equal to 0.05, 0.01 and 0.001 respectively. “NS” represents not significant.

[0145] FIG. 1 shows the glucose-stimulated insulin secretion (GSIS) for four comparative examples (i.e. one control sample, one sample with GLTX medium, one with liraglutide alone and one with nifedipine alone) and one inventive example (liraglutide and nifedipine in combination).

[0146] As can be seen from FIG. 1, glucolipotox treatment results in decreased GSIS and long-term treatment with 0.1 μM liraglutide alone does not improve the GSIS. On the other hand, long-term treatment with 0.2 μM nifedipine alone has a positive influence on GSIS. However, GSIS is significantly higher for the inventive examples (combination of 0.1 μM liraglutide and 0.2 μM nifedipine) compared to the comparative examples containing either liraglutide or nifedipine alone. Most significantly, despite lacking long-term positive effects on insulin secretion, liraglutide in combination with nifedipine synergistically improves GSIS.

[0147] FIG. 2 shows the glucose-stimulated insulin secretion (GSIS) for four comparative examples (i.e. one control sample, one sample with GLTX medium, one with semaglutide alone and one with nifedipine alone) and one inventive example (semaglutide and nifedipine in combination).

[0148] As can be seen from FIG. 2, glucolipotox treatment results in decreased GSIS and long-term treatment with 1 μM semaglutide alone does not improve the GSIS. On the other hand, long-term treatment with 0.2 μM nifedipine alone has a positive influence on GSIS. However, GSIS is significantly higher for the inventive example (combination of 1 μM semaglutide and 0.2 μM nifedipine) compared to the comparative examples containing either semaglutide or nifedipine alone. Most significantly, despite lacking long-term positive effects on insulin secretion, semaglutide in combination with nifedipine synergistically improves GSIS.

[0149] FIG. 3 shows the glucose-stimulated insulin secretion (GSIS) for four comparative examples (i.e. one control sample, one sample with GLTX medium, one with liraglutide alone and one with verapamil) and one inventive example (liraglutide and verapamil in combination).

[0150] As can be seen from FIG. 3, glucolipotox treatment results in decreased GSIS (significant only with t-test) and long-term treatment with 1 μM liraglutide alone does not improve the GSIS. On the other hand, long-term treatment with 10 μM verapamil alone has a positive influence on GSIS. However, GSIS is significantly higher for the inventive examples (combination of 1 μM liraglutide and 10 μM verapamil) compared to the comparative examples containing either liraglutide or verapamil alone. Most significantly, despite lacking long-term positive effects on insulin secretion, liraglutide in combination with verapamil synergistically improves GSIS.

[0151] The particular combinations of elements and features in the above detailed embodiments are exemplary only; the interchanging and substitution of these teachings with other teachings in this and the patents/applications incorporated by reference are also expressly contemplated. As those skilled in the art will recognize, variations, modifications, and other implementations of what is described herein can occur to those of ordinary skill in the art without departing from the spirit and the scope of the invention as claimed. Accordingly, the foregoing description is by way of example only and is not intended as limiting. In the claims, the word “comprising” does not exclude other elements or steps, and the indefinite article “a” or “an” does not exclude a plurality. The mere fact that certain measures are recited in mutually different dependent claims does not indicate that a combination of these measured cannot be used to advantage. The invention's scope is defined in the following claims and the equivalents thereto. Furthermore, reference signs used in the description and claims do not limit the scope of the invention as claimed.

LITERATURE

[0152] Aljuffali, I. A., Lin, C. F. Chen, C. H. Fang, J. Y. (2016) The codrug approach for facilitating drug delivery and bioactivity, Expert Opinion on Drug Delivery, 2016, 1311-1325 [0153] Aroda, V. R., Knowler, W. C., Crandall, J. P., Perreault, L., Edelstein, S. L., Jeffries, S. L., Molitch, M. E., Pi-Sunyer, X., Darwin, C., Heckman-Stoddard, B. M., et al. (2017). Metformin for diabetes prevention: insights gained from the diabetes prevention program/diabetes prevention program outcomes study. Diabetologia 60, 1601-1611. [0154] Chon, S., & Gautier, J. F. (2016). An Update on the Effect of Incretin-Based Therapies on β-Cell Function and Mass. Diabetes & metabolism journal, 40(2), 99-114. [0155] Das, N. Dhanawat M., Dash, B. Nagarwal, R. C, Shrivastava, S. K. (2010) Codrug: An efficient approach for drug optimization, European Journal of Pharmaceutical Sciences Volume 41, Issue 5, 23 Dec. 2010, Pages 571-588 [0156] Dogruel, H., & Balci, M. K. (2019). Development of therapeutic options on type 2 diabetes in years: Glucagon-like peptide-1 receptor agonist's role in treatment; from the past to future. World journal of diabetes, 10(8), 446-453. [0157] Drucker D. J. (2006). The biology of incretin hormones. Cell metabolism, 3(3), 153-165. [0158] Foretz, M., Guigas, B., Bertrand, L., Pollak, M., and Viollet, B. (2014). Metformin: from mechanisms of action to therapies. Cell Metabol. 20, 953-966. [0159] Frias, J. P., et al. (2018) Efficacy and safety of LY3298176, a novel dual GIP and GLP-1 receptor agonist, in patients with type 2 diabetes: a randomized, placebo-controlled and active comparator-controlled phase 2 trial. The Lancet 392(10160), 2180-2193. [0160] Kjems, L. L., Holst, J. J., Vølund, A., & Madsbad, S. (2003). The influence of GLP-1 on glucose-stimulated insulin secretion: effects on beta-cell sensitivity in type 2 and nondiabetic subjects. Diabetes, 52(2), 380-386. https://doi.org/10.2337/diabetes.52.2.380 [0161] Ligthart, S., van Herpt, T. T., Leening, M. J., Kavousi, M., Hofman, A., Stricker, B. H., van Hoek, M., Sijbrands, E. J., Franco, O. H., and Dehghan, A. (2016). Lifetime risk of developing impaired glucose metabolism and eventual pro-gression from prediabetes to type 2 diabetes: a prospective cohort study. Lancet Diabetes Endocrinol. 4, 44-51. [0162] Marselli, L., Suleiman, M., Masini, M., Campani, D., Bugliani, M., Syed, F., Martino, L., Focosi, D., Scatena, F., Olimpico, F., et al. (2014). Are we overestimating the loss of beta cells in type 2 diabetes? Diabetologia 57, 362-365. [0163] Meier, J. J., and Bonadonna, R. C. (2013). Role of reduced beta-cell mass versus impaired beta-cell function in the pathogenesis of type 2 diabetes. Diabetes Care 36 (Suppl 2), S113-S119. [0164] Meier, J. J., & Nauck, M. A. (2010). Is the diminished incretin effect in type 2 diabetes just an epi-phenomenon of impaired beta-cell function?. Diabetes, 59(5), 1117-1125. [0165] Misun, P. M., Yesildag, B., Forschler, F., Neelakandhan, A., Rousset, N., Biernath, A., Hierlemann, A., & Frey, O. (2020). In Vitro Platform for Studying Human Insulin Release Dynamics of Single Pancreatic Islet Microtissues at High Resolution. Advanced biosystems, 4(3), e1900291 [0166] Nauck, M. A. & Meierk, J. J. (2018) Incretin hormones: Their role in health and disease, Diabetes Obes Metab. 2018 February; 20 Suppl 1:5-21 [0167] Palanisamy, S., Yien, E. L. H., Shi, L. W., Si, L. Y., Qi, S. H., Ling, L. S. C., Lun, T. W., and Chen, Y. N. (2018). Systematic review of efficacy and safety of newer antidiabetic drugs approved from 2013 to 2017 in controlling HbA1c in diabetes patients. Pharmacy (Basel) 6 [0168] Wajchenberg, B. L. (2007). beta-Cell failure in diabetes and preservation by clinical treatment. Endocr. Rev. 28, 187-218. [0169] Weir, G. C., and Bonner-Weir, S. (2004). Five stages of evolving beta-cell dysfunction during progression to diabetes. Diabetes 53 (Suppl 3), S16-S21. [0170] Yabe, D., & Seino, Y. (2011). Two incretin hormones GLP-1 and GIP: comparison of their actions in insulin secretion and R cell preservation. Progress in biophysics and molecular biology, 107(2), 248-256. [0171] Zhang, E., Al-Amily, I., Mohammed, S. Luan, C. Asplung, O. Ahmed M, Ye, Y. Ben-hail, D. Soni, A., Vishnu, N, Bompada, P., De Marinis, Y., Groopl, L., Shoshan-Barmatz, V, Renstro, E, Wollheim, C. and Salehi, A., (2019) Preserving Insulin Secretion in Diabetes by Inhibiting VDAC1 Overexpression and Surface Translocation in β-Cells, Cell Metabolism 29, 64-77