New analogues from LASSBio‐1524, an IKK‐β inhibitor in Inflammatory Bowel Diseases
Authors: de Morais Cordeiro Natália , Freitas Rosana Helena Coimbra Nogueira , Manssour Carlos Alberto , Fernandes Patricia Dias 
Published: November 2023

LASSBio‐1524 is an ikB kinase β inhibitor that demonstrated acute anti‐inflammatory efficacy in vivo. As inflammatory bowel diseases (IBD) presents an increase in NF‐κB expression and high levels of cytokines our aim was to evaluate the effects of new analogues from LASSBio‐1524 IBD's models (i.e., Crohn's disease and ulcerative colitis).

Chemically induced colitis models with Dextran Sodium Sulfate (DSS) and Dinitrobenzene Sulfonic Acid (DNBS) were performed in Swiss webster mice (22–25 g, n=6–8). In order to investigate this effect, mice were treated once a day with 30 μmol/kg, (p.o.) of LASSBio‐1524 and its three analogues, LASSBio‐1760, LASSBio‐1763 or LASSBio‐1764. In DSS model, mice received drinking water supplemented with 3% DSS in 2 cycles of 5 days alternated with 10 days of regular water and were treated from day 1 to 7. At day 30, colons were collected and cytokines (TNF‐α, IL‐1β, IL‐12, IL‐6, IL‐10) and chemokine MCP‐1 were quantified by ELISA. LASSBio‐1524, LASSBio‐1760, LASSBio‐1763 and LASSBio‐1764 reduced TNF‐α when compared to DSS vehicle group in 91%, 91%, 97% and 92%, respectively. IL‐1β levels were also reduced in 86%, 91%, 68% and 91%. IL‐12 was reduced in 47%, 72% 71% and 76%. IL‐6 levels were reduced in 81%, 83%, 79% and 83%. IL‐10 increased its production in 893%, 1,500%, 340%, and 646%. Levels of MCP‐1 was not significantly affected. DNBS‐induced model consists in an intrarectal instillation of 1 mg DNBS/animal in 35% ethanol (100 μL), at day 1, treatment once a day and euthanasia in the 4th day. This protocol was also performed with 8 days and the inhibition profile remains. After 4 days, the cytokines listed above were also measured from the colonic tissue. LASSBio‐1524 and its three analogues reduced in 67%, 62%, 47% and 51% TNF‐α production. IL‐1β was also reduced, in 55%, 57%, 67% and 73%. IL‐12 levels were reduced in 84%, 72%, 57% and 54%. Treatment also reduced IL‐6 levels: 88%, 73%, 91% and 88%. IL‐10 levels were also increased in this model, and treatment with LASSBio‐1524 increased in 73%, LASSBio‐1760 in 67%, LASSBio‐1763 in 169% and LASSbio‐1764 in 7%. MCP‐1 reduced in 81%, 72%, 97% and 86%. Statistical analysis was performed by ANOVA followed by Bonferroni test. Procedures involving animals follow animal care guidelines and protocol for use (DFBCICB015‐04/16) was approved by CEUA/UFRJ.

Knowing that cytokines or cytokine signaling events are being seen as therapeutic targets in IBD, these compounds can be interesting once NF‐κB activation is associated with disease activity in IBD and they modulate some NF‐κB targets. Treatment with the compounds reduced proinflammatory cytokines TNF‐α, IL‐1 β and IL‐6, that are increased in Crohn's disease and ulcerative colitis as well as IL‐12, produced by APCs during intestinal inflammation. In addition, treatment increased IL‐10 levels, reported as an immunomodulatory cytokine in IBD. The new analogues showed anti‐inflammatory potential even better in some cytokines production than the original molecule, placing them as candidates for the development of new anti‐inflammatory prototypes.

Support or Funding Information

Financial support: CNPq, CAPES, FAPERJ and Instituto Vital Brazil (donation of animals).

Technical support: Alan Minho.

DSS‐ induced colitis cytokines

DSS‐ induced colitis
30 μmol/kg once a day ‐ 7 days

Cytokine (pg/μg of protein)
Saline‐Vehicle
DSS‐Vehicle
LASSBio‐1524
LASSBio‐1760
LASSBio‐1763
LASSBio‐1764

TNF‐α
14.3 ± 5.2
70.01 ± 21.3
6.3* ± 5.1
5.8* ± 3.6
1.9* ± 2.3
5.1* ± 3.3

IL‐1β
1.80 ± 09
51.5 ± 20.1
7.0* ± 9.8
4.6* ± 3.1
16.2* ± 11.7
4.4* ± 3.8

IL‐12
10.3 ± 5.3
69.6 ± 23.7
36.3 ± 11.3
19.0* ± 13.2
20.1* ± 8.3
16.5* ± 5.3

IL‐6
0.2 ± 0.18
4.3 ± 1.8
0.8* ± 0.5
0.7* ± 0.5
0.9* ± 0.7
0.7* ± 0.5

IL‐10
33.7 ± 10.4
1.5 ± 1.1
14.9* ± 4.4
25.4* ± 12.8
6.6* ± 5.4
11.2* ± 5.9

MCP‐1
2.3 ± 0.4
1.8 ± 1.3
1.9 ± 1.1
1.9 ± 1.0
2.1 ± 1.8
3.4 ± 2.0

ANOVA followed by Bonferroni test.

p<0.05 when compared to vehicle stimulated group.

DNBS‐induced colitis cytokines

DNBS‐ induced colitis
30 μmol/kg once a day ‐ 4 days

Cytokine (pg/μg of protein)
Saline‐Vehicle
DSS‐Vehicle
LASSBio‐1524
LASSBio‐1760
LASSBio‐1763
LASSBio‐1764

TNF‐α
10.7 ± 7.1
36.6 ± 17.5
12* ± 2.8
13.8* ± 6.5
19.3* ± 4.0
17.8* ± 10.5

IL‐1β
12.0 ± 6.1
34.1 ± 11.5
15.1* ± 8.6
14.5* ± 7.2
11.1* ± 2.5
8.9* ± 1.2

IL‐12
13.5 ± 1.4
26.1 ± 10.3
4.1* ± 2.2
7.2* ± 3.1
11.1* ± 1.5
11.8* ± 4.7

IL‐6
0.8 ± 0.2
4.5 ± 1.8
0.5* ± 0.1
1.2* ± 0.7
0.4* ± 0.2
0.5* ± 0.2

IL‐10
337 ± 110
34.1 ± 11.5
15.1* ± 8.6
14.5* ± 7.2
11.1* ± 2.5
8.9* ± 1.2

MCP‐1
1.3 ± 0.4
3.9 ± 1.5
0.7* ± 0.2
1.0* ± 0.6
0.1* ± 0.06
0.5* ± 0.2

ANOVA followed by Bonferroni test.

p<0.05 when compared to vehicle stimulated group.